trpc3  (Alomone Labs)


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    Structured Review

    Alomone Labs trpc3
    <t>TRPC3</t> antibody reduced mGluR1-mediated slow EPSC and blocked cerebellar LTD. A , Application of TRPC3 antibody reduced sEPSC. Representative current traces showing parallel fiber burst-evoked slow EPSC with TRPC3 antibody ( n = 6) and preincubated TRPC3 antibody ( n = 10). Time course graph showing sEPSC amplitude reduced over time, which indicates the diffusion time course from the patch pipette to the dendrite spine. Summary bar graph displays a significant decrease of sEPSC in the presence of TRPC3 antibody (*** p
    Trpc3, supplied by Alomone Labs, used in various techniques. Bioz Stars score: 86/100, based on 40 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/trpc3/product/Alomone Labs
    Average 86 stars, based on 40 article reviews
    Price from $9.99 to $1999.99
    trpc3 - by Bioz Stars, 2022-09
    86/100 stars

    Images

    1) Product Images from "Transient Receptor Potential Canonical Channels Regulate the Induction of Cerebellar Long-Term Depression"

    Article Title: Transient Receptor Potential Canonical Channels Regulate the Induction of Cerebellar Long-Term Depression

    Journal: The Journal of Neuroscience

    doi: 10.1523/JNEUROSCI.0073-12.2012

    TRPC3 antibody reduced mGluR1-mediated slow EPSC and blocked cerebellar LTD. A , Application of TRPC3 antibody reduced sEPSC. Representative current traces showing parallel fiber burst-evoked slow EPSC with TRPC3 antibody ( n = 6) and preincubated TRPC3 antibody ( n = 10). Time course graph showing sEPSC amplitude reduced over time, which indicates the diffusion time course from the patch pipette to the dendrite spine. Summary bar graph displays a significant decrease of sEPSC in the presence of TRPC3 antibody (*** p
    Figure Legend Snippet: TRPC3 antibody reduced mGluR1-mediated slow EPSC and blocked cerebellar LTD. A , Application of TRPC3 antibody reduced sEPSC. Representative current traces showing parallel fiber burst-evoked slow EPSC with TRPC3 antibody ( n = 6) and preincubated TRPC3 antibody ( n = 10). Time course graph showing sEPSC amplitude reduced over time, which indicates the diffusion time course from the patch pipette to the dendrite spine. Summary bar graph displays a significant decrease of sEPSC in the presence of TRPC3 antibody (*** p

    Techniques Used: Diffusion-based Assay, Transferring

    2) Product Images from "The involvement of TRPC3 channels in sinoatrial arrhythmias"

    Article Title: The involvement of TRPC3 channels in sinoatrial arrhythmias

    Journal: Frontiers in Physiology

    doi: 10.3389/fphys.2015.00086

    The effect of angiotension II (Ang II) and a selective TRPC3 channel blocker Pyr10 on pacemaker action potential after application of Ang II. (A–D) Intracellular recordings from WT mice; (E–H ) Intracellular recordings from TRPC3 −/− mice with conditions as indicated in each panel.
    Figure Legend Snippet: The effect of angiotension II (Ang II) and a selective TRPC3 channel blocker Pyr10 on pacemaker action potential after application of Ang II. (A–D) Intracellular recordings from WT mice; (E–H ) Intracellular recordings from TRPC3 −/− mice with conditions as indicated in each panel.

    Techniques Used: Mouse Assay

    Atrial arrhythmias induced by Ang II and pacing . Reconstructed excitation maps of data from multi-electrode array recordings (images below) from a Langendorff-perfused isolated heart. (A) sinus rhythm and (B) pacing induced atrial tachycardia. Note in AT, the ectopic beat starts from a different site to sinus rhythms. (C) Representative ECG recordings from isolated heart from WT and TRPC3 −/− mice under control conditions. (D) Atrial tachycardia and atrial fibrillation induced by burst pacing in WT mice and atrial-ventricular conduction block recorded from a TRPC −/− mouse heart respectively. (E) On average, the arrhythmia index was significant reduced in TRPC3KO mice ( n = 8) compared to WT mice ( n = 11, P = 0.004).
    Figure Legend Snippet: Atrial arrhythmias induced by Ang II and pacing . Reconstructed excitation maps of data from multi-electrode array recordings (images below) from a Langendorff-perfused isolated heart. (A) sinus rhythm and (B) pacing induced atrial tachycardia. Note in AT, the ectopic beat starts from a different site to sinus rhythms. (C) Representative ECG recordings from isolated heart from WT and TRPC3 −/− mice under control conditions. (D) Atrial tachycardia and atrial fibrillation induced by burst pacing in WT mice and atrial-ventricular conduction block recorded from a TRPC −/− mouse heart respectively. (E) On average, the arrhythmia index was significant reduced in TRPC3KO mice ( n = 8) compared to WT mice ( n = 11, P = 0.004).

    Techniques Used: Isolation, Mouse Assay, Blocking Assay

    Isolated single SAN cells express TRPC3, STIM1, and IP 3 R2 . Confocal immunofluorenscence images of isolated single pacemaker cells. Isolated group (A) and single (B) pacemaker cells were positively stained with anti HCN4 (in green). (C) Anti-TRPC3 in red, anti-STIM1 in green. (D) Anti-IP3R2 in red, anti-STIM1 in green. (E) 3D reconstruction image using N-SIM microscopy. Anti-IP 3 R2 in red, anti-STIM1 in green. Areas of co-localisation appear yellow due to color mixing.
    Figure Legend Snippet: Isolated single SAN cells express TRPC3, STIM1, and IP 3 R2 . Confocal immunofluorenscence images of isolated single pacemaker cells. Isolated group (A) and single (B) pacemaker cells were positively stained with anti HCN4 (in green). (C) Anti-TRPC3 in red, anti-STIM1 in green. (D) Anti-IP3R2 in red, anti-STIM1 in green. (E) 3D reconstruction image using N-SIM microscopy. Anti-IP 3 R2 in red, anti-STIM1 in green. Areas of co-localisation appear yellow due to color mixing.

    Techniques Used: Isolation, Staining, Microscopy

    The effect of Pyr10 on intracellular Ca 2+ after application of 1-oleoy1-2-acyl-sn-glycerol (OAG) 100 μM . The intact SANs were loaded with Ca 2+ indicator indo-1. (A,B) show intracellular Ca 2+ recordings from a WT mouse. (B,C) The statistics pool data shows percentage changes in resting Ca 2+ , Ca 2+ transient and firing rate in WT and TRPC3 KO mice respectively. (B) OAG treatment against control. (C) 2 μM Pyr10 treatment against OAG treatment. * P
    Figure Legend Snippet: The effect of Pyr10 on intracellular Ca 2+ after application of 1-oleoy1-2-acyl-sn-glycerol (OAG) 100 μM . The intact SANs were loaded with Ca 2+ indicator indo-1. (A,B) show intracellular Ca 2+ recordings from a WT mouse. (B,C) The statistics pool data shows percentage changes in resting Ca 2+ , Ca 2+ transient and firing rate in WT and TRPC3 KO mice respectively. (B) OAG treatment against control. (C) 2 μM Pyr10 treatment against OAG treatment. * P

    Techniques Used: Mouse Assay

    Signaling pathways involved in the activation of TRPC3 . The G-protein coupled receptor (GPCRs) activates phospholiase C (PLC), resulting in generation of IP3 and diacylglycerol (DAG). IP3 activates its receptor that leads to Ca 2+ release from SR/ER and the depletion of SR/ER Ca 2+ store. The Ca 2+ content change in the store can be sensed by STIM1, the ER Ca 2+ sensor and cause store-operated Ca 2+ entry (SOCE) through TRPC3 channels. An additional or alternate possibility is that DAG can directly activate TRPC3, and produce receptor-operated Ca 2+ entry (ROCE). GPCRs, G-protein coupled receptors; IP3R, inositol 1,4,5-trisphosphate receptors; PLC, Phospholipase C; ROCE, Receptor–operated Ca 2+ entry; SOCE, store-operated Ca 2+ entry; SR/ER, sarco- endo-plasmic reticulum; STIM1, stromal interacting molecule 1; TRPC3,4,6 canonical transient receptor potential channel types 3,4,6; NCX Na/Ca exchange.
    Figure Legend Snippet: Signaling pathways involved in the activation of TRPC3 . The G-protein coupled receptor (GPCRs) activates phospholiase C (PLC), resulting in generation of IP3 and diacylglycerol (DAG). IP3 activates its receptor that leads to Ca 2+ release from SR/ER and the depletion of SR/ER Ca 2+ store. The Ca 2+ content change in the store can be sensed by STIM1, the ER Ca 2+ sensor and cause store-operated Ca 2+ entry (SOCE) through TRPC3 channels. An additional or alternate possibility is that DAG can directly activate TRPC3, and produce receptor-operated Ca 2+ entry (ROCE). GPCRs, G-protein coupled receptors; IP3R, inositol 1,4,5-trisphosphate receptors; PLC, Phospholipase C; ROCE, Receptor–operated Ca 2+ entry; SOCE, store-operated Ca 2+ entry; SR/ER, sarco- endo-plasmic reticulum; STIM1, stromal interacting molecule 1; TRPC3,4,6 canonical transient receptor potential channel types 3,4,6; NCX Na/Ca exchange.

    Techniques Used: Activation Assay, Planar Chromatography

    3) Product Images from "TRPC3 positively regulates reactive oxygen species driving maladaptive cardiac remodeling"

    Article Title: TRPC3 positively regulates reactive oxygen species driving maladaptive cardiac remodeling

    Journal: Scientific Reports

    doi: 10.1038/srep37001

    TRPC3 forms a stable ternary complex with Nox2 and p22 phox . ( a , b ) Expression of Nox2 and p22 phox proteins in HEK293 cells that express a different combination of TRPC3-GFP and GFP. Results of a quantitative analysis are shown in ( b ) (n = 3). ( c ) Nox2 mRNA amounts in HEK293 cells co-expressing Nox2 with GFP or TRPC3-GFP (n = 3). ( d ) Increased Nox2 and p22 phox protein in HEK293 cells co-expressing pore-dead mutant of TRPC3 (n = 3). ( e ) Interaction of TRPC3 with Nox2 in HEK293 cells. Immunoprecipitation was performed using an anti-flag antibody. ( f ) Nox2 protein expression in HEK293 cells expressing TRPC3 alone or co-expressing TRPC3 and TRPC6 (n = 3). Error bars, s.e.m. *P
    Figure Legend Snippet: TRPC3 forms a stable ternary complex with Nox2 and p22 phox . ( a , b ) Expression of Nox2 and p22 phox proteins in HEK293 cells that express a different combination of TRPC3-GFP and GFP. Results of a quantitative analysis are shown in ( b ) (n = 3). ( c ) Nox2 mRNA amounts in HEK293 cells co-expressing Nox2 with GFP or TRPC3-GFP (n = 3). ( d ) Increased Nox2 and p22 phox protein in HEK293 cells co-expressing pore-dead mutant of TRPC3 (n = 3). ( e ) Interaction of TRPC3 with Nox2 in HEK293 cells. Immunoprecipitation was performed using an anti-flag antibody. ( f ) Nox2 protein expression in HEK293 cells expressing TRPC3 alone or co-expressing TRPC3 and TRPC6 (n = 3). Error bars, s.e.m. *P

    Techniques Used: Expressing, Mutagenesis, Immunoprecipitation

    TRPC3 plays a critical role in Mechanical stretch-induced ROS production. ( a , b ) Effects of siRNA targeting TRPC1, C3 or C6 on mechanical stretch (MS)-induced ROS production (n = 3). ( c ) mRNA expression of either TRPC1 or TRPC6 in NRCM transfected with siRNAs against either TRPC1 or TRPC6, respectively (n = 3). ( d,e ) Time courses of MS-induced ROS production in NRCMs treated with GsMTx-4 (1 μM; ( d ) or TRPV4 inhibitor (RN1734, 50 μM; ( e ) Reagents were added to cells 5 min before MS (n = 3). ( f ) MS-induced ROS production in TRPC(1–7)-deficient MEF cells expressing TRPC3, TRPC6, TRPC7, or LacZ (n = 30). Data are representative of three independent experiments. ( g–i ) Effect of TRPC3 siRNA on the protein abundances of TRPC3 ( h ) and Nox2 ( i ) protein expressions in NRCMs (n = 3). Error bars, s.e.m. *P
    Figure Legend Snippet: TRPC3 plays a critical role in Mechanical stretch-induced ROS production. ( a , b ) Effects of siRNA targeting TRPC1, C3 or C6 on mechanical stretch (MS)-induced ROS production (n = 3). ( c ) mRNA expression of either TRPC1 or TRPC6 in NRCM transfected with siRNAs against either TRPC1 or TRPC6, respectively (n = 3). ( d,e ) Time courses of MS-induced ROS production in NRCMs treated with GsMTx-4 (1 μM; ( d ) or TRPV4 inhibitor (RN1734, 50 μM; ( e ) Reagents were added to cells 5 min before MS (n = 3). ( f ) MS-induced ROS production in TRPC(1–7)-deficient MEF cells expressing TRPC3, TRPC6, TRPC7, or LacZ (n = 30). Data are representative of three independent experiments. ( g–i ) Effect of TRPC3 siRNA on the protein abundances of TRPC3 ( h ) and Nox2 ( i ) protein expressions in NRCMs (n = 3). Error bars, s.e.m. *P

    Techniques Used: Mass Spectrometry, Expressing, Transfection

    TRPC3 prevents Nox2 protein from proteasomal degradation. ( a–e ) Abundances of Nox2 protein ( a , b ) and mRNAs of TRPC3 ( c ), Nox2 ( d ), and p22 phox ( e ) in NRCM transfected with siRNAs targeting TRPC3 with or without MG132. Cells were treated with siRNAs and MG132 (1 μM) simultaneously (n = 3). ( f , g ) Effect of siRNA targeting TRPC3 on Nox2 protein abundance in cell surface (Surface) and total lysates (Total) from NRCMs (n = 3). GAPDH was used as an internal control. Error bars, s.e.m. *P
    Figure Legend Snippet: TRPC3 prevents Nox2 protein from proteasomal degradation. ( a–e ) Abundances of Nox2 protein ( a , b ) and mRNAs of TRPC3 ( c ), Nox2 ( d ), and p22 phox ( e ) in NRCM transfected with siRNAs targeting TRPC3 with or without MG132. Cells were treated with siRNAs and MG132 (1 μM) simultaneously (n = 3). ( f , g ) Effect of siRNA targeting TRPC3 on Nox2 protein abundance in cell surface (Surface) and total lysates (Total) from NRCMs (n = 3). GAPDH was used as an internal control. Error bars, s.e.m. *P

    Techniques Used: Transfection

    TRPC3 forms a stable ternary complex with Nox2 and p22 phox proteins in endogenously p22 phox -absent CHO cells. ( a ) Expression of Nox2 and p22 phox proteins in CHO cells that express a different combination of TRPC3-GFP and GFP. ( b ) Results of quantitative analysis (n = 3). ( c ) Expression of Nox2 and p22 phox co-expressed with either GFP or TRPC3-GFP in MG132 (10 μM)-treated CHO cells. ( d ) Graphs depict the relative expression of either Nox2 or p22 phox protein to that in non-treated cells. Band intensities were normalized by GAPDH. ( e–g ) Interaction of TRPC3 with p22 phox and Nox2 in CHO cells. ( h ) Localization of Nox2 in CHO cells co-expressing Nox2 with TRPC3-GFP (or GFP-F). Error bars, s.e.m. *P
    Figure Legend Snippet: TRPC3 forms a stable ternary complex with Nox2 and p22 phox proteins in endogenously p22 phox -absent CHO cells. ( a ) Expression of Nox2 and p22 phox proteins in CHO cells that express a different combination of TRPC3-GFP and GFP. ( b ) Results of quantitative analysis (n = 3). ( c ) Expression of Nox2 and p22 phox co-expressed with either GFP or TRPC3-GFP in MG132 (10 μM)-treated CHO cells. ( d ) Graphs depict the relative expression of either Nox2 or p22 phox protein to that in non-treated cells. Band intensities were normalized by GAPDH. ( e–g ) Interaction of TRPC3 with p22 phox and Nox2 in CHO cells. ( h ) Localization of Nox2 in CHO cells co-expressing Nox2 with TRPC3-GFP (or GFP-F). Error bars, s.e.m. *P

    Techniques Used: Expressing

    Formation of a TRPC3/Nox2 complex promotes TRPC3 channel activity through stabilization at the plasma membrane. ( a ) Effect of Nox2 siRNA on expression of TRPC3 in NRCMs (n = 3). ( b ) Representative images showing the levels of TRPC3-GFP and GFP expression in HEK293 cells co-expressing p22 phox or Nox2 (n = 3). ( c ) Expression of TRPC3-GFP mRNA in HEK293 cells co-expressing p22 phox or Nox2 (n = 3). ( d–f ) Representative time courses of TRPC3 currents ( d ) and the current-voltage (I-V) relationships ( e ) and peak TRPC3 current densities at −60 mV ( f ) induced by 100 μM carbachol (CCh) in HEK293 cells expressing TRPC3-mCherry alone or with p22 phox , Nox2, both p22 phox and Nox2, or Nox2 treated with DPI. DPI (0.3 μM) was treated 1 min before CCh stimulation. ( g , h ) Representative Ca 2+ responses in the presence ( g ) or absence ( h ) of pyrazole-3 (Pyr3, 1 μM) upon mechanical stretch (MS) application. ( i ) Peak Ca 2+ increases after MS in NRCMs treated with (n = 61) or without Pyr3 (n = 78). ( j ) Changes of minimal [Ca 2+ ] i before and after MS application. Minimal [Ca 2+ ] i from Ca 2+ responses in every 1 min were analyzed and represented as diastolic [Ca 2+ ] i . ( k ) Schematic images showing phosphorylation of p47 phox via TRPC3-PKCβ activation induced by MS in the heart. ( l–n ) Effects of TRPC3 ( l , m ) or PKCβ ( n) ; 10 μM Gö6976) inhibitors on p47 phox phosphorylation induced by MS in NRCMs (n = 3). ( o ) MS-induced ROS generation in NRCMs treated with a PKCβ inhibitor (n = 3). ( p ) Co-immunoprecipitation of TRPC3 with PKCβ, Nox2 and p22 phox in mouse hearts 1week after TAC operation (n = 3). Error bars, s.e.m.*P
    Figure Legend Snippet: Formation of a TRPC3/Nox2 complex promotes TRPC3 channel activity through stabilization at the plasma membrane. ( a ) Effect of Nox2 siRNA on expression of TRPC3 in NRCMs (n = 3). ( b ) Representative images showing the levels of TRPC3-GFP and GFP expression in HEK293 cells co-expressing p22 phox or Nox2 (n = 3). ( c ) Expression of TRPC3-GFP mRNA in HEK293 cells co-expressing p22 phox or Nox2 (n = 3). ( d–f ) Representative time courses of TRPC3 currents ( d ) and the current-voltage (I-V) relationships ( e ) and peak TRPC3 current densities at −60 mV ( f ) induced by 100 μM carbachol (CCh) in HEK293 cells expressing TRPC3-mCherry alone or with p22 phox , Nox2, both p22 phox and Nox2, or Nox2 treated with DPI. DPI (0.3 μM) was treated 1 min before CCh stimulation. ( g , h ) Representative Ca 2+ responses in the presence ( g ) or absence ( h ) of pyrazole-3 (Pyr3, 1 μM) upon mechanical stretch (MS) application. ( i ) Peak Ca 2+ increases after MS in NRCMs treated with (n = 61) or without Pyr3 (n = 78). ( j ) Changes of minimal [Ca 2+ ] i before and after MS application. Minimal [Ca 2+ ] i from Ca 2+ responses in every 1 min were analyzed and represented as diastolic [Ca 2+ ] i . ( k ) Schematic images showing phosphorylation of p47 phox via TRPC3-PKCβ activation induced by MS in the heart. ( l–n ) Effects of TRPC3 ( l , m ) or PKCβ ( n) ; 10 μM Gö6976) inhibitors on p47 phox phosphorylation induced by MS in NRCMs (n = 3). ( o ) MS-induced ROS generation in NRCMs treated with a PKCβ inhibitor (n = 3). ( p ) Co-immunoprecipitation of TRPC3 with PKCβ, Nox2 and p22 phox in mouse hearts 1week after TAC operation (n = 3). Error bars, s.e.m.*P

    Techniques Used: Activity Assay, Expressing, Mass Spectrometry, Activation Assay, Immunoprecipitation

    TRPC3 deletion suppresses TAC-induced LV dysfunction and dilation through Nox2 inhibition. ( a ) Left ventricular end-diastolic pressure (LVEDP; left) and dP/dT max (right) in TAC-operated TRPC3 (+/+) (n = 13) and TRPC3 (−/−) (n = 12) mice 6 week post-operation. ( b ) Myocardial malondialdehyde concentrations 1 week after TAC (n = 4). ( c ) Abundance of Nox2 protein in TRPC3 (+/+) and TRPC3 (−/−) hearts 1 week after TAC (n = 3). ( d ) Representative immunofluorescence images of TRPC3, p22 phox , and caveolin-3 (Cav-3) in adult mouse cardiomyocytes isolated from muscle LIM protein-deficient hearts. ( e ) Representative immunofluorescence images of p22 phox in adult mouse cardiomyocytes: green, anti-p22 phox ; blue, DAPI. ( f ) Relative abundances of p22 phox and Nox2 mRNA in mouse hearts 1 week after TAC (n = 4). ( g ) Abundance of Nox2 protein in TRPC6 (+/+) and TRPC6 (−/−) hearts 1 week after TAC (n = 3). Error bars, s.e.m. *P
    Figure Legend Snippet: TRPC3 deletion suppresses TAC-induced LV dysfunction and dilation through Nox2 inhibition. ( a ) Left ventricular end-diastolic pressure (LVEDP; left) and dP/dT max (right) in TAC-operated TRPC3 (+/+) (n = 13) and TRPC3 (−/−) (n = 12) mice 6 week post-operation. ( b ) Myocardial malondialdehyde concentrations 1 week after TAC (n = 4). ( c ) Abundance of Nox2 protein in TRPC3 (+/+) and TRPC3 (−/−) hearts 1 week after TAC (n = 3). ( d ) Representative immunofluorescence images of TRPC3, p22 phox , and caveolin-3 (Cav-3) in adult mouse cardiomyocytes isolated from muscle LIM protein-deficient hearts. ( e ) Representative immunofluorescence images of p22 phox in adult mouse cardiomyocytes: green, anti-p22 phox ; blue, DAPI. ( f ) Relative abundances of p22 phox and Nox2 mRNA in mouse hearts 1 week after TAC (n = 4). ( g ) Abundance of Nox2 protein in TRPC6 (+/+) and TRPC6 (−/−) hearts 1 week after TAC (n = 3). Error bars, s.e.m. *P

    Techniques Used: Inhibition, Mouse Assay, Immunofluorescence, Isolation

    Physical interaction between TRPC3 and Nox2 is critical for stabilization of Nox2. ( a ) Schematic illustration of TRPC3 terminal deletion mutants. ( b , c ) Expression of Nox2 and p22 phox co-expressed with TRPC3 deletion mutants in HEK293 cells (n = 3). ( d ) OAG-induced ROS production in NRCMs expressing Nox2-interacting TRPC3 C-terminal fragment (C3-C fragment) (n = 20–28). ( e ) Co-immunoprecipitation of TRPC3 with Nox2 in the presence or absence of C3-C fragment. Representative blot from three independent experiments was shown. ( f ) ATP (100 μM)-induced Ca 2+ responses in HEK293 cells expressing TRPC3 with or without C3-C fragment (n = 35–51). Timing of solution exchanges were indicated by horizontal bars above the graph. ( g ) Model of the regulation of TRPC3-Nox2 stability and induction of LV dysfunction induced by diastolic stretch of cardiomyocytes. Error bars, s.e.m.*P
    Figure Legend Snippet: Physical interaction between TRPC3 and Nox2 is critical for stabilization of Nox2. ( a ) Schematic illustration of TRPC3 terminal deletion mutants. ( b , c ) Expression of Nox2 and p22 phox co-expressed with TRPC3 deletion mutants in HEK293 cells (n = 3). ( d ) OAG-induced ROS production in NRCMs expressing Nox2-interacting TRPC3 C-terminal fragment (C3-C fragment) (n = 20–28). ( e ) Co-immunoprecipitation of TRPC3 with Nox2 in the presence or absence of C3-C fragment. Representative blot from three independent experiments was shown. ( f ) ATP (100 μM)-induced Ca 2+ responses in HEK293 cells expressing TRPC3 with or without C3-C fragment (n = 35–51). Timing of solution exchanges were indicated by horizontal bars above the graph. ( g ) Model of the regulation of TRPC3-Nox2 stability and induction of LV dysfunction induced by diastolic stretch of cardiomyocytes. Error bars, s.e.m.*P

    Techniques Used: Expressing, Immunoprecipitation

    4) Product Images from "The involvement of TRPC3 channels in sinoatrial arrhythmias"

    Article Title: The involvement of TRPC3 channels in sinoatrial arrhythmias

    Journal: Frontiers in Physiology

    doi: 10.3389/fphys.2015.00086

    The effect of angiotension II (Ang II) and a selective TRPC3 channel blocker Pyr10 on pacemaker action potential after application of Ang II. (A–D) Intracellular recordings from WT mice; (E–H ) Intracellular recordings from TRPC3 −/− mice with conditions as indicated in each panel.
    Figure Legend Snippet: The effect of angiotension II (Ang II) and a selective TRPC3 channel blocker Pyr10 on pacemaker action potential after application of Ang II. (A–D) Intracellular recordings from WT mice; (E–H ) Intracellular recordings from TRPC3 −/− mice with conditions as indicated in each panel.

    Techniques Used: Mouse Assay

    Atrial arrhythmias induced by Ang II and pacing . Reconstructed excitation maps of data from multi-electrode array recordings (images below) from a Langendorff-perfused isolated heart. (A) sinus rhythm and (B) pacing induced atrial tachycardia. Note in AT, the ectopic beat starts from a different site to sinus rhythms. (C) Representative ECG recordings from isolated heart from WT and TRPC3 −/− mice under control conditions. (D) Atrial tachycardia and atrial fibrillation induced by burst pacing in WT mice and atrial-ventricular conduction block recorded from a TRPC −/− mouse heart respectively. (E) On average, the arrhythmia index was significant reduced in TRPC3KO mice ( n = 8) compared to WT mice ( n = 11, P = 0.004).
    Figure Legend Snippet: Atrial arrhythmias induced by Ang II and pacing . Reconstructed excitation maps of data from multi-electrode array recordings (images below) from a Langendorff-perfused isolated heart. (A) sinus rhythm and (B) pacing induced atrial tachycardia. Note in AT, the ectopic beat starts from a different site to sinus rhythms. (C) Representative ECG recordings from isolated heart from WT and TRPC3 −/− mice under control conditions. (D) Atrial tachycardia and atrial fibrillation induced by burst pacing in WT mice and atrial-ventricular conduction block recorded from a TRPC −/− mouse heart respectively. (E) On average, the arrhythmia index was significant reduced in TRPC3KO mice ( n = 8) compared to WT mice ( n = 11, P = 0.004).

    Techniques Used: Isolation, Mouse Assay, Blocking Assay

    Isolated single SAN cells express TRPC3, STIM1, and IP 3 R2 . Confocal immunofluorenscence images of isolated single pacemaker cells. Isolated group (A) and single (B) pacemaker cells were positively stained with anti HCN4 (in green). (C) Anti-TRPC3 in red, anti-STIM1 in green. (D) Anti-IP3R2 in red, anti-STIM1 in green. (E) 3D reconstruction image using N-SIM microscopy. Anti-IP 3 R2 in red, anti-STIM1 in green. Areas of co-localisation appear yellow due to color mixing.
    Figure Legend Snippet: Isolated single SAN cells express TRPC3, STIM1, and IP 3 R2 . Confocal immunofluorenscence images of isolated single pacemaker cells. Isolated group (A) and single (B) pacemaker cells were positively stained with anti HCN4 (in green). (C) Anti-TRPC3 in red, anti-STIM1 in green. (D) Anti-IP3R2 in red, anti-STIM1 in green. (E) 3D reconstruction image using N-SIM microscopy. Anti-IP 3 R2 in red, anti-STIM1 in green. Areas of co-localisation appear yellow due to color mixing.

    Techniques Used: Isolation, Staining, Microscopy

    The effect of Pyr10 on intracellular Ca 2+ after application of 1-oleoy1-2-acyl-sn-glycerol (OAG) 100 μM . The intact SANs were loaded with Ca 2+ indicator indo-1. (A,B) show intracellular Ca 2+ recordings from a WT mouse. (B,C) The statistics pool data shows percentage changes in resting Ca 2+ , Ca 2+ transient and firing rate in WT and TRPC3 KO mice respectively. (B) OAG treatment against control. (C) 2 μM Pyr10 treatment against OAG treatment. * P
    Figure Legend Snippet: The effect of Pyr10 on intracellular Ca 2+ after application of 1-oleoy1-2-acyl-sn-glycerol (OAG) 100 μM . The intact SANs were loaded with Ca 2+ indicator indo-1. (A,B) show intracellular Ca 2+ recordings from a WT mouse. (B,C) The statistics pool data shows percentage changes in resting Ca 2+ , Ca 2+ transient and firing rate in WT and TRPC3 KO mice respectively. (B) OAG treatment against control. (C) 2 μM Pyr10 treatment against OAG treatment. * P

    Techniques Used: Mouse Assay

    Signaling pathways involved in the activation of TRPC3 . The G-protein coupled receptor (GPCRs) activates phospholiase C (PLC), resulting in generation of IP3 and diacylglycerol (DAG). IP3 activates its receptor that leads to Ca 2+ release from SR/ER and the depletion of SR/ER Ca 2+ store. The Ca 2+ content change in the store can be sensed by STIM1, the ER Ca 2+ sensor and cause store-operated Ca 2+ entry (SOCE) through TRPC3 channels. An additional or alternate possibility is that DAG can directly activate TRPC3, and produce receptor-operated Ca 2+ entry (ROCE). GPCRs, G-protein coupled receptors; IP3R, inositol 1,4,5-trisphosphate receptors; PLC, Phospholipase C; ROCE, Receptor–operated Ca 2+ entry; SOCE, store-operated Ca 2+ entry; SR/ER, sarco- endo-plasmic reticulum; STIM1, stromal interacting molecule 1; TRPC3,4,6 canonical transient receptor potential channel types 3,4,6; NCX Na/Ca exchange.
    Figure Legend Snippet: Signaling pathways involved in the activation of TRPC3 . The G-protein coupled receptor (GPCRs) activates phospholiase C (PLC), resulting in generation of IP3 and diacylglycerol (DAG). IP3 activates its receptor that leads to Ca 2+ release from SR/ER and the depletion of SR/ER Ca 2+ store. The Ca 2+ content change in the store can be sensed by STIM1, the ER Ca 2+ sensor and cause store-operated Ca 2+ entry (SOCE) through TRPC3 channels. An additional or alternate possibility is that DAG can directly activate TRPC3, and produce receptor-operated Ca 2+ entry (ROCE). GPCRs, G-protein coupled receptors; IP3R, inositol 1,4,5-trisphosphate receptors; PLC, Phospholipase C; ROCE, Receptor–operated Ca 2+ entry; SOCE, store-operated Ca 2+ entry; SR/ER, sarco- endo-plasmic reticulum; STIM1, stromal interacting molecule 1; TRPC3,4,6 canonical transient receptor potential channel types 3,4,6; NCX Na/Ca exchange.

    Techniques Used: Activation Assay, Planar Chromatography

    5) Product Images from "Resistance to Store Depletion-induced Endothelial Injury in Rat Lung after Chronic Heart Failure"

    Article Title: Resistance to Store Depletion-induced Endothelial Injury in Rat Lung after Chronic Heart Failure

    Journal:

    doi: 10.1164/rccm.200506-847OC

    Immunostaining for transient receptor potential proteins TRPC1, TRPC3, TRPC4, and TRPC6/7 in rat lungs. Expression of each TRPC isoform in endothelium was confirmed in small muscular or partially muscularized extraalveolar vessels ( A ) and conduit artery
    Figure Legend Snippet: Immunostaining for transient receptor potential proteins TRPC1, TRPC3, TRPC4, and TRPC6/7 in rat lungs. Expression of each TRPC isoform in endothelium was confirmed in small muscular or partially muscularized extraalveolar vessels ( A ) and conduit artery

    Techniques Used: Immunostaining, Expressing

    6) Product Images from "The involvement of TRPC3 channels in sinoatrial arrhythmias"

    Article Title: The involvement of TRPC3 channels in sinoatrial arrhythmias

    Journal: Frontiers in Physiology

    doi: 10.3389/fphys.2015.00086

    The effect of angiotension II (Ang II) and a selective TRPC3 channel blocker Pyr10 on pacemaker action potential after application of Ang II. (A–D) Intracellular recordings from WT mice; (E–H ) Intracellular recordings from TRPC3 −/− mice with conditions as indicated in each panel.
    Figure Legend Snippet: The effect of angiotension II (Ang II) and a selective TRPC3 channel blocker Pyr10 on pacemaker action potential after application of Ang II. (A–D) Intracellular recordings from WT mice; (E–H ) Intracellular recordings from TRPC3 −/− mice with conditions as indicated in each panel.

    Techniques Used: Mouse Assay

    Atrial arrhythmias induced by Ang II and pacing . Reconstructed excitation maps of data from multi-electrode array recordings (images below) from a Langendorff-perfused isolated heart. (A) sinus rhythm and (B) pacing induced atrial tachycardia. Note in AT, the ectopic beat starts from a different site to sinus rhythms. (C) Representative ECG recordings from isolated heart from WT and TRPC3 −/− mice under control conditions. (D) Atrial tachycardia and atrial fibrillation induced by burst pacing in WT mice and atrial-ventricular conduction block recorded from a TRPC −/− mouse heart respectively. (E) On average, the arrhythmia index was significant reduced in TRPC3KO mice ( n = 8) compared to WT mice ( n = 11, P = 0.004).
    Figure Legend Snippet: Atrial arrhythmias induced by Ang II and pacing . Reconstructed excitation maps of data from multi-electrode array recordings (images below) from a Langendorff-perfused isolated heart. (A) sinus rhythm and (B) pacing induced atrial tachycardia. Note in AT, the ectopic beat starts from a different site to sinus rhythms. (C) Representative ECG recordings from isolated heart from WT and TRPC3 −/− mice under control conditions. (D) Atrial tachycardia and atrial fibrillation induced by burst pacing in WT mice and atrial-ventricular conduction block recorded from a TRPC −/− mouse heart respectively. (E) On average, the arrhythmia index was significant reduced in TRPC3KO mice ( n = 8) compared to WT mice ( n = 11, P = 0.004).

    Techniques Used: Isolation, Mouse Assay, Blocking Assay

    Isolated single SAN cells express TRPC3, STIM1, and IP 3 R2 . Confocal immunofluorenscence images of isolated single pacemaker cells. Isolated group (A) and single (B) pacemaker cells were positively stained with anti HCN4 (in green). (C) Anti-TRPC3 in red, anti-STIM1 in green. (D) Anti-IP3R2 in red, anti-STIM1 in green. (E) 3D reconstruction image using N-SIM microscopy. Anti-IP 3 R2 in red, anti-STIM1 in green. Areas of co-localisation appear yellow due to color mixing.
    Figure Legend Snippet: Isolated single SAN cells express TRPC3, STIM1, and IP 3 R2 . Confocal immunofluorenscence images of isolated single pacemaker cells. Isolated group (A) and single (B) pacemaker cells were positively stained with anti HCN4 (in green). (C) Anti-TRPC3 in red, anti-STIM1 in green. (D) Anti-IP3R2 in red, anti-STIM1 in green. (E) 3D reconstruction image using N-SIM microscopy. Anti-IP 3 R2 in red, anti-STIM1 in green. Areas of co-localisation appear yellow due to color mixing.

    Techniques Used: Isolation, Staining, Microscopy

    The effect of Pyr10 on intracellular Ca 2+ after application of 1-oleoy1-2-acyl-sn-glycerol (OAG) 100 μM . The intact SANs were loaded with Ca 2+ indicator indo-1. (A,B) show intracellular Ca 2+ recordings from a WT mouse. (B,C) The statistics pool data shows percentage changes in resting Ca 2+ , Ca 2+ transient and firing rate in WT and TRPC3 KO mice respectively. (B) OAG treatment against control. (C) 2 μM Pyr10 treatment against OAG treatment. * P
    Figure Legend Snippet: The effect of Pyr10 on intracellular Ca 2+ after application of 1-oleoy1-2-acyl-sn-glycerol (OAG) 100 μM . The intact SANs were loaded with Ca 2+ indicator indo-1. (A,B) show intracellular Ca 2+ recordings from a WT mouse. (B,C) The statistics pool data shows percentage changes in resting Ca 2+ , Ca 2+ transient and firing rate in WT and TRPC3 KO mice respectively. (B) OAG treatment against control. (C) 2 μM Pyr10 treatment against OAG treatment. * P

    Techniques Used: Mouse Assay

    Signaling pathways involved in the activation of TRPC3 . The G-protein coupled receptor (GPCRs) activates phospholiase C (PLC), resulting in generation of IP3 and diacylglycerol (DAG). IP3 activates its receptor that leads to Ca 2+ release from SR/ER and the depletion of SR/ER Ca 2+ store. The Ca 2+ content change in the store can be sensed by STIM1, the ER Ca 2+ sensor and cause store-operated Ca 2+ entry (SOCE) through TRPC3 channels. An additional or alternate possibility is that DAG can directly activate TRPC3, and produce receptor-operated Ca 2+ entry (ROCE). GPCRs, G-protein coupled receptors; IP3R, inositol 1,4,5-trisphosphate receptors; PLC, Phospholipase C; ROCE, Receptor–operated Ca 2+ entry; SOCE, store-operated Ca 2+ entry; SR/ER, sarco- endo-plasmic reticulum; STIM1, stromal interacting molecule 1; TRPC3,4,6 canonical transient receptor potential channel types 3,4,6; NCX Na/Ca exchange.
    Figure Legend Snippet: Signaling pathways involved in the activation of TRPC3 . The G-protein coupled receptor (GPCRs) activates phospholiase C (PLC), resulting in generation of IP3 and diacylglycerol (DAG). IP3 activates its receptor that leads to Ca 2+ release from SR/ER and the depletion of SR/ER Ca 2+ store. The Ca 2+ content change in the store can be sensed by STIM1, the ER Ca 2+ sensor and cause store-operated Ca 2+ entry (SOCE) through TRPC3 channels. An additional or alternate possibility is that DAG can directly activate TRPC3, and produce receptor-operated Ca 2+ entry (ROCE). GPCRs, G-protein coupled receptors; IP3R, inositol 1,4,5-trisphosphate receptors; PLC, Phospholipase C; ROCE, Receptor–operated Ca 2+ entry; SOCE, store-operated Ca 2+ entry; SR/ER, sarco- endo-plasmic reticulum; STIM1, stromal interacting molecule 1; TRPC3,4,6 canonical transient receptor potential channel types 3,4,6; NCX Na/Ca exchange.

    Techniques Used: Activation Assay, Planar Chromatography

    7) Product Images from "Overexpression of TRPC3 increases apoptosis but not necrosis in response to ischemia/reperfusion in adult mouse cardiomyocytes"

    Article Title: Overexpression of TRPC3 increases apoptosis but not necrosis in response to ischemia/reperfusion in adult mouse cardiomyocytes

    Journal: American journal of physiology. Cell physiology

    doi: 10.1152/ajpcell.00313.2007

    After 1h of isolation, cardiomyocytes from WT and TRPC3 transgenic mice were treated with 10 ng/ml TNF-α for 2h or 18h, viability and apoptosis were measured by Annexin V-propidium iodide staining. Data presented as mean ± SEM of six individual experiments, i.e., six separate mouse cardiomyocytes isolations; with at least 300 cells counted per experiment under each condition. * = p
    Figure Legend Snippet: After 1h of isolation, cardiomyocytes from WT and TRPC3 transgenic mice were treated with 10 ng/ml TNF-α for 2h or 18h, viability and apoptosis were measured by Annexin V-propidium iodide staining. Data presented as mean ± SEM of six individual experiments, i.e., six separate mouse cardiomyocytes isolations; with at least 300 cells counted per experiment under each condition. * = p

    Techniques Used: Isolation, Transgenic Assay, Mouse Assay, Staining

    A) Bright field phase contrast images of cardiomyocytes from wild-type (WT) and TRPC3 transgenic mice at baseline, at the end of 90 min ischemia and after 90 min ischemia and 3 hours reperfusion; B) cardiomyocytes from wild-type (WT) and TRPC3 transgenic mice after 90 min ischemia and 3 hours reperfusion showing Annexin V and propidium iodide staining. Arrows in merged image indicate necrotic cells, staining positive for both Annexin V and propidium iodide.
    Figure Legend Snippet: A) Bright field phase contrast images of cardiomyocytes from wild-type (WT) and TRPC3 transgenic mice at baseline, at the end of 90 min ischemia and after 90 min ischemia and 3 hours reperfusion; B) cardiomyocytes from wild-type (WT) and TRPC3 transgenic mice after 90 min ischemia and 3 hours reperfusion showing Annexin V and propidium iodide staining. Arrows in merged image indicate necrotic cells, staining positive for both Annexin V and propidium iodide.

    Techniques Used: Transgenic Assay, Mouse Assay, Staining

    A) Cell viability assessed by % rod shaped cells; B) % apoptotic cells indicated by Annexin V positive and propidium iodide negative staining; C) % necrotic cells indicated by Annexin V and propidium iodide positive staining in cardiomyocytes from wild-type (WT) and TRPC3 transgenic mice at baseline, at the end of 90 min ischemia and after 90 min ischemia and 3 hours reperfusion (I/R). Data presented as mean ± SEM of six individual experiments (i.e., six separate mouse cardiomyocytes isolations; with at least 300 cells counted per experiment under each condition). # = p
    Figure Legend Snippet: A) Cell viability assessed by % rod shaped cells; B) % apoptotic cells indicated by Annexin V positive and propidium iodide negative staining; C) % necrotic cells indicated by Annexin V and propidium iodide positive staining in cardiomyocytes from wild-type (WT) and TRPC3 transgenic mice at baseline, at the end of 90 min ischemia and after 90 min ischemia and 3 hours reperfusion (I/R). Data presented as mean ± SEM of six individual experiments (i.e., six separate mouse cardiomyocytes isolations; with at least 300 cells counted per experiment under each condition). # = p

    Techniques Used: Negative Staining, Staining, Transgenic Assay, Mouse Assay

    Cardiomyocytes from wild-type (WT) and TRPC3 transgenic mice were treated with 5 μM thapsigargin for 5 mins in the absence of extracellular Ca 2+ and were then exposed to 2.5 mM extracellular calcium. Upper panels show a typical time course of myocyte rounding following addition of 2.5 mM Ca 2+ in WT and TRPC3 cardiomyocytes. Data are mean ± SEM from 15 cells of at least 3 individual experiments. # = p
    Figure Legend Snippet: Cardiomyocytes from wild-type (WT) and TRPC3 transgenic mice were treated with 5 μM thapsigargin for 5 mins in the absence of extracellular Ca 2+ and were then exposed to 2.5 mM extracellular calcium. Upper panels show a typical time course of myocyte rounding following addition of 2.5 mM Ca 2+ in WT and TRPC3 cardiomyocytes. Data are mean ± SEM from 15 cells of at least 3 individual experiments. # = p

    Techniques Used: Transgenic Assay, Mouse Assay

    A) TRPC3 and B) TRPC1 protein expression in the hearts from wild-type (WT) and TRPC3 transgenic mice. Upper panels are representative immunoblots and the lower panels are mean densitometric data from 6 individual experiments normalized to β-actin. # = p
    Figure Legend Snippet: A) TRPC3 and B) TRPC1 protein expression in the hearts from wild-type (WT) and TRPC3 transgenic mice. Upper panels are representative immunoblots and the lower panels are mean densitometric data from 6 individual experiments normalized to β-actin. # = p

    Techniques Used: Expressing, Transgenic Assay, Mouse Assay, Western Blot

    8) Product Images from "The histone variant MacroH2A regulates Ca2+ influx through TRPC3 and TRPC6 channels"

    Article Title: The histone variant MacroH2A regulates Ca2+ influx through TRPC3 and TRPC6 channels

    Journal: Oncogenesis

    doi: 10.1038/oncsis.2013.40

    MacroH2A1 depletion enhances transcriptional potential of Ca 2+ binding protein-related genes. ( a ) MacroH2A1-regulated genes were analyzed by DAVID bioinformatics resources ( http://david.abcc.ncifcrf.gov ), and ontological classification of genes based on molecular function is presented as upregulated or downregulated gene groups. ( b ) For validation of microarray data, 12 genes that are related to Ca 2+ binding proteins and are upregulated in macroH2A1-depleted cells were subjected to qRT–PCR. Gapdh was used as an internal control gene. All expression values were normalized to the average of β-actin . ( c ) Trpc gene expression in control and macroH2A1-depleted LD611 cells was analyzed by qRT–PCR. ND, not detected. ( d ) Cell extracts from control and macroH2A1-depleted cells were immunoblotted with antibodies against TRPC3 and TRPC6. β-Actin was used as the internal control for loading. The analysis was performed in duplicates with comparable results. ( e ) Changes in intracellular cytosolic Ca 2+ concentration after macroH2A1 depletion were measured with the Ca 2+ -sensitive dye Fluo-8NW. ( f , g ) Control and macroH2A1-delepeted LD611 cells loaded with Fura-2 AM were stimulated with 100 μ M ATP. Representative traces of Ca 2+ in response to ATP are shown in ( f ), and changes in intracellular Ca 2+ were quantified in ( g ). ( h ) LD611 cells were stably transfected with control or macroH2A1.2 expression vectors, and the expression of macroH2A1.2 at the mRNA and protein levels was analyzed by qRT–PCR (left) and western blotting (right). ( i ) qRT–PCR was performed to check relative expressions of Ca 2+ binding-related genes, which are downregulated after macroH2A1.2 expression. ( j ) TRPC3 and TRPC6 protein levels in control and macroH2A1.2-transfected cell were evaluated by western blotting. ( k ) The intracellular Ca 2+ concentration was determined as in ( e ), but after macroH2A1.2 expression. Each bar in ( b , c , e , g – i , k ) represents the mean s.d. of three replicates in two independent experiments. * P
    Figure Legend Snippet: MacroH2A1 depletion enhances transcriptional potential of Ca 2+ binding protein-related genes. ( a ) MacroH2A1-regulated genes were analyzed by DAVID bioinformatics resources ( http://david.abcc.ncifcrf.gov ), and ontological classification of genes based on molecular function is presented as upregulated or downregulated gene groups. ( b ) For validation of microarray data, 12 genes that are related to Ca 2+ binding proteins and are upregulated in macroH2A1-depleted cells were subjected to qRT–PCR. Gapdh was used as an internal control gene. All expression values were normalized to the average of β-actin . ( c ) Trpc gene expression in control and macroH2A1-depleted LD611 cells was analyzed by qRT–PCR. ND, not detected. ( d ) Cell extracts from control and macroH2A1-depleted cells were immunoblotted with antibodies against TRPC3 and TRPC6. β-Actin was used as the internal control for loading. The analysis was performed in duplicates with comparable results. ( e ) Changes in intracellular cytosolic Ca 2+ concentration after macroH2A1 depletion were measured with the Ca 2+ -sensitive dye Fluo-8NW. ( f , g ) Control and macroH2A1-delepeted LD611 cells loaded with Fura-2 AM were stimulated with 100 μ M ATP. Representative traces of Ca 2+ in response to ATP are shown in ( f ), and changes in intracellular Ca 2+ were quantified in ( g ). ( h ) LD611 cells were stably transfected with control or macroH2A1.2 expression vectors, and the expression of macroH2A1.2 at the mRNA and protein levels was analyzed by qRT–PCR (left) and western blotting (right). ( i ) qRT–PCR was performed to check relative expressions of Ca 2+ binding-related genes, which are downregulated after macroH2A1.2 expression. ( j ) TRPC3 and TRPC6 protein levels in control and macroH2A1.2-transfected cell were evaluated by western blotting. ( k ) The intracellular Ca 2+ concentration was determined as in ( e ), but after macroH2A1.2 expression. Each bar in ( b , c , e , g – i , k ) represents the mean s.d. of three replicates in two independent experiments. * P

    Techniques Used: Binding Assay, Microarray, Quantitative RT-PCR, Expressing, Concentration Assay, Stable Transfection, Transfection, Western Blot

    Immunohistochemical staining of macroH2A1, TRPC3 and TRPC6 in tissue microarray. ( a ) Tissue microarrays containing 36 cases of bladder tumor with 12 normal tissues were subjected to immunohistochemistry with antibodies against macroH2A1, TRPC3 and TRPC6. High-power magnifications are shown for representative immunostaining samples. Bar, 50 μm. ( b ) Immunostaining scores of macroH2A1, TRPC3 and TRPC6 in bladder normal and malignant tissues. The graph indicates the percentage of sections with different scores (negative, weak, moderate and strong).
    Figure Legend Snippet: Immunohistochemical staining of macroH2A1, TRPC3 and TRPC6 in tissue microarray. ( a ) Tissue microarrays containing 36 cases of bladder tumor with 12 normal tissues were subjected to immunohistochemistry with antibodies against macroH2A1, TRPC3 and TRPC6. High-power magnifications are shown for representative immunostaining samples. Bar, 50 μm. ( b ) Immunostaining scores of macroH2A1, TRPC3 and TRPC6 in bladder normal and malignant tissues. The graph indicates the percentage of sections with different scores (negative, weak, moderate and strong).

    Techniques Used: Immunohistochemistry, Staining, Microarray, Immunostaining

    TRPC3/TRPC6 silencing results in loss of macroH2A1 function. ( a ) Cell proliferation assays were carried out in quadruplicate using cells depleted of TRPC3, TRPC6 and/or macroH2A1 as indicated. Each bar represents the mean s.d. of four replicates in three independent experiments. ( b ) Cell invasion assays were performed using cells depleted of TRPC3, TRPC6 and/or macroH2A1. Each bar represents the mean s.d. of three replicates in two independent experiments. * P
    Figure Legend Snippet: TRPC3/TRPC6 silencing results in loss of macroH2A1 function. ( a ) Cell proliferation assays were carried out in quadruplicate using cells depleted of TRPC3, TRPC6 and/or macroH2A1 as indicated. Each bar represents the mean s.d. of four replicates in three independent experiments. ( b ) Cell invasion assays were performed using cells depleted of TRPC3, TRPC6 and/or macroH2A1. Each bar represents the mean s.d. of three replicates in two independent experiments. * P

    Techniques Used:

    9) Product Images from "The involvement of TRPC3 channels in sinoatrial arrhythmias"

    Article Title: The involvement of TRPC3 channels in sinoatrial arrhythmias

    Journal: Frontiers in Physiology

    doi: 10.3389/fphys.2015.00086

    The effect of angiotension II (Ang II) and a selective TRPC3 channel blocker Pyr10 on pacemaker action potential after application of Ang II. (A–D) Intracellular recordings from WT mice; (E–H ) Intracellular recordings from TRPC3 −/− mice with conditions as indicated in each panel.
    Figure Legend Snippet: The effect of angiotension II (Ang II) and a selective TRPC3 channel blocker Pyr10 on pacemaker action potential after application of Ang II. (A–D) Intracellular recordings from WT mice; (E–H ) Intracellular recordings from TRPC3 −/− mice with conditions as indicated in each panel.

    Techniques Used: Mouse Assay

    Atrial arrhythmias induced by Ang II and pacing . Reconstructed excitation maps of data from multi-electrode array recordings (images below) from a Langendorff-perfused isolated heart. (A) sinus rhythm and (B) pacing induced atrial tachycardia. Note in AT, the ectopic beat starts from a different site to sinus rhythms. (C) Representative ECG recordings from isolated heart from WT and TRPC3 −/− mice under control conditions. (D) Atrial tachycardia and atrial fibrillation induced by burst pacing in WT mice and atrial-ventricular conduction block recorded from a TRPC −/− mouse heart respectively. (E) On average, the arrhythmia index was significant reduced in TRPC3KO mice ( n = 8) compared to WT mice ( n = 11, P = 0.004).
    Figure Legend Snippet: Atrial arrhythmias induced by Ang II and pacing . Reconstructed excitation maps of data from multi-electrode array recordings (images below) from a Langendorff-perfused isolated heart. (A) sinus rhythm and (B) pacing induced atrial tachycardia. Note in AT, the ectopic beat starts from a different site to sinus rhythms. (C) Representative ECG recordings from isolated heart from WT and TRPC3 −/− mice under control conditions. (D) Atrial tachycardia and atrial fibrillation induced by burst pacing in WT mice and atrial-ventricular conduction block recorded from a TRPC −/− mouse heart respectively. (E) On average, the arrhythmia index was significant reduced in TRPC3KO mice ( n = 8) compared to WT mice ( n = 11, P = 0.004).

    Techniques Used: Isolation, Mouse Assay, Blocking Assay

    Isolated single SAN cells express TRPC3, STIM1, and IP 3 R2 . Confocal immunofluorenscence images of isolated single pacemaker cells. Isolated group (A) and single (B) pacemaker cells were positively stained with anti HCN4 (in green). (C) Anti-TRPC3 in red, anti-STIM1 in green. (D) Anti-IP3R2 in red, anti-STIM1 in green. (E) 3D reconstruction image using N-SIM microscopy. Anti-IP 3 R2 in red, anti-STIM1 in green. Areas of co-localisation appear yellow due to color mixing.
    Figure Legend Snippet: Isolated single SAN cells express TRPC3, STIM1, and IP 3 R2 . Confocal immunofluorenscence images of isolated single pacemaker cells. Isolated group (A) and single (B) pacemaker cells were positively stained with anti HCN4 (in green). (C) Anti-TRPC3 in red, anti-STIM1 in green. (D) Anti-IP3R2 in red, anti-STIM1 in green. (E) 3D reconstruction image using N-SIM microscopy. Anti-IP 3 R2 in red, anti-STIM1 in green. Areas of co-localisation appear yellow due to color mixing.

    Techniques Used: Isolation, Staining, Microscopy

    The effect of Pyr10 on intracellular Ca 2+ after application of 1-oleoy1-2-acyl-sn-glycerol (OAG) 100 μM . The intact SANs were loaded with Ca 2+ indicator indo-1. (A,B) show intracellular Ca 2+ recordings from a WT mouse. (B,C) The statistics pool data shows percentage changes in resting Ca 2+ , Ca 2+ transient and firing rate in WT and TRPC3 KO mice respectively. (B) OAG treatment against control. (C) 2 μM Pyr10 treatment against OAG treatment. * P
    Figure Legend Snippet: The effect of Pyr10 on intracellular Ca 2+ after application of 1-oleoy1-2-acyl-sn-glycerol (OAG) 100 μM . The intact SANs were loaded with Ca 2+ indicator indo-1. (A,B) show intracellular Ca 2+ recordings from a WT mouse. (B,C) The statistics pool data shows percentage changes in resting Ca 2+ , Ca 2+ transient and firing rate in WT and TRPC3 KO mice respectively. (B) OAG treatment against control. (C) 2 μM Pyr10 treatment against OAG treatment. * P

    Techniques Used: Mouse Assay

    Signaling pathways involved in the activation of TRPC3 . The G-protein coupled receptor (GPCRs) activates phospholiase C (PLC), resulting in generation of IP3 and diacylglycerol (DAG). IP3 activates its receptor that leads to Ca 2+ release from SR/ER and the depletion of SR/ER Ca 2+ store. The Ca 2+ content change in the store can be sensed by STIM1, the ER Ca 2+ sensor and cause store-operated Ca 2+ entry (SOCE) through TRPC3 channels. An additional or alternate possibility is that DAG can directly activate TRPC3, and produce receptor-operated Ca 2+ entry (ROCE). GPCRs, G-protein coupled receptors; IP3R, inositol 1,4,5-trisphosphate receptors; PLC, Phospholipase C; ROCE, Receptor–operated Ca 2+ entry; SOCE, store-operated Ca 2+ entry; SR/ER, sarco- endo-plasmic reticulum; STIM1, stromal interacting molecule 1; TRPC3,4,6 canonical transient receptor potential channel types 3,4,6; NCX Na/Ca exchange.
    Figure Legend Snippet: Signaling pathways involved in the activation of TRPC3 . The G-protein coupled receptor (GPCRs) activates phospholiase C (PLC), resulting in generation of IP3 and diacylglycerol (DAG). IP3 activates its receptor that leads to Ca 2+ release from SR/ER and the depletion of SR/ER Ca 2+ store. The Ca 2+ content change in the store can be sensed by STIM1, the ER Ca 2+ sensor and cause store-operated Ca 2+ entry (SOCE) through TRPC3 channels. An additional or alternate possibility is that DAG can directly activate TRPC3, and produce receptor-operated Ca 2+ entry (ROCE). GPCRs, G-protein coupled receptors; IP3R, inositol 1,4,5-trisphosphate receptors; PLC, Phospholipase C; ROCE, Receptor–operated Ca 2+ entry; SOCE, store-operated Ca 2+ entry; SR/ER, sarco- endo-plasmic reticulum; STIM1, stromal interacting molecule 1; TRPC3,4,6 canonical transient receptor potential channel types 3,4,6; NCX Na/Ca exchange.

    Techniques Used: Activation Assay, Planar Chromatography

    10) Product Images from "The involvement of TRPC3 channels in sinoatrial arrhythmias"

    Article Title: The involvement of TRPC3 channels in sinoatrial arrhythmias

    Journal: Frontiers in Physiology

    doi: 10.3389/fphys.2015.00086

    The effect of angiotension II (Ang II) and a selective TRPC3 channel blocker Pyr10 on pacemaker action potential after application of Ang II. (A–D) Intracellular recordings from WT mice; (E–H ) Intracellular recordings from TRPC3 −/− mice with conditions as indicated in each panel.
    Figure Legend Snippet: The effect of angiotension II (Ang II) and a selective TRPC3 channel blocker Pyr10 on pacemaker action potential after application of Ang II. (A–D) Intracellular recordings from WT mice; (E–H ) Intracellular recordings from TRPC3 −/− mice with conditions as indicated in each panel.

    Techniques Used: Mouse Assay

    Atrial arrhythmias induced by Ang II and pacing . Reconstructed excitation maps of data from multi-electrode array recordings (images below) from a Langendorff-perfused isolated heart. (A) sinus rhythm and (B) pacing induced atrial tachycardia. Note in AT, the ectopic beat starts from a different site to sinus rhythms. (C) Representative ECG recordings from isolated heart from WT and TRPC3 −/− mice under control conditions. (D) Atrial tachycardia and atrial fibrillation induced by burst pacing in WT mice and atrial-ventricular conduction block recorded from a TRPC −/− mouse heart respectively. (E) On average, the arrhythmia index was significant reduced in TRPC3KO mice ( n = 8) compared to WT mice ( n = 11, P = 0.004).
    Figure Legend Snippet: Atrial arrhythmias induced by Ang II and pacing . Reconstructed excitation maps of data from multi-electrode array recordings (images below) from a Langendorff-perfused isolated heart. (A) sinus rhythm and (B) pacing induced atrial tachycardia. Note in AT, the ectopic beat starts from a different site to sinus rhythms. (C) Representative ECG recordings from isolated heart from WT and TRPC3 −/− mice under control conditions. (D) Atrial tachycardia and atrial fibrillation induced by burst pacing in WT mice and atrial-ventricular conduction block recorded from a TRPC −/− mouse heart respectively. (E) On average, the arrhythmia index was significant reduced in TRPC3KO mice ( n = 8) compared to WT mice ( n = 11, P = 0.004).

    Techniques Used: Isolation, Mouse Assay, Blocking Assay

    Isolated single SAN cells express TRPC3, STIM1, and IP 3 R2 . Confocal immunofluorenscence images of isolated single pacemaker cells. Isolated group (A) and single (B) pacemaker cells were positively stained with anti HCN4 (in green). (C) Anti-TRPC3 in red, anti-STIM1 in green. (D) Anti-IP3R2 in red, anti-STIM1 in green. (E) 3D reconstruction image using N-SIM microscopy. Anti-IP 3 R2 in red, anti-STIM1 in green. Areas of co-localisation appear yellow due to color mixing.
    Figure Legend Snippet: Isolated single SAN cells express TRPC3, STIM1, and IP 3 R2 . Confocal immunofluorenscence images of isolated single pacemaker cells. Isolated group (A) and single (B) pacemaker cells were positively stained with anti HCN4 (in green). (C) Anti-TRPC3 in red, anti-STIM1 in green. (D) Anti-IP3R2 in red, anti-STIM1 in green. (E) 3D reconstruction image using N-SIM microscopy. Anti-IP 3 R2 in red, anti-STIM1 in green. Areas of co-localisation appear yellow due to color mixing.

    Techniques Used: Isolation, Staining, Microscopy

    The effect of Pyr10 on intracellular Ca 2+ after application of 1-oleoy1-2-acyl-sn-glycerol (OAG) 100 μM . The intact SANs were loaded with Ca 2+ indicator indo-1. (A,B) show intracellular Ca 2+ recordings from a WT mouse. (B,C) The statistics pool data shows percentage changes in resting Ca 2+ , Ca 2+ transient and firing rate in WT and TRPC3 KO mice respectively. (B) OAG treatment against control. (C) 2 μM Pyr10 treatment against OAG treatment. * P
    Figure Legend Snippet: The effect of Pyr10 on intracellular Ca 2+ after application of 1-oleoy1-2-acyl-sn-glycerol (OAG) 100 μM . The intact SANs were loaded with Ca 2+ indicator indo-1. (A,B) show intracellular Ca 2+ recordings from a WT mouse. (B,C) The statistics pool data shows percentage changes in resting Ca 2+ , Ca 2+ transient and firing rate in WT and TRPC3 KO mice respectively. (B) OAG treatment against control. (C) 2 μM Pyr10 treatment against OAG treatment. * P

    Techniques Used: Mouse Assay

    Signaling pathways involved in the activation of TRPC3 . The G-protein coupled receptor (GPCRs) activates phospholiase C (PLC), resulting in generation of IP3 and diacylglycerol (DAG). IP3 activates its receptor that leads to Ca 2+ release from SR/ER and the depletion of SR/ER Ca 2+ store. The Ca 2+ content change in the store can be sensed by STIM1, the ER Ca 2+ sensor and cause store-operated Ca 2+ entry (SOCE) through TRPC3 channels. An additional or alternate possibility is that DAG can directly activate TRPC3, and produce receptor-operated Ca 2+ entry (ROCE). GPCRs, G-protein coupled receptors; IP3R, inositol 1,4,5-trisphosphate receptors; PLC, Phospholipase C; ROCE, Receptor–operated Ca 2+ entry; SOCE, store-operated Ca 2+ entry; SR/ER, sarco- endo-plasmic reticulum; STIM1, stromal interacting molecule 1; TRPC3,4,6 canonical transient receptor potential channel types 3,4,6; NCX Na/Ca exchange.
    Figure Legend Snippet: Signaling pathways involved in the activation of TRPC3 . The G-protein coupled receptor (GPCRs) activates phospholiase C (PLC), resulting in generation of IP3 and diacylglycerol (DAG). IP3 activates its receptor that leads to Ca 2+ release from SR/ER and the depletion of SR/ER Ca 2+ store. The Ca 2+ content change in the store can be sensed by STIM1, the ER Ca 2+ sensor and cause store-operated Ca 2+ entry (SOCE) through TRPC3 channels. An additional or alternate possibility is that DAG can directly activate TRPC3, and produce receptor-operated Ca 2+ entry (ROCE). GPCRs, G-protein coupled receptors; IP3R, inositol 1,4,5-trisphosphate receptors; PLC, Phospholipase C; ROCE, Receptor–operated Ca 2+ entry; SOCE, store-operated Ca 2+ entry; SR/ER, sarco- endo-plasmic reticulum; STIM1, stromal interacting molecule 1; TRPC3,4,6 canonical transient receptor potential channel types 3,4,6; NCX Na/Ca exchange.

    Techniques Used: Activation Assay, Planar Chromatography

    11) Product Images from "The involvement of TRPC3 channels in sinoatrial arrhythmias"

    Article Title: The involvement of TRPC3 channels in sinoatrial arrhythmias

    Journal: Frontiers in Physiology

    doi: 10.3389/fphys.2015.00086

    The effect of angiotension II (Ang II) and a selective TRPC3 channel blocker Pyr10 on pacemaker action potential after application of Ang II. (A–D) Intracellular recordings from WT mice; (E–H ) Intracellular recordings from TRPC3 −/− mice with conditions as indicated in each panel.
    Figure Legend Snippet: The effect of angiotension II (Ang II) and a selective TRPC3 channel blocker Pyr10 on pacemaker action potential after application of Ang II. (A–D) Intracellular recordings from WT mice; (E–H ) Intracellular recordings from TRPC3 −/− mice with conditions as indicated in each panel.

    Techniques Used: Mouse Assay

    Atrial arrhythmias induced by Ang II and pacing . Reconstructed excitation maps of data from multi-electrode array recordings (images below) from a Langendorff-perfused isolated heart. (A) sinus rhythm and (B) pacing induced atrial tachycardia. Note in AT, the ectopic beat starts from a different site to sinus rhythms. (C) Representative ECG recordings from isolated heart from WT and TRPC3 −/− mice under control conditions. (D) Atrial tachycardia and atrial fibrillation induced by burst pacing in WT mice and atrial-ventricular conduction block recorded from a TRPC −/− mouse heart respectively. (E) On average, the arrhythmia index was significant reduced in TRPC3KO mice ( n = 8) compared to WT mice ( n = 11, P = 0.004).
    Figure Legend Snippet: Atrial arrhythmias induced by Ang II and pacing . Reconstructed excitation maps of data from multi-electrode array recordings (images below) from a Langendorff-perfused isolated heart. (A) sinus rhythm and (B) pacing induced atrial tachycardia. Note in AT, the ectopic beat starts from a different site to sinus rhythms. (C) Representative ECG recordings from isolated heart from WT and TRPC3 −/− mice under control conditions. (D) Atrial tachycardia and atrial fibrillation induced by burst pacing in WT mice and atrial-ventricular conduction block recorded from a TRPC −/− mouse heart respectively. (E) On average, the arrhythmia index was significant reduced in TRPC3KO mice ( n = 8) compared to WT mice ( n = 11, P = 0.004).

    Techniques Used: Isolation, Mouse Assay, Blocking Assay

    Isolated single SAN cells express TRPC3, STIM1, and IP 3 R2 . Confocal immunofluorenscence images of isolated single pacemaker cells. Isolated group (A) and single (B) pacemaker cells were positively stained with anti HCN4 (in green). (C) Anti-TRPC3 in red, anti-STIM1 in green. (D) Anti-IP3R2 in red, anti-STIM1 in green. (E) 3D reconstruction image using N-SIM microscopy. Anti-IP 3 R2 in red, anti-STIM1 in green. Areas of co-localisation appear yellow due to color mixing.
    Figure Legend Snippet: Isolated single SAN cells express TRPC3, STIM1, and IP 3 R2 . Confocal immunofluorenscence images of isolated single pacemaker cells. Isolated group (A) and single (B) pacemaker cells were positively stained with anti HCN4 (in green). (C) Anti-TRPC3 in red, anti-STIM1 in green. (D) Anti-IP3R2 in red, anti-STIM1 in green. (E) 3D reconstruction image using N-SIM microscopy. Anti-IP 3 R2 in red, anti-STIM1 in green. Areas of co-localisation appear yellow due to color mixing.

    Techniques Used: Isolation, Staining, Microscopy

    The effect of Pyr10 on intracellular Ca 2+ after application of 1-oleoy1-2-acyl-sn-glycerol (OAG) 100 μM . The intact SANs were loaded with Ca 2+ indicator indo-1. (A,B) show intracellular Ca 2+ recordings from a WT mouse. (B,C) The statistics pool data shows percentage changes in resting Ca 2+ , Ca 2+ transient and firing rate in WT and TRPC3 KO mice respectively. (B) OAG treatment against control. (C) 2 μM Pyr10 treatment against OAG treatment. * P
    Figure Legend Snippet: The effect of Pyr10 on intracellular Ca 2+ after application of 1-oleoy1-2-acyl-sn-glycerol (OAG) 100 μM . The intact SANs were loaded with Ca 2+ indicator indo-1. (A,B) show intracellular Ca 2+ recordings from a WT mouse. (B,C) The statistics pool data shows percentage changes in resting Ca 2+ , Ca 2+ transient and firing rate in WT and TRPC3 KO mice respectively. (B) OAG treatment against control. (C) 2 μM Pyr10 treatment against OAG treatment. * P

    Techniques Used: Mouse Assay

    Signaling pathways involved in the activation of TRPC3 . The G-protein coupled receptor (GPCRs) activates phospholiase C (PLC), resulting in generation of IP3 and diacylglycerol (DAG). IP3 activates its receptor that leads to Ca 2+ release from SR/ER and the depletion of SR/ER Ca 2+ store. The Ca 2+ content change in the store can be sensed by STIM1, the ER Ca 2+ sensor and cause store-operated Ca 2+ entry (SOCE) through TRPC3 channels. An additional or alternate possibility is that DAG can directly activate TRPC3, and produce receptor-operated Ca 2+ entry (ROCE). GPCRs, G-protein coupled receptors; IP3R, inositol 1,4,5-trisphosphate receptors; PLC, Phospholipase C; ROCE, Receptor–operated Ca 2+ entry; SOCE, store-operated Ca 2+ entry; SR/ER, sarco- endo-plasmic reticulum; STIM1, stromal interacting molecule 1; TRPC3,4,6 canonical transient receptor potential channel types 3,4,6; NCX Na/Ca exchange.
    Figure Legend Snippet: Signaling pathways involved in the activation of TRPC3 . The G-protein coupled receptor (GPCRs) activates phospholiase C (PLC), resulting in generation of IP3 and diacylglycerol (DAG). IP3 activates its receptor that leads to Ca 2+ release from SR/ER and the depletion of SR/ER Ca 2+ store. The Ca 2+ content change in the store can be sensed by STIM1, the ER Ca 2+ sensor and cause store-operated Ca 2+ entry (SOCE) through TRPC3 channels. An additional or alternate possibility is that DAG can directly activate TRPC3, and produce receptor-operated Ca 2+ entry (ROCE). GPCRs, G-protein coupled receptors; IP3R, inositol 1,4,5-trisphosphate receptors; PLC, Phospholipase C; ROCE, Receptor–operated Ca 2+ entry; SOCE, store-operated Ca 2+ entry; SR/ER, sarco- endo-plasmic reticulum; STIM1, stromal interacting molecule 1; TRPC3,4,6 canonical transient receptor potential channel types 3,4,6; NCX Na/Ca exchange.

    Techniques Used: Activation Assay, Planar Chromatography

    12) Product Images from "The involvement of TRPC3 channels in sinoatrial arrhythmias"

    Article Title: The involvement of TRPC3 channels in sinoatrial arrhythmias

    Journal: Frontiers in Physiology

    doi: 10.3389/fphys.2015.00086

    The effect of angiotension II (Ang II) and a selective TRPC3 channel blocker Pyr10 on pacemaker action potential after application of Ang II. (A–D) Intracellular recordings from WT mice; (E–H ) Intracellular recordings from TRPC3 −/− mice with conditions as indicated in each panel.
    Figure Legend Snippet: The effect of angiotension II (Ang II) and a selective TRPC3 channel blocker Pyr10 on pacemaker action potential after application of Ang II. (A–D) Intracellular recordings from WT mice; (E–H ) Intracellular recordings from TRPC3 −/− mice with conditions as indicated in each panel.

    Techniques Used: Mouse Assay

    Atrial arrhythmias induced by Ang II and pacing . Reconstructed excitation maps of data from multi-electrode array recordings (images below) from a Langendorff-perfused isolated heart. (A) sinus rhythm and (B) pacing induced atrial tachycardia. Note in AT, the ectopic beat starts from a different site to sinus rhythms. (C) Representative ECG recordings from isolated heart from WT and TRPC3 −/− mice under control conditions. (D) Atrial tachycardia and atrial fibrillation induced by burst pacing in WT mice and atrial-ventricular conduction block recorded from a TRPC −/− mouse heart respectively. (E) On average, the arrhythmia index was significant reduced in TRPC3KO mice ( n = 8) compared to WT mice ( n = 11, P = 0.004).
    Figure Legend Snippet: Atrial arrhythmias induced by Ang II and pacing . Reconstructed excitation maps of data from multi-electrode array recordings (images below) from a Langendorff-perfused isolated heart. (A) sinus rhythm and (B) pacing induced atrial tachycardia. Note in AT, the ectopic beat starts from a different site to sinus rhythms. (C) Representative ECG recordings from isolated heart from WT and TRPC3 −/− mice under control conditions. (D) Atrial tachycardia and atrial fibrillation induced by burst pacing in WT mice and atrial-ventricular conduction block recorded from a TRPC −/− mouse heart respectively. (E) On average, the arrhythmia index was significant reduced in TRPC3KO mice ( n = 8) compared to WT mice ( n = 11, P = 0.004).

    Techniques Used: Isolation, Mouse Assay, Blocking Assay

    Isolated single SAN cells express TRPC3, STIM1, and IP 3 R2 . Confocal immunofluorenscence images of isolated single pacemaker cells. Isolated group (A) and single (B) pacemaker cells were positively stained with anti HCN4 (in green). (C) Anti-TRPC3 in red, anti-STIM1 in green. (D) Anti-IP3R2 in red, anti-STIM1 in green. (E) 3D reconstruction image using N-SIM microscopy. Anti-IP 3 R2 in red, anti-STIM1 in green. Areas of co-localisation appear yellow due to color mixing.
    Figure Legend Snippet: Isolated single SAN cells express TRPC3, STIM1, and IP 3 R2 . Confocal immunofluorenscence images of isolated single pacemaker cells. Isolated group (A) and single (B) pacemaker cells were positively stained with anti HCN4 (in green). (C) Anti-TRPC3 in red, anti-STIM1 in green. (D) Anti-IP3R2 in red, anti-STIM1 in green. (E) 3D reconstruction image using N-SIM microscopy. Anti-IP 3 R2 in red, anti-STIM1 in green. Areas of co-localisation appear yellow due to color mixing.

    Techniques Used: Isolation, Staining, Microscopy

    The effect of Pyr10 on intracellular Ca 2+ after application of 1-oleoy1-2-acyl-sn-glycerol (OAG) 100 μM . The intact SANs were loaded with Ca 2+ indicator indo-1. (A,B) show intracellular Ca 2+ recordings from a WT mouse. (B,C) The statistics pool data shows percentage changes in resting Ca 2+ , Ca 2+ transient and firing rate in WT and TRPC3 KO mice respectively. (B) OAG treatment against control. (C) 2 μM Pyr10 treatment against OAG treatment. * P
    Figure Legend Snippet: The effect of Pyr10 on intracellular Ca 2+ after application of 1-oleoy1-2-acyl-sn-glycerol (OAG) 100 μM . The intact SANs were loaded with Ca 2+ indicator indo-1. (A,B) show intracellular Ca 2+ recordings from a WT mouse. (B,C) The statistics pool data shows percentage changes in resting Ca 2+ , Ca 2+ transient and firing rate in WT and TRPC3 KO mice respectively. (B) OAG treatment against control. (C) 2 μM Pyr10 treatment against OAG treatment. * P

    Techniques Used: Mouse Assay

    Signaling pathways involved in the activation of TRPC3 . The G-protein coupled receptor (GPCRs) activates phospholiase C (PLC), resulting in generation of IP3 and diacylglycerol (DAG). IP3 activates its receptor that leads to Ca 2+ release from SR/ER and the depletion of SR/ER Ca 2+ store. The Ca 2+ content change in the store can be sensed by STIM1, the ER Ca 2+ sensor and cause store-operated Ca 2+ entry (SOCE) through TRPC3 channels. An additional or alternate possibility is that DAG can directly activate TRPC3, and produce receptor-operated Ca 2+ entry (ROCE). GPCRs, G-protein coupled receptors; IP3R, inositol 1,4,5-trisphosphate receptors; PLC, Phospholipase C; ROCE, Receptor–operated Ca 2+ entry; SOCE, store-operated Ca 2+ entry; SR/ER, sarco- endo-plasmic reticulum; STIM1, stromal interacting molecule 1; TRPC3,4,6 canonical transient receptor potential channel types 3,4,6; NCX Na/Ca exchange.
    Figure Legend Snippet: Signaling pathways involved in the activation of TRPC3 . The G-protein coupled receptor (GPCRs) activates phospholiase C (PLC), resulting in generation of IP3 and diacylglycerol (DAG). IP3 activates its receptor that leads to Ca 2+ release from SR/ER and the depletion of SR/ER Ca 2+ store. The Ca 2+ content change in the store can be sensed by STIM1, the ER Ca 2+ sensor and cause store-operated Ca 2+ entry (SOCE) through TRPC3 channels. An additional or alternate possibility is that DAG can directly activate TRPC3, and produce receptor-operated Ca 2+ entry (ROCE). GPCRs, G-protein coupled receptors; IP3R, inositol 1,4,5-trisphosphate receptors; PLC, Phospholipase C; ROCE, Receptor–operated Ca 2+ entry; SOCE, store-operated Ca 2+ entry; SR/ER, sarco- endo-plasmic reticulum; STIM1, stromal interacting molecule 1; TRPC3,4,6 canonical transient receptor potential channel types 3,4,6; NCX Na/Ca exchange.

    Techniques Used: Activation Assay, Planar Chromatography

    13) Product Images from "TRPC3 cation channel plays an important role in proliferation and differentiation of skeletal muscle myoblasts"

    Article Title: TRPC3 cation channel plays an important role in proliferation and differentiation of skeletal muscle myoblasts

    Journal: Experimental & Molecular Medicine

    doi: 10.3858/emm.2010.42.9.061

    An increase in the SR Ca 2+ content in MDG myoblasts in the presence of Pyr3, a specific blocker of TRPC3. The SR Ca 2+ of MDG myoblasts loaded with fura-2 was depleted by treatment with 10 µM CPA in the absence of extracellular Ca 2+ and presence
    Figure Legend Snippet: An increase in the SR Ca 2+ content in MDG myoblasts in the presence of Pyr3, a specific blocker of TRPC3. The SR Ca 2+ of MDG myoblasts loaded with fura-2 was depleted by treatment with 10 µM CPA in the absence of extracellular Ca 2+ and presence

    Techniques Used:

    Morphological changes in MDG/TRPC3 KD myoblasts. (A) Images of MDG/TRPC3 KD myoblasts plated on 10-cm dishes coated with collagen were obtained. Myoblasts in numbered boxes were enlarged and are presented in the lower panel (two boxes for wild-type myoblasts
    Figure Legend Snippet: Morphological changes in MDG/TRPC3 KD myoblasts. (A) Images of MDG/TRPC3 KD myoblasts plated on 10-cm dishes coated with collagen were obtained. Myoblasts in numbered boxes were enlarged and are presented in the lower panel (two boxes for wild-type myoblasts

    Techniques Used:

    Apoptotic cell death of MDG/TRPC3 KD myoblasts during exposure to differentiation conditions. (A) Fully and successfully differentiated wild-type myoblasts are shown in the upper panel. D0 to D6 means differentiation day zero to six. MDG/TRPC3 KD myoblasts
    Figure Legend Snippet: Apoptotic cell death of MDG/TRPC3 KD myoblasts during exposure to differentiation conditions. (A) Fully and successfully differentiated wild-type myoblasts are shown in the upper panel. D0 to D6 means differentiation day zero to six. MDG/TRPC3 KD myoblasts

    Techniques Used:

    TRPC3 and DHPR could be a system of checks and double-checks for skeletal myoblast differentiation by playing a redundant role
    Figure Legend Snippet: TRPC3 and DHPR could be a system of checks and double-checks for skeletal myoblast differentiation by playing a redundant role

    Techniques Used:

    Increases in both the SR Ca 2+ content and resting cytoplasmic Ca 2+ level in MDG/TRPC3 KD myoblasts. (A) the SR Ca 2+ of wild-type or MDG/TRPC3 KD myoblasts loaded with fura-2 was depleted by treatment with 10 µM cyclopiazonic acid (CPA) in the
    Figure Legend Snippet: Increases in both the SR Ca 2+ content and resting cytoplasmic Ca 2+ level in MDG/TRPC3 KD myoblasts. (A) the SR Ca 2+ of wild-type or MDG/TRPC3 KD myoblasts loaded with fura-2 was depleted by treatment with 10 µM cyclopiazonic acid (CPA) in the

    Techniques Used:

    Decreases in the expression level of TRPC4, Orai1, and MG29 in MDG/TRPC3 KD myoblasts. Solubilized cell lysate from wild-type or MDG/TRPC3 KD myoblasts (30 µg of total protein) was subjected to SDS-PAGE (10% or 12% gel) and immunoblot assay with
    Figure Legend Snippet: Decreases in the expression level of TRPC4, Orai1, and MG29 in MDG/TRPC3 KD myoblasts. Solubilized cell lysate from wild-type or MDG/TRPC3 KD myoblasts (30 µg of total protein) was subjected to SDS-PAGE (10% or 12% gel) and immunoblot assay with

    Techniques Used: Expressing, SDS Page

    Creation of the MDG/TRPC3 KD myoblast line. (A) To obtain the virus to interfere with TRPC3 mRNA in α1 S DHPR-null MDG myoblasts, a retroviral vector and two different sequences (Sequences I and II) complementary to regions of TRPC3 mRNA were used.
    Figure Legend Snippet: Creation of the MDG/TRPC3 KD myoblast line. (A) To obtain the virus to interfere with TRPC3 mRNA in α1 S DHPR-null MDG myoblasts, a retroviral vector and two different sequences (Sequences I and II) complementary to regions of TRPC3 mRNA were used.

    Techniques Used: Plasmid Preparation

    14) Product Images from "Resistance to Store Depletion-induced Endothelial Injury in Rat Lung after Chronic Heart Failure"

    Article Title: Resistance to Store Depletion-induced Endothelial Injury in Rat Lung after Chronic Heart Failure

    Journal:

    doi: 10.1164/rccm.200506-847OC

    Immunostaining for transient receptor potential proteins TRPC1, TRPC3, TRPC4, and TRPC6/7 in rat lungs. Expression of each TRPC isoform in endothelium was confirmed in small muscular or partially muscularized extraalveolar vessels ( A ) and conduit artery
    Figure Legend Snippet: Immunostaining for transient receptor potential proteins TRPC1, TRPC3, TRPC4, and TRPC6/7 in rat lungs. Expression of each TRPC isoform in endothelium was confirmed in small muscular or partially muscularized extraalveolar vessels ( A ) and conduit artery

    Techniques Used: Immunostaining, Expressing

    15) Product Images from "The involvement of TRPC3 channels in sinoatrial arrhythmias"

    Article Title: The involvement of TRPC3 channels in sinoatrial arrhythmias

    Journal: Frontiers in Physiology

    doi: 10.3389/fphys.2015.00086

    The effect of angiotension II (Ang II) and a selective TRPC3 channel blocker Pyr10 on pacemaker action potential after application of Ang II. (A–D) Intracellular recordings from WT mice; (E–H ) Intracellular recordings from TRPC3 −/− mice with conditions as indicated in each panel.
    Figure Legend Snippet: The effect of angiotension II (Ang II) and a selective TRPC3 channel blocker Pyr10 on pacemaker action potential after application of Ang II. (A–D) Intracellular recordings from WT mice; (E–H ) Intracellular recordings from TRPC3 −/− mice with conditions as indicated in each panel.

    Techniques Used: Mouse Assay

    Atrial arrhythmias induced by Ang II and pacing . Reconstructed excitation maps of data from multi-electrode array recordings (images below) from a Langendorff-perfused isolated heart. (A) sinus rhythm and (B) pacing induced atrial tachycardia. Note in AT, the ectopic beat starts from a different site to sinus rhythms. (C) Representative ECG recordings from isolated heart from WT and TRPC3 −/− mice under control conditions. (D) Atrial tachycardia and atrial fibrillation induced by burst pacing in WT mice and atrial-ventricular conduction block recorded from a TRPC −/− mouse heart respectively. (E) On average, the arrhythmia index was significant reduced in TRPC3KO mice ( n = 8) compared to WT mice ( n = 11, P = 0.004).
    Figure Legend Snippet: Atrial arrhythmias induced by Ang II and pacing . Reconstructed excitation maps of data from multi-electrode array recordings (images below) from a Langendorff-perfused isolated heart. (A) sinus rhythm and (B) pacing induced atrial tachycardia. Note in AT, the ectopic beat starts from a different site to sinus rhythms. (C) Representative ECG recordings from isolated heart from WT and TRPC3 −/− mice under control conditions. (D) Atrial tachycardia and atrial fibrillation induced by burst pacing in WT mice and atrial-ventricular conduction block recorded from a TRPC −/− mouse heart respectively. (E) On average, the arrhythmia index was significant reduced in TRPC3KO mice ( n = 8) compared to WT mice ( n = 11, P = 0.004).

    Techniques Used: Isolation, Mouse Assay, Blocking Assay

    Isolated single SAN cells express TRPC3, STIM1, and IP 3 R2 . Confocal immunofluorenscence images of isolated single pacemaker cells. Isolated group (A) and single (B) pacemaker cells were positively stained with anti HCN4 (in green). (C) Anti-TRPC3 in red, anti-STIM1 in green. (D) Anti-IP3R2 in red, anti-STIM1 in green. (E) 3D reconstruction image using N-SIM microscopy. Anti-IP 3 R2 in red, anti-STIM1 in green. Areas of co-localisation appear yellow due to color mixing.
    Figure Legend Snippet: Isolated single SAN cells express TRPC3, STIM1, and IP 3 R2 . Confocal immunofluorenscence images of isolated single pacemaker cells. Isolated group (A) and single (B) pacemaker cells were positively stained with anti HCN4 (in green). (C) Anti-TRPC3 in red, anti-STIM1 in green. (D) Anti-IP3R2 in red, anti-STIM1 in green. (E) 3D reconstruction image using N-SIM microscopy. Anti-IP 3 R2 in red, anti-STIM1 in green. Areas of co-localisation appear yellow due to color mixing.

    Techniques Used: Isolation, Staining, Microscopy

    The effect of Pyr10 on intracellular Ca 2+ after application of 1-oleoy1-2-acyl-sn-glycerol (OAG) 100 μM . The intact SANs were loaded with Ca 2+ indicator indo-1. (A,B) show intracellular Ca 2+ recordings from a WT mouse. (B,C) The statistics pool data shows percentage changes in resting Ca 2+ , Ca 2+ transient and firing rate in WT and TRPC3 KO mice respectively. (B) OAG treatment against control. (C) 2 μM Pyr10 treatment against OAG treatment. * P
    Figure Legend Snippet: The effect of Pyr10 on intracellular Ca 2+ after application of 1-oleoy1-2-acyl-sn-glycerol (OAG) 100 μM . The intact SANs were loaded with Ca 2+ indicator indo-1. (A,B) show intracellular Ca 2+ recordings from a WT mouse. (B,C) The statistics pool data shows percentage changes in resting Ca 2+ , Ca 2+ transient and firing rate in WT and TRPC3 KO mice respectively. (B) OAG treatment against control. (C) 2 μM Pyr10 treatment against OAG treatment. * P

    Techniques Used: Mouse Assay

    Signaling pathways involved in the activation of TRPC3 . The G-protein coupled receptor (GPCRs) activates phospholiase C (PLC), resulting in generation of IP3 and diacylglycerol (DAG). IP3 activates its receptor that leads to Ca 2+ release from SR/ER and the depletion of SR/ER Ca 2+ store. The Ca 2+ content change in the store can be sensed by STIM1, the ER Ca 2+ sensor and cause store-operated Ca 2+ entry (SOCE) through TRPC3 channels. An additional or alternate possibility is that DAG can directly activate TRPC3, and produce receptor-operated Ca 2+ entry (ROCE). GPCRs, G-protein coupled receptors; IP3R, inositol 1,4,5-trisphosphate receptors; PLC, Phospholipase C; ROCE, Receptor–operated Ca 2+ entry; SOCE, store-operated Ca 2+ entry; SR/ER, sarco- endo-plasmic reticulum; STIM1, stromal interacting molecule 1; TRPC3,4,6 canonical transient receptor potential channel types 3,4,6; NCX Na/Ca exchange.
    Figure Legend Snippet: Signaling pathways involved in the activation of TRPC3 . The G-protein coupled receptor (GPCRs) activates phospholiase C (PLC), resulting in generation of IP3 and diacylglycerol (DAG). IP3 activates its receptor that leads to Ca 2+ release from SR/ER and the depletion of SR/ER Ca 2+ store. The Ca 2+ content change in the store can be sensed by STIM1, the ER Ca 2+ sensor and cause store-operated Ca 2+ entry (SOCE) through TRPC3 channels. An additional or alternate possibility is that DAG can directly activate TRPC3, and produce receptor-operated Ca 2+ entry (ROCE). GPCRs, G-protein coupled receptors; IP3R, inositol 1,4,5-trisphosphate receptors; PLC, Phospholipase C; ROCE, Receptor–operated Ca 2+ entry; SOCE, store-operated Ca 2+ entry; SR/ER, sarco- endo-plasmic reticulum; STIM1, stromal interacting molecule 1; TRPC3,4,6 canonical transient receptor potential channel types 3,4,6; NCX Na/Ca exchange.

    Techniques Used: Activation Assay, Planar Chromatography

    16) Product Images from "The involvement of TRPC3 channels in sinoatrial arrhythmias"

    Article Title: The involvement of TRPC3 channels in sinoatrial arrhythmias

    Journal: Frontiers in Physiology

    doi: 10.3389/fphys.2015.00086

    The effect of angiotension II (Ang II) and a selective TRPC3 channel blocker Pyr10 on pacemaker action potential after application of Ang II. (A–D) Intracellular recordings from WT mice; (E–H ) Intracellular recordings from TRPC3 −/− mice with conditions as indicated in each panel.
    Figure Legend Snippet: The effect of angiotension II (Ang II) and a selective TRPC3 channel blocker Pyr10 on pacemaker action potential after application of Ang II. (A–D) Intracellular recordings from WT mice; (E–H ) Intracellular recordings from TRPC3 −/− mice with conditions as indicated in each panel.

    Techniques Used: Mouse Assay

    Atrial arrhythmias induced by Ang II and pacing . Reconstructed excitation maps of data from multi-electrode array recordings (images below) from a Langendorff-perfused isolated heart. (A) sinus rhythm and (B) pacing induced atrial tachycardia. Note in AT, the ectopic beat starts from a different site to sinus rhythms. (C) Representative ECG recordings from isolated heart from WT and TRPC3 −/− mice under control conditions. (D) Atrial tachycardia and atrial fibrillation induced by burst pacing in WT mice and atrial-ventricular conduction block recorded from a TRPC −/− mouse heart respectively. (E) On average, the arrhythmia index was significant reduced in TRPC3KO mice ( n = 8) compared to WT mice ( n = 11, P = 0.004).
    Figure Legend Snippet: Atrial arrhythmias induced by Ang II and pacing . Reconstructed excitation maps of data from multi-electrode array recordings (images below) from a Langendorff-perfused isolated heart. (A) sinus rhythm and (B) pacing induced atrial tachycardia. Note in AT, the ectopic beat starts from a different site to sinus rhythms. (C) Representative ECG recordings from isolated heart from WT and TRPC3 −/− mice under control conditions. (D) Atrial tachycardia and atrial fibrillation induced by burst pacing in WT mice and atrial-ventricular conduction block recorded from a TRPC −/− mouse heart respectively. (E) On average, the arrhythmia index was significant reduced in TRPC3KO mice ( n = 8) compared to WT mice ( n = 11, P = 0.004).

    Techniques Used: Isolation, Mouse Assay, Blocking Assay

    Isolated single SAN cells express TRPC3, STIM1, and IP 3 R2 . Confocal immunofluorenscence images of isolated single pacemaker cells. Isolated group (A) and single (B) pacemaker cells were positively stained with anti HCN4 (in green). (C) Anti-TRPC3 in red, anti-STIM1 in green. (D) Anti-IP3R2 in red, anti-STIM1 in green. (E) 3D reconstruction image using N-SIM microscopy. Anti-IP 3 R2 in red, anti-STIM1 in green. Areas of co-localisation appear yellow due to color mixing.
    Figure Legend Snippet: Isolated single SAN cells express TRPC3, STIM1, and IP 3 R2 . Confocal immunofluorenscence images of isolated single pacemaker cells. Isolated group (A) and single (B) pacemaker cells were positively stained with anti HCN4 (in green). (C) Anti-TRPC3 in red, anti-STIM1 in green. (D) Anti-IP3R2 in red, anti-STIM1 in green. (E) 3D reconstruction image using N-SIM microscopy. Anti-IP 3 R2 in red, anti-STIM1 in green. Areas of co-localisation appear yellow due to color mixing.

    Techniques Used: Isolation, Staining, Microscopy

    The effect of Pyr10 on intracellular Ca 2+ after application of 1-oleoy1-2-acyl-sn-glycerol (OAG) 100 μM . The intact SANs were loaded with Ca 2+ indicator indo-1. (A,B) show intracellular Ca 2+ recordings from a WT mouse. (B,C) The statistics pool data shows percentage changes in resting Ca 2+ , Ca 2+ transient and firing rate in WT and TRPC3 KO mice respectively. (B) OAG treatment against control. (C) 2 μM Pyr10 treatment against OAG treatment. * P
    Figure Legend Snippet: The effect of Pyr10 on intracellular Ca 2+ after application of 1-oleoy1-2-acyl-sn-glycerol (OAG) 100 μM . The intact SANs were loaded with Ca 2+ indicator indo-1. (A,B) show intracellular Ca 2+ recordings from a WT mouse. (B,C) The statistics pool data shows percentage changes in resting Ca 2+ , Ca 2+ transient and firing rate in WT and TRPC3 KO mice respectively. (B) OAG treatment against control. (C) 2 μM Pyr10 treatment against OAG treatment. * P

    Techniques Used: Mouse Assay

    Signaling pathways involved in the activation of TRPC3 . The G-protein coupled receptor (GPCRs) activates phospholiase C (PLC), resulting in generation of IP3 and diacylglycerol (DAG). IP3 activates its receptor that leads to Ca 2+ release from SR/ER and the depletion of SR/ER Ca 2+ store. The Ca 2+ content change in the store can be sensed by STIM1, the ER Ca 2+ sensor and cause store-operated Ca 2+ entry (SOCE) through TRPC3 channels. An additional or alternate possibility is that DAG can directly activate TRPC3, and produce receptor-operated Ca 2+ entry (ROCE). GPCRs, G-protein coupled receptors; IP3R, inositol 1,4,5-trisphosphate receptors; PLC, Phospholipase C; ROCE, Receptor–operated Ca 2+ entry; SOCE, store-operated Ca 2+ entry; SR/ER, sarco- endo-plasmic reticulum; STIM1, stromal interacting molecule 1; TRPC3,4,6 canonical transient receptor potential channel types 3,4,6; NCX Na/Ca exchange.
    Figure Legend Snippet: Signaling pathways involved in the activation of TRPC3 . The G-protein coupled receptor (GPCRs) activates phospholiase C (PLC), resulting in generation of IP3 and diacylglycerol (DAG). IP3 activates its receptor that leads to Ca 2+ release from SR/ER and the depletion of SR/ER Ca 2+ store. The Ca 2+ content change in the store can be sensed by STIM1, the ER Ca 2+ sensor and cause store-operated Ca 2+ entry (SOCE) through TRPC3 channels. An additional or alternate possibility is that DAG can directly activate TRPC3, and produce receptor-operated Ca 2+ entry (ROCE). GPCRs, G-protein coupled receptors; IP3R, inositol 1,4,5-trisphosphate receptors; PLC, Phospholipase C; ROCE, Receptor–operated Ca 2+ entry; SOCE, store-operated Ca 2+ entry; SR/ER, sarco- endo-plasmic reticulum; STIM1, stromal interacting molecule 1; TRPC3,4,6 canonical transient receptor potential channel types 3,4,6; NCX Na/Ca exchange.

    Techniques Used: Activation Assay, Planar Chromatography

    17) Product Images from "The involvement of TRPC3 channels in sinoatrial arrhythmias"

    Article Title: The involvement of TRPC3 channels in sinoatrial arrhythmias

    Journal: Frontiers in Physiology

    doi: 10.3389/fphys.2015.00086

    The effect of angiotension II (Ang II) and a selective TRPC3 channel blocker Pyr10 on pacemaker action potential after application of Ang II. (A–D) Intracellular recordings from WT mice; (E–H ) Intracellular recordings from TRPC3 −/− mice with conditions as indicated in each panel.
    Figure Legend Snippet: The effect of angiotension II (Ang II) and a selective TRPC3 channel blocker Pyr10 on pacemaker action potential after application of Ang II. (A–D) Intracellular recordings from WT mice; (E–H ) Intracellular recordings from TRPC3 −/− mice with conditions as indicated in each panel.

    Techniques Used: Mouse Assay

    Atrial arrhythmias induced by Ang II and pacing . Reconstructed excitation maps of data from multi-electrode array recordings (images below) from a Langendorff-perfused isolated heart. (A) sinus rhythm and (B) pacing induced atrial tachycardia. Note in AT, the ectopic beat starts from a different site to sinus rhythms. (C) Representative ECG recordings from isolated heart from WT and TRPC3 −/− mice under control conditions. (D) Atrial tachycardia and atrial fibrillation induced by burst pacing in WT mice and atrial-ventricular conduction block recorded from a TRPC −/− mouse heart respectively. (E) On average, the arrhythmia index was significant reduced in TRPC3KO mice ( n = 8) compared to WT mice ( n = 11, P = 0.004).
    Figure Legend Snippet: Atrial arrhythmias induced by Ang II and pacing . Reconstructed excitation maps of data from multi-electrode array recordings (images below) from a Langendorff-perfused isolated heart. (A) sinus rhythm and (B) pacing induced atrial tachycardia. Note in AT, the ectopic beat starts from a different site to sinus rhythms. (C) Representative ECG recordings from isolated heart from WT and TRPC3 −/− mice under control conditions. (D) Atrial tachycardia and atrial fibrillation induced by burst pacing in WT mice and atrial-ventricular conduction block recorded from a TRPC −/− mouse heart respectively. (E) On average, the arrhythmia index was significant reduced in TRPC3KO mice ( n = 8) compared to WT mice ( n = 11, P = 0.004).

    Techniques Used: Isolation, Mouse Assay, Blocking Assay

    Isolated single SAN cells express TRPC3, STIM1, and IP 3 R2 . Confocal immunofluorenscence images of isolated single pacemaker cells. Isolated group (A) and single (B) pacemaker cells were positively stained with anti HCN4 (in green). (C) Anti-TRPC3 in red, anti-STIM1 in green. (D) Anti-IP3R2 in red, anti-STIM1 in green. (E) 3D reconstruction image using N-SIM microscopy. Anti-IP 3 R2 in red, anti-STIM1 in green. Areas of co-localisation appear yellow due to color mixing.
    Figure Legend Snippet: Isolated single SAN cells express TRPC3, STIM1, and IP 3 R2 . Confocal immunofluorenscence images of isolated single pacemaker cells. Isolated group (A) and single (B) pacemaker cells were positively stained with anti HCN4 (in green). (C) Anti-TRPC3 in red, anti-STIM1 in green. (D) Anti-IP3R2 in red, anti-STIM1 in green. (E) 3D reconstruction image using N-SIM microscopy. Anti-IP 3 R2 in red, anti-STIM1 in green. Areas of co-localisation appear yellow due to color mixing.

    Techniques Used: Isolation, Staining, Microscopy

    The effect of Pyr10 on intracellular Ca 2+ after application of 1-oleoy1-2-acyl-sn-glycerol (OAG) 100 μM . The intact SANs were loaded with Ca 2+ indicator indo-1. (A,B) show intracellular Ca 2+ recordings from a WT mouse. (B,C) The statistics pool data shows percentage changes in resting Ca 2+ , Ca 2+ transient and firing rate in WT and TRPC3 KO mice respectively. (B) OAG treatment against control. (C) 2 μM Pyr10 treatment against OAG treatment. * P
    Figure Legend Snippet: The effect of Pyr10 on intracellular Ca 2+ after application of 1-oleoy1-2-acyl-sn-glycerol (OAG) 100 μM . The intact SANs were loaded with Ca 2+ indicator indo-1. (A,B) show intracellular Ca 2+ recordings from a WT mouse. (B,C) The statistics pool data shows percentage changes in resting Ca 2+ , Ca 2+ transient and firing rate in WT and TRPC3 KO mice respectively. (B) OAG treatment against control. (C) 2 μM Pyr10 treatment against OAG treatment. * P

    Techniques Used: Mouse Assay

    Signaling pathways involved in the activation of TRPC3 . The G-protein coupled receptor (GPCRs) activates phospholiase C (PLC), resulting in generation of IP3 and diacylglycerol (DAG). IP3 activates its receptor that leads to Ca 2+ release from SR/ER and the depletion of SR/ER Ca 2+ store. The Ca 2+ content change in the store can be sensed by STIM1, the ER Ca 2+ sensor and cause store-operated Ca 2+ entry (SOCE) through TRPC3 channels. An additional or alternate possibility is that DAG can directly activate TRPC3, and produce receptor-operated Ca 2+ entry (ROCE). GPCRs, G-protein coupled receptors; IP3R, inositol 1,4,5-trisphosphate receptors; PLC, Phospholipase C; ROCE, Receptor–operated Ca 2+ entry; SOCE, store-operated Ca 2+ entry; SR/ER, sarco- endo-plasmic reticulum; STIM1, stromal interacting molecule 1; TRPC3,4,6 canonical transient receptor potential channel types 3,4,6; NCX Na/Ca exchange.
    Figure Legend Snippet: Signaling pathways involved in the activation of TRPC3 . The G-protein coupled receptor (GPCRs) activates phospholiase C (PLC), resulting in generation of IP3 and diacylglycerol (DAG). IP3 activates its receptor that leads to Ca 2+ release from SR/ER and the depletion of SR/ER Ca 2+ store. The Ca 2+ content change in the store can be sensed by STIM1, the ER Ca 2+ sensor and cause store-operated Ca 2+ entry (SOCE) through TRPC3 channels. An additional or alternate possibility is that DAG can directly activate TRPC3, and produce receptor-operated Ca 2+ entry (ROCE). GPCRs, G-protein coupled receptors; IP3R, inositol 1,4,5-trisphosphate receptors; PLC, Phospholipase C; ROCE, Receptor–operated Ca 2+ entry; SOCE, store-operated Ca 2+ entry; SR/ER, sarco- endo-plasmic reticulum; STIM1, stromal interacting molecule 1; TRPC3,4,6 canonical transient receptor potential channel types 3,4,6; NCX Na/Ca exchange.

    Techniques Used: Activation Assay, Planar Chromatography

    18) Product Images from "TRPC3 positively regulates reactive oxygen species driving maladaptive cardiac remodeling"

    Article Title: TRPC3 positively regulates reactive oxygen species driving maladaptive cardiac remodeling

    Journal: Scientific Reports

    doi: 10.1038/srep37001

    TRPC3 forms a stable ternary complex with Nox2 and p22 phox . ( a , b ) Expression of Nox2 and p22 phox proteins in HEK293 cells that express a different combination of TRPC3-GFP and GFP. Results of a quantitative analysis are shown in ( b ) (n = 3). ( c ) Nox2 mRNA amounts in HEK293 cells co-expressing Nox2 with GFP or TRPC3-GFP (n = 3). ( d ) Increased Nox2 and p22 phox protein in HEK293 cells co-expressing pore-dead mutant of TRPC3 (n = 3). ( e ) Interaction of TRPC3 with Nox2 in HEK293 cells. Immunoprecipitation was performed using an anti-flag antibody. ( f ) Nox2 protein expression in HEK293 cells expressing TRPC3 alone or co-expressing TRPC3 and TRPC6 (n = 3). Error bars, s.e.m. *P
    Figure Legend Snippet: TRPC3 forms a stable ternary complex with Nox2 and p22 phox . ( a , b ) Expression of Nox2 and p22 phox proteins in HEK293 cells that express a different combination of TRPC3-GFP and GFP. Results of a quantitative analysis are shown in ( b ) (n = 3). ( c ) Nox2 mRNA amounts in HEK293 cells co-expressing Nox2 with GFP or TRPC3-GFP (n = 3). ( d ) Increased Nox2 and p22 phox protein in HEK293 cells co-expressing pore-dead mutant of TRPC3 (n = 3). ( e ) Interaction of TRPC3 with Nox2 in HEK293 cells. Immunoprecipitation was performed using an anti-flag antibody. ( f ) Nox2 protein expression in HEK293 cells expressing TRPC3 alone or co-expressing TRPC3 and TRPC6 (n = 3). Error bars, s.e.m. *P

    Techniques Used: Expressing, Mutagenesis, Immunoprecipitation

    TRPC3 plays a critical role in Mechanical stretch-induced ROS production. ( a , b ) Effects of siRNA targeting TRPC1, C3 or C6 on mechanical stretch (MS)-induced ROS production (n = 3). ( c ) mRNA expression of either TRPC1 or TRPC6 in NRCM transfected with siRNAs against either TRPC1 or TRPC6, respectively (n = 3). ( d,e ) Time courses of MS-induced ROS production in NRCMs treated with GsMTx-4 (1 μM; ( d ) or TRPV4 inhibitor (RN1734, 50 μM; ( e ) Reagents were added to cells 5 min before MS (n = 3). ( f ) MS-induced ROS production in TRPC(1–7)-deficient MEF cells expressing TRPC3, TRPC6, TRPC7, or LacZ (n = 30). Data are representative of three independent experiments. ( g–i ) Effect of TRPC3 siRNA on the protein abundances of TRPC3 ( h ) and Nox2 ( i ) protein expressions in NRCMs (n = 3). Error bars, s.e.m. *P
    Figure Legend Snippet: TRPC3 plays a critical role in Mechanical stretch-induced ROS production. ( a , b ) Effects of siRNA targeting TRPC1, C3 or C6 on mechanical stretch (MS)-induced ROS production (n = 3). ( c ) mRNA expression of either TRPC1 or TRPC6 in NRCM transfected with siRNAs against either TRPC1 or TRPC6, respectively (n = 3). ( d,e ) Time courses of MS-induced ROS production in NRCMs treated with GsMTx-4 (1 μM; ( d ) or TRPV4 inhibitor (RN1734, 50 μM; ( e ) Reagents were added to cells 5 min before MS (n = 3). ( f ) MS-induced ROS production in TRPC(1–7)-deficient MEF cells expressing TRPC3, TRPC6, TRPC7, or LacZ (n = 30). Data are representative of three independent experiments. ( g–i ) Effect of TRPC3 siRNA on the protein abundances of TRPC3 ( h ) and Nox2 ( i ) protein expressions in NRCMs (n = 3). Error bars, s.e.m. *P

    Techniques Used: Mass Spectrometry, Expressing, Transfection

    TRPC3 prevents Nox2 protein from proteasomal degradation. ( a–e ) Abundances of Nox2 protein ( a , b ) and mRNAs of TRPC3 ( c ), Nox2 ( d ), and p22 phox ( e ) in NRCM transfected with siRNAs targeting TRPC3 with or without MG132. Cells were treated with siRNAs and MG132 (1 μM) simultaneously (n = 3). ( f , g ) Effect of siRNA targeting TRPC3 on Nox2 protein abundance in cell surface (Surface) and total lysates (Total) from NRCMs (n = 3). GAPDH was used as an internal control. Error bars, s.e.m. *P
    Figure Legend Snippet: TRPC3 prevents Nox2 protein from proteasomal degradation. ( a–e ) Abundances of Nox2 protein ( a , b ) and mRNAs of TRPC3 ( c ), Nox2 ( d ), and p22 phox ( e ) in NRCM transfected with siRNAs targeting TRPC3 with or without MG132. Cells were treated with siRNAs and MG132 (1 μM) simultaneously (n = 3). ( f , g ) Effect of siRNA targeting TRPC3 on Nox2 protein abundance in cell surface (Surface) and total lysates (Total) from NRCMs (n = 3). GAPDH was used as an internal control. Error bars, s.e.m. *P

    Techniques Used: Transfection

    TRPC3 forms a stable ternary complex with Nox2 and p22 phox proteins in endogenously p22 phox -absent CHO cells. ( a ) Expression of Nox2 and p22 phox proteins in CHO cells that express a different combination of TRPC3-GFP and GFP. ( b ) Results of quantitative analysis (n = 3). ( c ) Expression of Nox2 and p22 phox co-expressed with either GFP or TRPC3-GFP in MG132 (10 μM)-treated CHO cells. ( d ) Graphs depict the relative expression of either Nox2 or p22 phox protein to that in non-treated cells. Band intensities were normalized by GAPDH. ( e–g ) Interaction of TRPC3 with p22 phox and Nox2 in CHO cells. ( h ) Localization of Nox2 in CHO cells co-expressing Nox2 with TRPC3-GFP (or GFP-F). Error bars, s.e.m. *P
    Figure Legend Snippet: TRPC3 forms a stable ternary complex with Nox2 and p22 phox proteins in endogenously p22 phox -absent CHO cells. ( a ) Expression of Nox2 and p22 phox proteins in CHO cells that express a different combination of TRPC3-GFP and GFP. ( b ) Results of quantitative analysis (n = 3). ( c ) Expression of Nox2 and p22 phox co-expressed with either GFP or TRPC3-GFP in MG132 (10 μM)-treated CHO cells. ( d ) Graphs depict the relative expression of either Nox2 or p22 phox protein to that in non-treated cells. Band intensities were normalized by GAPDH. ( e–g ) Interaction of TRPC3 with p22 phox and Nox2 in CHO cells. ( h ) Localization of Nox2 in CHO cells co-expressing Nox2 with TRPC3-GFP (or GFP-F). Error bars, s.e.m. *P

    Techniques Used: Expressing

    Formation of a TRPC3/Nox2 complex promotes TRPC3 channel activity through stabilization at the plasma membrane. ( a ) Effect of Nox2 siRNA on expression of TRPC3 in NRCMs (n = 3). ( b ) Representative images showing the levels of TRPC3-GFP and GFP expression in HEK293 cells co-expressing p22 phox or Nox2 (n = 3). ( c ) Expression of TRPC3-GFP mRNA in HEK293 cells co-expressing p22 phox or Nox2 (n = 3). ( d–f ) Representative time courses of TRPC3 currents ( d ) and the current-voltage (I-V) relationships ( e ) and peak TRPC3 current densities at −60 mV ( f ) induced by 100 μM carbachol (CCh) in HEK293 cells expressing TRPC3-mCherry alone or with p22 phox , Nox2, both p22 phox and Nox2, or Nox2 treated with DPI. DPI (0.3 μM) was treated 1 min before CCh stimulation. ( g , h ) Representative Ca 2+ responses in the presence ( g ) or absence ( h ) of pyrazole-3 (Pyr3, 1 μM) upon mechanical stretch (MS) application. ( i ) Peak Ca 2+ increases after MS in NRCMs treated with (n = 61) or without Pyr3 (n = 78). ( j ) Changes of minimal [Ca 2+ ] i before and after MS application. Minimal [Ca 2+ ] i from Ca 2+ responses in every 1 min were analyzed and represented as diastolic [Ca 2+ ] i . ( k ) Schematic images showing phosphorylation of p47 phox via TRPC3-PKCβ activation induced by MS in the heart. ( l–n ) Effects of TRPC3 ( l , m ) or PKCβ ( n) ; 10 μM Gö6976) inhibitors on p47 phox phosphorylation induced by MS in NRCMs (n = 3). ( o ) MS-induced ROS generation in NRCMs treated with a PKCβ inhibitor (n = 3). ( p ) Co-immunoprecipitation of TRPC3 with PKCβ, Nox2 and p22 phox in mouse hearts 1week after TAC operation (n = 3). Error bars, s.e.m.*P
    Figure Legend Snippet: Formation of a TRPC3/Nox2 complex promotes TRPC3 channel activity through stabilization at the plasma membrane. ( a ) Effect of Nox2 siRNA on expression of TRPC3 in NRCMs (n = 3). ( b ) Representative images showing the levels of TRPC3-GFP and GFP expression in HEK293 cells co-expressing p22 phox or Nox2 (n = 3). ( c ) Expression of TRPC3-GFP mRNA in HEK293 cells co-expressing p22 phox or Nox2 (n = 3). ( d–f ) Representative time courses of TRPC3 currents ( d ) and the current-voltage (I-V) relationships ( e ) and peak TRPC3 current densities at −60 mV ( f ) induced by 100 μM carbachol (CCh) in HEK293 cells expressing TRPC3-mCherry alone or with p22 phox , Nox2, both p22 phox and Nox2, or Nox2 treated with DPI. DPI (0.3 μM) was treated 1 min before CCh stimulation. ( g , h ) Representative Ca 2+ responses in the presence ( g ) or absence ( h ) of pyrazole-3 (Pyr3, 1 μM) upon mechanical stretch (MS) application. ( i ) Peak Ca 2+ increases after MS in NRCMs treated with (n = 61) or without Pyr3 (n = 78). ( j ) Changes of minimal [Ca 2+ ] i before and after MS application. Minimal [Ca 2+ ] i from Ca 2+ responses in every 1 min were analyzed and represented as diastolic [Ca 2+ ] i . ( k ) Schematic images showing phosphorylation of p47 phox via TRPC3-PKCβ activation induced by MS in the heart. ( l–n ) Effects of TRPC3 ( l , m ) or PKCβ ( n) ; 10 μM Gö6976) inhibitors on p47 phox phosphorylation induced by MS in NRCMs (n = 3). ( o ) MS-induced ROS generation in NRCMs treated with a PKCβ inhibitor (n = 3). ( p ) Co-immunoprecipitation of TRPC3 with PKCβ, Nox2 and p22 phox in mouse hearts 1week after TAC operation (n = 3). Error bars, s.e.m.*P

    Techniques Used: Activity Assay, Expressing, Mass Spectrometry, Activation Assay, Immunoprecipitation

    TRPC3 deletion suppresses TAC-induced LV dysfunction and dilation through Nox2 inhibition. ( a ) Left ventricular end-diastolic pressure (LVEDP; left) and dP/dT max (right) in TAC-operated TRPC3 (+/+) (n = 13) and TRPC3 (−/−) (n = 12) mice 6 week post-operation. ( b ) Myocardial malondialdehyde concentrations 1 week after TAC (n = 4). ( c ) Abundance of Nox2 protein in TRPC3 (+/+) and TRPC3 (−/−) hearts 1 week after TAC (n = 3). ( d ) Representative immunofluorescence images of TRPC3, p22 phox , and caveolin-3 (Cav-3) in adult mouse cardiomyocytes isolated from muscle LIM protein-deficient hearts. ( e ) Representative immunofluorescence images of p22 phox in adult mouse cardiomyocytes: green, anti-p22 phox ; blue, DAPI. ( f ) Relative abundances of p22 phox and Nox2 mRNA in mouse hearts 1 week after TAC (n = 4). ( g ) Abundance of Nox2 protein in TRPC6 (+/+) and TRPC6 (−/−) hearts 1 week after TAC (n = 3). Error bars, s.e.m. *P
    Figure Legend Snippet: TRPC3 deletion suppresses TAC-induced LV dysfunction and dilation through Nox2 inhibition. ( a ) Left ventricular end-diastolic pressure (LVEDP; left) and dP/dT max (right) in TAC-operated TRPC3 (+/+) (n = 13) and TRPC3 (−/−) (n = 12) mice 6 week post-operation. ( b ) Myocardial malondialdehyde concentrations 1 week after TAC (n = 4). ( c ) Abundance of Nox2 protein in TRPC3 (+/+) and TRPC3 (−/−) hearts 1 week after TAC (n = 3). ( d ) Representative immunofluorescence images of TRPC3, p22 phox , and caveolin-3 (Cav-3) in adult mouse cardiomyocytes isolated from muscle LIM protein-deficient hearts. ( e ) Representative immunofluorescence images of p22 phox in adult mouse cardiomyocytes: green, anti-p22 phox ; blue, DAPI. ( f ) Relative abundances of p22 phox and Nox2 mRNA in mouse hearts 1 week after TAC (n = 4). ( g ) Abundance of Nox2 protein in TRPC6 (+/+) and TRPC6 (−/−) hearts 1 week after TAC (n = 3). Error bars, s.e.m. *P

    Techniques Used: Inhibition, Mouse Assay, Immunofluorescence, Isolation

    Physical interaction between TRPC3 and Nox2 is critical for stabilization of Nox2. ( a ) Schematic illustration of TRPC3 terminal deletion mutants. ( b , c ) Expression of Nox2 and p22 phox co-expressed with TRPC3 deletion mutants in HEK293 cells (n = 3). ( d ) OAG-induced ROS production in NRCMs expressing Nox2-interacting TRPC3 C-terminal fragment (C3-C fragment) (n = 20–28). ( e ) Co-immunoprecipitation of TRPC3 with Nox2 in the presence or absence of C3-C fragment. Representative blot from three independent experiments was shown. ( f ) ATP (100 μM)-induced Ca 2+ responses in HEK293 cells expressing TRPC3 with or without C3-C fragment (n = 35–51). Timing of solution exchanges were indicated by horizontal bars above the graph. ( g ) Model of the regulation of TRPC3-Nox2 stability and induction of LV dysfunction induced by diastolic stretch of cardiomyocytes. Error bars, s.e.m.*P
    Figure Legend Snippet: Physical interaction between TRPC3 and Nox2 is critical for stabilization of Nox2. ( a ) Schematic illustration of TRPC3 terminal deletion mutants. ( b , c ) Expression of Nox2 and p22 phox co-expressed with TRPC3 deletion mutants in HEK293 cells (n = 3). ( d ) OAG-induced ROS production in NRCMs expressing Nox2-interacting TRPC3 C-terminal fragment (C3-C fragment) (n = 20–28). ( e ) Co-immunoprecipitation of TRPC3 with Nox2 in the presence or absence of C3-C fragment. Representative blot from three independent experiments was shown. ( f ) ATP (100 μM)-induced Ca 2+ responses in HEK293 cells expressing TRPC3 with or without C3-C fragment (n = 35–51). Timing of solution exchanges were indicated by horizontal bars above the graph. ( g ) Model of the regulation of TRPC3-Nox2 stability and induction of LV dysfunction induced by diastolic stretch of cardiomyocytes. Error bars, s.e.m.*P

    Techniques Used: Expressing, Immunoprecipitation

    19) Product Images from "Hypertrophic scar contracture is mediated by the TRPC3 mechanical force transducer via NFkB activation"

    Article Title: Hypertrophic scar contracture is mediated by the TRPC3 mechanical force transducer via NFkB activation

    Journal: Scientific Reports

    doi: 10.1038/srep11620

    Fibroblast Populated Collagen Lattice with TRPC3 overexpressing fibroblasts appeared to have more contractile activity than a collagen lattice with control fibroblasts. The contractile activities of TRPC3 overexpressing fibroblasts were analyzed using a gel contraction assay. Each gel contained either 8 × 10 5 TRPC3 overexpressing cells or empty vector transfected control cells and was incubated for 24 hours. After starvation, the gels were treated with either a TRPC3 agonist (OAG) or a TRPC3 inhibitor (Pyr3) and the contraction ratio was observed. Gels with TRPC3 overexpressing cells exhibited more contraction than control gels. This contraction effect was enhanced with OAG and attenuated by Pyr3. The data is expressed as the mean ± SD (n = 4). * p
    Figure Legend Snippet: Fibroblast Populated Collagen Lattice with TRPC3 overexpressing fibroblasts appeared to have more contractile activity than a collagen lattice with control fibroblasts. The contractile activities of TRPC3 overexpressing fibroblasts were analyzed using a gel contraction assay. Each gel contained either 8 × 10 5 TRPC3 overexpressing cells or empty vector transfected control cells and was incubated for 24 hours. After starvation, the gels were treated with either a TRPC3 agonist (OAG) or a TRPC3 inhibitor (Pyr3) and the contraction ratio was observed. Gels with TRPC3 overexpressing cells exhibited more contraction than control gels. This contraction effect was enhanced with OAG and attenuated by Pyr3. The data is expressed as the mean ± SD (n = 4). * p

    Techniques Used: Activity Assay, Collagen Gel Contraction Assay, Plasmid Preparation, Transfection, Incubation

    Proposed model for the role of TRPC3 in hypertrophic scar contracture. TRPC3 gene transcription is upregulated by mechanical stretching forces. Increased TRPC3 expression leads to increased calcium influx, which induces NFκB phosphorylation. Activated NFκB translocates into the nucleus and promotes Fibronectin gene expression, which ultimately enhances wound contraction.
    Figure Legend Snippet: Proposed model for the role of TRPC3 in hypertrophic scar contracture. TRPC3 gene transcription is upregulated by mechanical stretching forces. Increased TRPC3 expression leads to increased calcium influx, which induces NFκB phosphorylation. Activated NFκB translocates into the nucleus and promotes Fibronectin gene expression, which ultimately enhances wound contraction.

    Techniques Used: Expressing

    Expression of TRPC3 channel in human hypertrophic scar tissue. In order to investigate the expression levels of TRPC3 in human hypertrophic scar tissue, 3 surgical specimens were obtained from scar revision surgery and underwent qRT PCR and immunohistochemical analysis as described in the Materials and Methods. a ; qRT-PCR showed human hypertrophic scar tissue had more TRPC3 expression compared to normal skin. Values were standardized against cyclophilin. Data represents means ± SD of 3 samples. * p
    Figure Legend Snippet: Expression of TRPC3 channel in human hypertrophic scar tissue. In order to investigate the expression levels of TRPC3 in human hypertrophic scar tissue, 3 surgical specimens were obtained from scar revision surgery and underwent qRT PCR and immunohistochemical analysis as described in the Materials and Methods. a ; qRT-PCR showed human hypertrophic scar tissue had more TRPC3 expression compared to normal skin. Values were standardized against cyclophilin. Data represents means ± SD of 3 samples. * p

    Techniques Used: Expressing, Quantitative RT-PCR, Immunohistochemistry

    TRPC3 mediates cyclic stretching forces via NFκB activation in fibroblasts. To elucidate the mechanism by which Fibronectin expression is increased in TRPC3 overexpressing cells in response to mechanical stretching, the activity of transcriptional factor NFκB was assessed. To clarify the data, the blots were cropped. Uncropped, full-length blots for Western Blot and EMSA are presented in Supplementary Figures S11–20 . a ; In order to determine whether NFκB is involved in the regulation of Fibronectin production, TRPC3 overexpressing cells and control cells were stretched (20%, 10 Hz) with/without an NFκB inhibitor (Wedelolactone) for 24 hours. Western blot analysis demonstrated that the increased expression of Fibronectin in TRPC3 overexpressing cells in response to mechanical stretching was blocked by Wedelolactone. b ; The phosphorylation of NFκB in TRPC3 overexpressing cells was upregulated in a time dependent manner compared to control cells c ; The phosphorylation of NFκB in response to stretching stimuli was attenuated by the TRPC3 specific inhibitor, Pyr3. d ; The phosphorylation pattern of IκB, which is a regulator of NFκB activation, was assessed. TPC3 overexpressing fibroblasts showed more activation of IκB in response to mechanical stretching compared to control fibroblasts. The activation of IκB was attenuated by Pyr3 in dose dependent manner. e ; Translocation of phosphorylated NFκB was evident in TRPC3 overexpressing fibroblasts stretched for 15 minutes. Arrows indicate translocated phosphorylated NFκB. Scale bars for each images = 50 μm. f ; TRPC3 overexpressing fibroblasts and control fibroblasts were stretched for 15 minutes and the nuclear proteins were extracted. A gel shift assay was then performed, as described in the Materials and Methods section. Gel shift was most apparent in stretched TRPC3 overexpressing fibroblasts, which was attenuated by the addition of Pyr3.
    Figure Legend Snippet: TRPC3 mediates cyclic stretching forces via NFκB activation in fibroblasts. To elucidate the mechanism by which Fibronectin expression is increased in TRPC3 overexpressing cells in response to mechanical stretching, the activity of transcriptional factor NFκB was assessed. To clarify the data, the blots were cropped. Uncropped, full-length blots for Western Blot and EMSA are presented in Supplementary Figures S11–20 . a ; In order to determine whether NFκB is involved in the regulation of Fibronectin production, TRPC3 overexpressing cells and control cells were stretched (20%, 10 Hz) with/without an NFκB inhibitor (Wedelolactone) for 24 hours. Western blot analysis demonstrated that the increased expression of Fibronectin in TRPC3 overexpressing cells in response to mechanical stretching was blocked by Wedelolactone. b ; The phosphorylation of NFκB in TRPC3 overexpressing cells was upregulated in a time dependent manner compared to control cells c ; The phosphorylation of NFκB in response to stretching stimuli was attenuated by the TRPC3 specific inhibitor, Pyr3. d ; The phosphorylation pattern of IκB, which is a regulator of NFκB activation, was assessed. TPC3 overexpressing fibroblasts showed more activation of IκB in response to mechanical stretching compared to control fibroblasts. The activation of IκB was attenuated by Pyr3 in dose dependent manner. e ; Translocation of phosphorylated NFκB was evident in TRPC3 overexpressing fibroblasts stretched for 15 minutes. Arrows indicate translocated phosphorylated NFκB. Scale bars for each images = 50 μm. f ; TRPC3 overexpressing fibroblasts and control fibroblasts were stretched for 15 minutes and the nuclear proteins were extracted. A gel shift assay was then performed, as described in the Materials and Methods section. Gel shift was most apparent in stretched TRPC3 overexpressing fibroblasts, which was attenuated by the addition of Pyr3.

    Techniques Used: Activation Assay, Expressing, Activity Assay, Western Blot, Translocation Assay, Electrophoretic Mobility Shift Assay

    Fibronectin production was increased in TRPC3 overexpressing fibroblasts when they were subjected to repetitive stretching. TRPC3 overexpressing fibroblasts and control cells were stretched for 24 hours and the expression levels of Fibronectin were assessed using qRT PCR and Western Blot analysis. The blots presented here were cropped to improve clarity. Uncropped, full-length blots are presented in Supplementary Figures S6–10 . a ; qRT PCR demonstrated that the gene expression level of Fibronectin was significantly increased after 24 hours of stretching in TRPC3 overexpressing cells compared to control cells. Data represents means ± SD of 3 samples. * p
    Figure Legend Snippet: Fibronectin production was increased in TRPC3 overexpressing fibroblasts when they were subjected to repetitive stretching. TRPC3 overexpressing fibroblasts and control cells were stretched for 24 hours and the expression levels of Fibronectin were assessed using qRT PCR and Western Blot analysis. The blots presented here were cropped to improve clarity. Uncropped, full-length blots are presented in Supplementary Figures S6–10 . a ; qRT PCR demonstrated that the gene expression level of Fibronectin was significantly increased after 24 hours of stretching in TRPC3 overexpressing cells compared to control cells. Data represents means ± SD of 3 samples. * p

    Techniques Used: Expressing, Quantitative RT-PCR, Western Blot

    Real-time calcium imaging of TRPC3 overexpressing fibroblasts and control fibroblasts during mechanical stretching. Live imaging of calcium influx in fibroblasts was obtained as described in the Materials and Methods section. Scale bars for each images = 50 μm. a ; Control fibroblasts showed a weak increase in fluorescence upon repetitive mechanical stretching. b ; TRPC3 overexpressing fibroblasts exhibited increased fluorescence upon repetitive mechanical stretching. The increased fluorescence continued throughout the period in which they were stretched. c ; When TRPC3 overexpressing fibroblasts were stretched once and then held in a stationary position, the fluorescence decreased over time.
    Figure Legend Snippet: Real-time calcium imaging of TRPC3 overexpressing fibroblasts and control fibroblasts during mechanical stretching. Live imaging of calcium influx in fibroblasts was obtained as described in the Materials and Methods section. Scale bars for each images = 50 μm. a ; Control fibroblasts showed a weak increase in fluorescence upon repetitive mechanical stretching. b ; TRPC3 overexpressing fibroblasts exhibited increased fluorescence upon repetitive mechanical stretching. The increased fluorescence continued throughout the period in which they were stretched. c ; When TRPC3 overexpressing fibroblasts were stretched once and then held in a stationary position, the fluorescence decreased over time.

    Techniques Used: Imaging, Fluorescence

    Wound contraction was enhanced by the transplantation of TRPC3 overexpressing fibroblasts in mice. To assess the effect of TRPC3 overexpressing fibroblasts on wound healing in vivo , TRPC3 overexpressing fibroblasts, empty-vector transfected control fibroblasts or the same amount of saline were injected into the dorsum of nude mice. 10 days post-transplantation, 6 mm wounds were created and the size of the wounds were subsequently measured on Days 1, 3, 5, 7 and 9. The wound tissue was ultimately harvested at Day 9 post-wounding and immunochemistry with anti-TRPC3 and Fibronectin was performed. a ; Representative photographs of cutaneous wounds. Differential healing was observed at all time points checked (n = 6 for each time point). b ; Quantification of the wound area from Days 0–9. The data is expressed as the mean ± SD. n = 6 per group. * p
    Figure Legend Snippet: Wound contraction was enhanced by the transplantation of TRPC3 overexpressing fibroblasts in mice. To assess the effect of TRPC3 overexpressing fibroblasts on wound healing in vivo , TRPC3 overexpressing fibroblasts, empty-vector transfected control fibroblasts or the same amount of saline were injected into the dorsum of nude mice. 10 days post-transplantation, 6 mm wounds were created and the size of the wounds were subsequently measured on Days 1, 3, 5, 7 and 9. The wound tissue was ultimately harvested at Day 9 post-wounding and immunochemistry with anti-TRPC3 and Fibronectin was performed. a ; Representative photographs of cutaneous wounds. Differential healing was observed at all time points checked (n = 6 for each time point). b ; Quantification of the wound area from Days 0–9. The data is expressed as the mean ± SD. n = 6 per group. * p

    Techniques Used: Transplantation Assay, Mouse Assay, In Vivo, Plasmid Preparation, Transfection, Injection

    20) Product Images from "Resistance to Store Depletion-induced Endothelial Injury in Rat Lung after Chronic Heart Failure"

    Article Title: Resistance to Store Depletion-induced Endothelial Injury in Rat Lung after Chronic Heart Failure

    Journal:

    doi: 10.1164/rccm.200506-847OC

    Immunostaining for transient receptor potential proteins TRPC1, TRPC3, TRPC4, and TRPC6/7 in rat lungs. Expression of each TRPC isoform in endothelium was confirmed in small muscular or partially muscularized extraalveolar vessels ( A ) and conduit artery
    Figure Legend Snippet: Immunostaining for transient receptor potential proteins TRPC1, TRPC3, TRPC4, and TRPC6/7 in rat lungs. Expression of each TRPC isoform in endothelium was confirmed in small muscular or partially muscularized extraalveolar vessels ( A ) and conduit artery

    Techniques Used: Immunostaining, Expressing

    21) Product Images from "The involvement of TRPC3 channels in sinoatrial arrhythmias"

    Article Title: The involvement of TRPC3 channels in sinoatrial arrhythmias

    Journal: Frontiers in Physiology

    doi: 10.3389/fphys.2015.00086

    The effect of angiotension II (Ang II) and a selective TRPC3 channel blocker Pyr10 on pacemaker action potential after application of Ang II. (A–D) Intracellular recordings from WT mice; (E–H ) Intracellular recordings from TRPC3 −/− mice with conditions as indicated in each panel.
    Figure Legend Snippet: The effect of angiotension II (Ang II) and a selective TRPC3 channel blocker Pyr10 on pacemaker action potential after application of Ang II. (A–D) Intracellular recordings from WT mice; (E–H ) Intracellular recordings from TRPC3 −/− mice with conditions as indicated in each panel.

    Techniques Used: Mouse Assay

    Atrial arrhythmias induced by Ang II and pacing . Reconstructed excitation maps of data from multi-electrode array recordings (images below) from a Langendorff-perfused isolated heart. (A) sinus rhythm and (B) pacing induced atrial tachycardia. Note in AT, the ectopic beat starts from a different site to sinus rhythms. (C) Representative ECG recordings from isolated heart from WT and TRPC3 −/− mice under control conditions. (D) Atrial tachycardia and atrial fibrillation induced by burst pacing in WT mice and atrial-ventricular conduction block recorded from a TRPC −/− mouse heart respectively. (E) On average, the arrhythmia index was significant reduced in TRPC3KO mice ( n = 8) compared to WT mice ( n = 11, P = 0.004).
    Figure Legend Snippet: Atrial arrhythmias induced by Ang II and pacing . Reconstructed excitation maps of data from multi-electrode array recordings (images below) from a Langendorff-perfused isolated heart. (A) sinus rhythm and (B) pacing induced atrial tachycardia. Note in AT, the ectopic beat starts from a different site to sinus rhythms. (C) Representative ECG recordings from isolated heart from WT and TRPC3 −/− mice under control conditions. (D) Atrial tachycardia and atrial fibrillation induced by burst pacing in WT mice and atrial-ventricular conduction block recorded from a TRPC −/− mouse heart respectively. (E) On average, the arrhythmia index was significant reduced in TRPC3KO mice ( n = 8) compared to WT mice ( n = 11, P = 0.004).

    Techniques Used: Isolation, Mouse Assay, Blocking Assay

    Isolated single SAN cells express TRPC3, STIM1, and IP 3 R2 . Confocal immunofluorenscence images of isolated single pacemaker cells. Isolated group (A) and single (B) pacemaker cells were positively stained with anti HCN4 (in green). (C) Anti-TRPC3 in red, anti-STIM1 in green. (D) Anti-IP3R2 in red, anti-STIM1 in green. (E) 3D reconstruction image using N-SIM microscopy. Anti-IP 3 R2 in red, anti-STIM1 in green. Areas of co-localisation appear yellow due to color mixing.
    Figure Legend Snippet: Isolated single SAN cells express TRPC3, STIM1, and IP 3 R2 . Confocal immunofluorenscence images of isolated single pacemaker cells. Isolated group (A) and single (B) pacemaker cells were positively stained with anti HCN4 (in green). (C) Anti-TRPC3 in red, anti-STIM1 in green. (D) Anti-IP3R2 in red, anti-STIM1 in green. (E) 3D reconstruction image using N-SIM microscopy. Anti-IP 3 R2 in red, anti-STIM1 in green. Areas of co-localisation appear yellow due to color mixing.

    Techniques Used: Isolation, Staining, Microscopy

    The effect of Pyr10 on intracellular Ca 2+ after application of 1-oleoy1-2-acyl-sn-glycerol (OAG) 100 μM . The intact SANs were loaded with Ca 2+ indicator indo-1. (A,B) show intracellular Ca 2+ recordings from a WT mouse. (B,C) The statistics pool data shows percentage changes in resting Ca 2+ , Ca 2+ transient and firing rate in WT and TRPC3 KO mice respectively. (B) OAG treatment against control. (C) 2 μM Pyr10 treatment against OAG treatment. * P
    Figure Legend Snippet: The effect of Pyr10 on intracellular Ca 2+ after application of 1-oleoy1-2-acyl-sn-glycerol (OAG) 100 μM . The intact SANs were loaded with Ca 2+ indicator indo-1. (A,B) show intracellular Ca 2+ recordings from a WT mouse. (B,C) The statistics pool data shows percentage changes in resting Ca 2+ , Ca 2+ transient and firing rate in WT and TRPC3 KO mice respectively. (B) OAG treatment against control. (C) 2 μM Pyr10 treatment against OAG treatment. * P

    Techniques Used: Mouse Assay

    Signaling pathways involved in the activation of TRPC3 . The G-protein coupled receptor (GPCRs) activates phospholiase C (PLC), resulting in generation of IP3 and diacylglycerol (DAG). IP3 activates its receptor that leads to Ca 2+ release from SR/ER and the depletion of SR/ER Ca 2+ store. The Ca 2+ content change in the store can be sensed by STIM1, the ER Ca 2+ sensor and cause store-operated Ca 2+ entry (SOCE) through TRPC3 channels. An additional or alternate possibility is that DAG can directly activate TRPC3, and produce receptor-operated Ca 2+ entry (ROCE). GPCRs, G-protein coupled receptors; IP3R, inositol 1,4,5-trisphosphate receptors; PLC, Phospholipase C; ROCE, Receptor–operated Ca 2+ entry; SOCE, store-operated Ca 2+ entry; SR/ER, sarco- endo-plasmic reticulum; STIM1, stromal interacting molecule 1; TRPC3,4,6 canonical transient receptor potential channel types 3,4,6; NCX Na/Ca exchange.
    Figure Legend Snippet: Signaling pathways involved in the activation of TRPC3 . The G-protein coupled receptor (GPCRs) activates phospholiase C (PLC), resulting in generation of IP3 and diacylglycerol (DAG). IP3 activates its receptor that leads to Ca 2+ release from SR/ER and the depletion of SR/ER Ca 2+ store. The Ca 2+ content change in the store can be sensed by STIM1, the ER Ca 2+ sensor and cause store-operated Ca 2+ entry (SOCE) through TRPC3 channels. An additional or alternate possibility is that DAG can directly activate TRPC3, and produce receptor-operated Ca 2+ entry (ROCE). GPCRs, G-protein coupled receptors; IP3R, inositol 1,4,5-trisphosphate receptors; PLC, Phospholipase C; ROCE, Receptor–operated Ca 2+ entry; SOCE, store-operated Ca 2+ entry; SR/ER, sarco- endo-plasmic reticulum; STIM1, stromal interacting molecule 1; TRPC3,4,6 canonical transient receptor potential channel types 3,4,6; NCX Na/Ca exchange.

    Techniques Used: Activation Assay, Planar Chromatography

    22) Product Images from "The involvement of TRPC3 channels in sinoatrial arrhythmias"

    Article Title: The involvement of TRPC3 channels in sinoatrial arrhythmias

    Journal: Frontiers in Physiology

    doi: 10.3389/fphys.2015.00086

    The effect of angiotension II (Ang II) and a selective TRPC3 channel blocker Pyr10 on pacemaker action potential after application of Ang II. (A–D) Intracellular recordings from WT mice; (E–H ) Intracellular recordings from TRPC3 −/− mice with conditions as indicated in each panel.
    Figure Legend Snippet: The effect of angiotension II (Ang II) and a selective TRPC3 channel blocker Pyr10 on pacemaker action potential after application of Ang II. (A–D) Intracellular recordings from WT mice; (E–H ) Intracellular recordings from TRPC3 −/− mice with conditions as indicated in each panel.

    Techniques Used: Mouse Assay

    Atrial arrhythmias induced by Ang II and pacing . Reconstructed excitation maps of data from multi-electrode array recordings (images below) from a Langendorff-perfused isolated heart. (A) sinus rhythm and (B) pacing induced atrial tachycardia. Note in AT, the ectopic beat starts from a different site to sinus rhythms. (C) Representative ECG recordings from isolated heart from WT and TRPC3 −/− mice under control conditions. (D) Atrial tachycardia and atrial fibrillation induced by burst pacing in WT mice and atrial-ventricular conduction block recorded from a TRPC −/− mouse heart respectively. (E) On average, the arrhythmia index was significant reduced in TRPC3KO mice ( n = 8) compared to WT mice ( n = 11, P = 0.004).
    Figure Legend Snippet: Atrial arrhythmias induced by Ang II and pacing . Reconstructed excitation maps of data from multi-electrode array recordings (images below) from a Langendorff-perfused isolated heart. (A) sinus rhythm and (B) pacing induced atrial tachycardia. Note in AT, the ectopic beat starts from a different site to sinus rhythms. (C) Representative ECG recordings from isolated heart from WT and TRPC3 −/− mice under control conditions. (D) Atrial tachycardia and atrial fibrillation induced by burst pacing in WT mice and atrial-ventricular conduction block recorded from a TRPC −/− mouse heart respectively. (E) On average, the arrhythmia index was significant reduced in TRPC3KO mice ( n = 8) compared to WT mice ( n = 11, P = 0.004).

    Techniques Used: Isolation, Mouse Assay, Blocking Assay

    Isolated single SAN cells express TRPC3, STIM1, and IP 3 R2 . Confocal immunofluorenscence images of isolated single pacemaker cells. Isolated group (A) and single (B) pacemaker cells were positively stained with anti HCN4 (in green). (C) Anti-TRPC3 in red, anti-STIM1 in green. (D) Anti-IP3R2 in red, anti-STIM1 in green. (E) 3D reconstruction image using N-SIM microscopy. Anti-IP 3 R2 in red, anti-STIM1 in green. Areas of co-localisation appear yellow due to color mixing.
    Figure Legend Snippet: Isolated single SAN cells express TRPC3, STIM1, and IP 3 R2 . Confocal immunofluorenscence images of isolated single pacemaker cells. Isolated group (A) and single (B) pacemaker cells were positively stained with anti HCN4 (in green). (C) Anti-TRPC3 in red, anti-STIM1 in green. (D) Anti-IP3R2 in red, anti-STIM1 in green. (E) 3D reconstruction image using N-SIM microscopy. Anti-IP 3 R2 in red, anti-STIM1 in green. Areas of co-localisation appear yellow due to color mixing.

    Techniques Used: Isolation, Staining, Microscopy

    The effect of Pyr10 on intracellular Ca 2+ after application of 1-oleoy1-2-acyl-sn-glycerol (OAG) 100 μM . The intact SANs were loaded with Ca 2+ indicator indo-1. (A,B) show intracellular Ca 2+ recordings from a WT mouse. (B,C) The statistics pool data shows percentage changes in resting Ca 2+ , Ca 2+ transient and firing rate in WT and TRPC3 KO mice respectively. (B) OAG treatment against control. (C) 2 μM Pyr10 treatment against OAG treatment. * P
    Figure Legend Snippet: The effect of Pyr10 on intracellular Ca 2+ after application of 1-oleoy1-2-acyl-sn-glycerol (OAG) 100 μM . The intact SANs were loaded with Ca 2+ indicator indo-1. (A,B) show intracellular Ca 2+ recordings from a WT mouse. (B,C) The statistics pool data shows percentage changes in resting Ca 2+ , Ca 2+ transient and firing rate in WT and TRPC3 KO mice respectively. (B) OAG treatment against control. (C) 2 μM Pyr10 treatment against OAG treatment. * P

    Techniques Used: Mouse Assay

    Signaling pathways involved in the activation of TRPC3 . The G-protein coupled receptor (GPCRs) activates phospholiase C (PLC), resulting in generation of IP3 and diacylglycerol (DAG). IP3 activates its receptor that leads to Ca 2+ release from SR/ER and the depletion of SR/ER Ca 2+ store. The Ca 2+ content change in the store can be sensed by STIM1, the ER Ca 2+ sensor and cause store-operated Ca 2+ entry (SOCE) through TRPC3 channels. An additional or alternate possibility is that DAG can directly activate TRPC3, and produce receptor-operated Ca 2+ entry (ROCE). GPCRs, G-protein coupled receptors; IP3R, inositol 1,4,5-trisphosphate receptors; PLC, Phospholipase C; ROCE, Receptor–operated Ca 2+ entry; SOCE, store-operated Ca 2+ entry; SR/ER, sarco- endo-plasmic reticulum; STIM1, stromal interacting molecule 1; TRPC3,4,6 canonical transient receptor potential channel types 3,4,6; NCX Na/Ca exchange.
    Figure Legend Snippet: Signaling pathways involved in the activation of TRPC3 . The G-protein coupled receptor (GPCRs) activates phospholiase C (PLC), resulting in generation of IP3 and diacylglycerol (DAG). IP3 activates its receptor that leads to Ca 2+ release from SR/ER and the depletion of SR/ER Ca 2+ store. The Ca 2+ content change in the store can be sensed by STIM1, the ER Ca 2+ sensor and cause store-operated Ca 2+ entry (SOCE) through TRPC3 channels. An additional or alternate possibility is that DAG can directly activate TRPC3, and produce receptor-operated Ca 2+ entry (ROCE). GPCRs, G-protein coupled receptors; IP3R, inositol 1,4,5-trisphosphate receptors; PLC, Phospholipase C; ROCE, Receptor–operated Ca 2+ entry; SOCE, store-operated Ca 2+ entry; SR/ER, sarco- endo-plasmic reticulum; STIM1, stromal interacting molecule 1; TRPC3,4,6 canonical transient receptor potential channel types 3,4,6; NCX Na/Ca exchange.

    Techniques Used: Activation Assay, Planar Chromatography

    23) Product Images from "Resistance to Store Depletion-induced Endothelial Injury in Rat Lung after Chronic Heart Failure"

    Article Title: Resistance to Store Depletion-induced Endothelial Injury in Rat Lung after Chronic Heart Failure

    Journal:

    doi: 10.1164/rccm.200506-847OC

    Immunostaining for transient receptor potential proteins TRPC1, TRPC3, TRPC4, and TRPC6/7 in rat lungs. Expression of each TRPC isoform in endothelium was confirmed in small muscular or partially muscularized extraalveolar vessels ( A ) and conduit artery
    Figure Legend Snippet: Immunostaining for transient receptor potential proteins TRPC1, TRPC3, TRPC4, and TRPC6/7 in rat lungs. Expression of each TRPC isoform in endothelium was confirmed in small muscular or partially muscularized extraalveolar vessels ( A ) and conduit artery

    Techniques Used: Immunostaining, Expressing

    24) Product Images from "The involvement of TRPC3 channels in sinoatrial arrhythmias"

    Article Title: The involvement of TRPC3 channels in sinoatrial arrhythmias

    Journal: Frontiers in Physiology

    doi: 10.3389/fphys.2015.00086

    The effect of angiotension II (Ang II) and a selective TRPC3 channel blocker Pyr10 on pacemaker action potential after application of Ang II. (A–D) Intracellular recordings from WT mice; (E–H ) Intracellular recordings from TRPC3 −/− mice with conditions as indicated in each panel.
    Figure Legend Snippet: The effect of angiotension II (Ang II) and a selective TRPC3 channel blocker Pyr10 on pacemaker action potential after application of Ang II. (A–D) Intracellular recordings from WT mice; (E–H ) Intracellular recordings from TRPC3 −/− mice with conditions as indicated in each panel.

    Techniques Used: Mouse Assay

    Atrial arrhythmias induced by Ang II and pacing . Reconstructed excitation maps of data from multi-electrode array recordings (images below) from a Langendorff-perfused isolated heart. (A) sinus rhythm and (B) pacing induced atrial tachycardia. Note in AT, the ectopic beat starts from a different site to sinus rhythms. (C) Representative ECG recordings from isolated heart from WT and TRPC3 −/− mice under control conditions. (D) Atrial tachycardia and atrial fibrillation induced by burst pacing in WT mice and atrial-ventricular conduction block recorded from a TRPC −/− mouse heart respectively. (E) On average, the arrhythmia index was significant reduced in TRPC3KO mice ( n = 8) compared to WT mice ( n = 11, P = 0.004).
    Figure Legend Snippet: Atrial arrhythmias induced by Ang II and pacing . Reconstructed excitation maps of data from multi-electrode array recordings (images below) from a Langendorff-perfused isolated heart. (A) sinus rhythm and (B) pacing induced atrial tachycardia. Note in AT, the ectopic beat starts from a different site to sinus rhythms. (C) Representative ECG recordings from isolated heart from WT and TRPC3 −/− mice under control conditions. (D) Atrial tachycardia and atrial fibrillation induced by burst pacing in WT mice and atrial-ventricular conduction block recorded from a TRPC −/− mouse heart respectively. (E) On average, the arrhythmia index was significant reduced in TRPC3KO mice ( n = 8) compared to WT mice ( n = 11, P = 0.004).

    Techniques Used: Isolation, Mouse Assay, Blocking Assay

    Isolated single SAN cells express TRPC3, STIM1, and IP 3 R2 . Confocal immunofluorenscence images of isolated single pacemaker cells. Isolated group (A) and single (B) pacemaker cells were positively stained with anti HCN4 (in green). (C) Anti-TRPC3 in red, anti-STIM1 in green. (D) Anti-IP3R2 in red, anti-STIM1 in green. (E) 3D reconstruction image using N-SIM microscopy. Anti-IP 3 R2 in red, anti-STIM1 in green. Areas of co-localisation appear yellow due to color mixing.
    Figure Legend Snippet: Isolated single SAN cells express TRPC3, STIM1, and IP 3 R2 . Confocal immunofluorenscence images of isolated single pacemaker cells. Isolated group (A) and single (B) pacemaker cells were positively stained with anti HCN4 (in green). (C) Anti-TRPC3 in red, anti-STIM1 in green. (D) Anti-IP3R2 in red, anti-STIM1 in green. (E) 3D reconstruction image using N-SIM microscopy. Anti-IP 3 R2 in red, anti-STIM1 in green. Areas of co-localisation appear yellow due to color mixing.

    Techniques Used: Isolation, Staining, Microscopy

    The effect of Pyr10 on intracellular Ca 2+ after application of 1-oleoy1-2-acyl-sn-glycerol (OAG) 100 μM . The intact SANs were loaded with Ca 2+ indicator indo-1. (A,B) show intracellular Ca 2+ recordings from a WT mouse. (B,C) The statistics pool data shows percentage changes in resting Ca 2+ , Ca 2+ transient and firing rate in WT and TRPC3 KO mice respectively. (B) OAG treatment against control. (C) 2 μM Pyr10 treatment against OAG treatment. * P
    Figure Legend Snippet: The effect of Pyr10 on intracellular Ca 2+ after application of 1-oleoy1-2-acyl-sn-glycerol (OAG) 100 μM . The intact SANs were loaded with Ca 2+ indicator indo-1. (A,B) show intracellular Ca 2+ recordings from a WT mouse. (B,C) The statistics pool data shows percentage changes in resting Ca 2+ , Ca 2+ transient and firing rate in WT and TRPC3 KO mice respectively. (B) OAG treatment against control. (C) 2 μM Pyr10 treatment against OAG treatment. * P

    Techniques Used: Mouse Assay

    Signaling pathways involved in the activation of TRPC3 . The G-protein coupled receptor (GPCRs) activates phospholiase C (PLC), resulting in generation of IP3 and diacylglycerol (DAG). IP3 activates its receptor that leads to Ca 2+ release from SR/ER and the depletion of SR/ER Ca 2+ store. The Ca 2+ content change in the store can be sensed by STIM1, the ER Ca 2+ sensor and cause store-operated Ca 2+ entry (SOCE) through TRPC3 channels. An additional or alternate possibility is that DAG can directly activate TRPC3, and produce receptor-operated Ca 2+ entry (ROCE). GPCRs, G-protein coupled receptors; IP3R, inositol 1,4,5-trisphosphate receptors; PLC, Phospholipase C; ROCE, Receptor–operated Ca 2+ entry; SOCE, store-operated Ca 2+ entry; SR/ER, sarco- endo-plasmic reticulum; STIM1, stromal interacting molecule 1; TRPC3,4,6 canonical transient receptor potential channel types 3,4,6; NCX Na/Ca exchange.
    Figure Legend Snippet: Signaling pathways involved in the activation of TRPC3 . The G-protein coupled receptor (GPCRs) activates phospholiase C (PLC), resulting in generation of IP3 and diacylglycerol (DAG). IP3 activates its receptor that leads to Ca 2+ release from SR/ER and the depletion of SR/ER Ca 2+ store. The Ca 2+ content change in the store can be sensed by STIM1, the ER Ca 2+ sensor and cause store-operated Ca 2+ entry (SOCE) through TRPC3 channels. An additional or alternate possibility is that DAG can directly activate TRPC3, and produce receptor-operated Ca 2+ entry (ROCE). GPCRs, G-protein coupled receptors; IP3R, inositol 1,4,5-trisphosphate receptors; PLC, Phospholipase C; ROCE, Receptor–operated Ca 2+ entry; SOCE, store-operated Ca 2+ entry; SR/ER, sarco- endo-plasmic reticulum; STIM1, stromal interacting molecule 1; TRPC3,4,6 canonical transient receptor potential channel types 3,4,6; NCX Na/Ca exchange.

    Techniques Used: Activation Assay, Planar Chromatography

    25) Product Images from "Resistance to Store Depletion-induced Endothelial Injury in Rat Lung after Chronic Heart Failure"

    Article Title: Resistance to Store Depletion-induced Endothelial Injury in Rat Lung after Chronic Heart Failure

    Journal:

    doi: 10.1164/rccm.200506-847OC

    Immunostaining for transient receptor potential proteins TRPC1, TRPC3, TRPC4, and TRPC6/7 in rat lungs. Expression of each TRPC isoform in endothelium was confirmed in small muscular or partially muscularized extraalveolar vessels ( A ) and conduit artery
    Figure Legend Snippet: Immunostaining for transient receptor potential proteins TRPC1, TRPC3, TRPC4, and TRPC6/7 in rat lungs. Expression of each TRPC isoform in endothelium was confirmed in small muscular or partially muscularized extraalveolar vessels ( A ) and conduit artery

    Techniques Used: Immunostaining, Expressing

    26) Product Images from "The involvement of TRPC3 channels in sinoatrial arrhythmias"

    Article Title: The involvement of TRPC3 channels in sinoatrial arrhythmias

    Journal: Frontiers in Physiology

    doi: 10.3389/fphys.2015.00086

    The effect of angiotension II (Ang II) and a selective TRPC3 channel blocker Pyr10 on pacemaker action potential after application of Ang II. (A–D) Intracellular recordings from WT mice; (E–H ) Intracellular recordings from TRPC3 −/− mice with conditions as indicated in each panel.
    Figure Legend Snippet: The effect of angiotension II (Ang II) and a selective TRPC3 channel blocker Pyr10 on pacemaker action potential after application of Ang II. (A–D) Intracellular recordings from WT mice; (E–H ) Intracellular recordings from TRPC3 −/− mice with conditions as indicated in each panel.

    Techniques Used: Mouse Assay

    Atrial arrhythmias induced by Ang II and pacing . Reconstructed excitation maps of data from multi-electrode array recordings (images below) from a Langendorff-perfused isolated heart. (A) sinus rhythm and (B) pacing induced atrial tachycardia. Note in AT, the ectopic beat starts from a different site to sinus rhythms. (C) Representative ECG recordings from isolated heart from WT and TRPC3 −/− mice under control conditions. (D) Atrial tachycardia and atrial fibrillation induced by burst pacing in WT mice and atrial-ventricular conduction block recorded from a TRPC −/− mouse heart respectively. (E) On average, the arrhythmia index was significant reduced in TRPC3KO mice ( n = 8) compared to WT mice ( n = 11, P = 0.004).
    Figure Legend Snippet: Atrial arrhythmias induced by Ang II and pacing . Reconstructed excitation maps of data from multi-electrode array recordings (images below) from a Langendorff-perfused isolated heart. (A) sinus rhythm and (B) pacing induced atrial tachycardia. Note in AT, the ectopic beat starts from a different site to sinus rhythms. (C) Representative ECG recordings from isolated heart from WT and TRPC3 −/− mice under control conditions. (D) Atrial tachycardia and atrial fibrillation induced by burst pacing in WT mice and atrial-ventricular conduction block recorded from a TRPC −/− mouse heart respectively. (E) On average, the arrhythmia index was significant reduced in TRPC3KO mice ( n = 8) compared to WT mice ( n = 11, P = 0.004).

    Techniques Used: Isolation, Mouse Assay, Blocking Assay

    Isolated single SAN cells express TRPC3, STIM1, and IP 3 R2 . Confocal immunofluorenscence images of isolated single pacemaker cells. Isolated group (A) and single (B) pacemaker cells were positively stained with anti HCN4 (in green). (C) Anti-TRPC3 in red, anti-STIM1 in green. (D) Anti-IP3R2 in red, anti-STIM1 in green. (E) 3D reconstruction image using N-SIM microscopy. Anti-IP 3 R2 in red, anti-STIM1 in green. Areas of co-localisation appear yellow due to color mixing.
    Figure Legend Snippet: Isolated single SAN cells express TRPC3, STIM1, and IP 3 R2 . Confocal immunofluorenscence images of isolated single pacemaker cells. Isolated group (A) and single (B) pacemaker cells were positively stained with anti HCN4 (in green). (C) Anti-TRPC3 in red, anti-STIM1 in green. (D) Anti-IP3R2 in red, anti-STIM1 in green. (E) 3D reconstruction image using N-SIM microscopy. Anti-IP 3 R2 in red, anti-STIM1 in green. Areas of co-localisation appear yellow due to color mixing.

    Techniques Used: Isolation, Staining, Microscopy

    The effect of Pyr10 on intracellular Ca 2+ after application of 1-oleoy1-2-acyl-sn-glycerol (OAG) 100 μM . The intact SANs were loaded with Ca 2+ indicator indo-1. (A,B) show intracellular Ca 2+ recordings from a WT mouse. (B,C) The statistics pool data shows percentage changes in resting Ca 2+ , Ca 2+ transient and firing rate in WT and TRPC3 KO mice respectively. (B) OAG treatment against control. (C) 2 μM Pyr10 treatment against OAG treatment. * P
    Figure Legend Snippet: The effect of Pyr10 on intracellular Ca 2+ after application of 1-oleoy1-2-acyl-sn-glycerol (OAG) 100 μM . The intact SANs were loaded with Ca 2+ indicator indo-1. (A,B) show intracellular Ca 2+ recordings from a WT mouse. (B,C) The statistics pool data shows percentage changes in resting Ca 2+ , Ca 2+ transient and firing rate in WT and TRPC3 KO mice respectively. (B) OAG treatment against control. (C) 2 μM Pyr10 treatment against OAG treatment. * P

    Techniques Used: Mouse Assay

    Signaling pathways involved in the activation of TRPC3 . The G-protein coupled receptor (GPCRs) activates phospholiase C (PLC), resulting in generation of IP3 and diacylglycerol (DAG). IP3 activates its receptor that leads to Ca 2+ release from SR/ER and the depletion of SR/ER Ca 2+ store. The Ca 2+ content change in the store can be sensed by STIM1, the ER Ca 2+ sensor and cause store-operated Ca 2+ entry (SOCE) through TRPC3 channels. An additional or alternate possibility is that DAG can directly activate TRPC3, and produce receptor-operated Ca 2+ entry (ROCE). GPCRs, G-protein coupled receptors; IP3R, inositol 1,4,5-trisphosphate receptors; PLC, Phospholipase C; ROCE, Receptor–operated Ca 2+ entry; SOCE, store-operated Ca 2+ entry; SR/ER, sarco- endo-plasmic reticulum; STIM1, stromal interacting molecule 1; TRPC3,4,6 canonical transient receptor potential channel types 3,4,6; NCX Na/Ca exchange.
    Figure Legend Snippet: Signaling pathways involved in the activation of TRPC3 . The G-protein coupled receptor (GPCRs) activates phospholiase C (PLC), resulting in generation of IP3 and diacylglycerol (DAG). IP3 activates its receptor that leads to Ca 2+ release from SR/ER and the depletion of SR/ER Ca 2+ store. The Ca 2+ content change in the store can be sensed by STIM1, the ER Ca 2+ sensor and cause store-operated Ca 2+ entry (SOCE) through TRPC3 channels. An additional or alternate possibility is that DAG can directly activate TRPC3, and produce receptor-operated Ca 2+ entry (ROCE). GPCRs, G-protein coupled receptors; IP3R, inositol 1,4,5-trisphosphate receptors; PLC, Phospholipase C; ROCE, Receptor–operated Ca 2+ entry; SOCE, store-operated Ca 2+ entry; SR/ER, sarco- endo-plasmic reticulum; STIM1, stromal interacting molecule 1; TRPC3,4,6 canonical transient receptor potential channel types 3,4,6; NCX Na/Ca exchange.

    Techniques Used: Activation Assay, Planar Chromatography

    27) Product Images from "Resistance to Store Depletion-induced Endothelial Injury in Rat Lung after Chronic Heart Failure"

    Article Title: Resistance to Store Depletion-induced Endothelial Injury in Rat Lung after Chronic Heart Failure

    Journal:

    doi: 10.1164/rccm.200506-847OC

    Immunostaining for transient receptor potential proteins TRPC1, TRPC3, TRPC4, and TRPC6/7 in rat lungs. Expression of each TRPC isoform in endothelium was confirmed in small muscular or partially muscularized extraalveolar vessels ( A ) and conduit artery
    Figure Legend Snippet: Immunostaining for transient receptor potential proteins TRPC1, TRPC3, TRPC4, and TRPC6/7 in rat lungs. Expression of each TRPC isoform in endothelium was confirmed in small muscular or partially muscularized extraalveolar vessels ( A ) and conduit artery

    Techniques Used: Immunostaining, Expressing

    28) Product Images from "Characterization of pressure-mediated vascular tone in resistance arteries from bile duct-ligated rats"

    Article Title: Characterization of pressure-mediated vascular tone in resistance arteries from bile duct-ligated rats

    Journal: Oncotarget

    doi: 10.18632/oncotarget.15409

    Expression of ion channels in small mesenteric resistance arteries from SHAM- and BDL-rats A . Immunoblots for Ca v 1.2, TRPC3 and β-actin (loading control) from small mesenteric artery homogenates indicate no differences among both groups. The bar graphs show densitometry analyses for Ca v 1.2 and TRPC3. B . qPCR for Ca v 1.2. Representative IHC photographs and qPCR indicate that in the arteries from the BDL-rats, expressions of C ., D . BK Ca and E ., F . TRPC6 are reduced when compared to SHAM-rats ( n = 3-5/group). G . The immunoblots for both BK Ca and TRPC6 and their loading controls are shown. The bar graphs show densitometry analyses for BK Ca and TRPC6 ( n = 3/group). * P
    Figure Legend Snippet: Expression of ion channels in small mesenteric resistance arteries from SHAM- and BDL-rats A . Immunoblots for Ca v 1.2, TRPC3 and β-actin (loading control) from small mesenteric artery homogenates indicate no differences among both groups. The bar graphs show densitometry analyses for Ca v 1.2 and TRPC3. B . qPCR for Ca v 1.2. Representative IHC photographs and qPCR indicate that in the arteries from the BDL-rats, expressions of C ., D . BK Ca and E ., F . TRPC6 are reduced when compared to SHAM-rats ( n = 3-5/group). G . The immunoblots for both BK Ca and TRPC6 and their loading controls are shown. The bar graphs show densitometry analyses for BK Ca and TRPC6 ( n = 3/group). * P

    Techniques Used: Expressing, Western Blot, Real-time Polymerase Chain Reaction, Immunohistochemistry

    29) Product Images from "New Features on the Expression and Trafficking of mGluR1 Splice Variants Exposed by Two Novel Mutant Mouse Lines"

    Article Title: New Features on the Expression and Trafficking of mGluR1 Splice Variants Exposed by Two Novel Mutant Mouse Lines

    Journal: Frontiers in Molecular Neuroscience

    doi: 10.3389/fnmol.2018.00439

    Generation of mGluR1a-P/E-rescue mice. (A) Schematic drawing of the L7-mGluR1a-P/E transgene construct. Proline 1153 was replaced by a glutamate residue in the Homer binding domain of mGluR1α. Rat mGluR1α-P/E cDNA was inserted into the L7 promoter vector. Open and yellow boxes represent exons of the L7 gene and an insulator sequence, respectively. The expected size of Dra I- Pvu II restriction fragment from the transgene is shown below the drawing. (B) Southern blot analysis of genomic DNA isolated from tail biopsies of hemizygous L7-mGluR1a-P/E transgenic (Tg) mice. Genomic integration of the transgene was confirmed by the presence of a 2.6-kb Dra I- Pvu II fragment. Among four founder mice, #28 line was intercrossed to generate homozygous Tg mice for further experiments. (C) Immunofluorescence analysis of mGluR1 protein expression. Parasagittal sections from WT (left), mGluR1-knock out (KO; middle) and mGluR1a-P/E-rescue (right) mice were stained with an antibody against the extracellular domain of mGluR1. Scale bar, 1 mm. (D) Double immunofluorescence analysis of mGluR1α expression in cerebellar Purkinje cells (PCs; somata indicated by *) of 10 weeks-old WT (upper) and mGluR1a-P/E-rescue (lower) mice. Sections were stained with antibodies against mGluR1α (RED), calbindin (green) and counterstained with DAPI (blue). Mo, molecular layer. Scale bars, 20 μm (left panels) and 5 μm (right panels). (E) Immunoprecipitation analysis of the mGluR1 protein complex. GluRδ2 and transient receptor potential channel 3 (TRPC3), but not long Homers, were immunoprecipitated (IP) by the mGluR1 antibody from mouse cerebellar protein extracts, confirming that the P/E mutation indeed prevents in vivo the interaction between the C-terminal tail of mGluR1α and long Homers. (F) Rotarod task in WT ( n = 4), mGluR1a-P/E-rescue ( n = 6) and mGluR1-KO ( n = 6) mice. Each mouse was subjected to three training sessions per day for 5 days on the accelerating rotarod (4–40 rpm over 300 s). No significant differences were detected between WT and mGluR1a-P/E-rescue mice (Two-way ANOVA, interaction F (8,65) = 1.389, p = 0.2181), whereas mGluR1-KO mice were unable to remain on the rod (genotype F (2,65) = 122.8, p
    Figure Legend Snippet: Generation of mGluR1a-P/E-rescue mice. (A) Schematic drawing of the L7-mGluR1a-P/E transgene construct. Proline 1153 was replaced by a glutamate residue in the Homer binding domain of mGluR1α. Rat mGluR1α-P/E cDNA was inserted into the L7 promoter vector. Open and yellow boxes represent exons of the L7 gene and an insulator sequence, respectively. The expected size of Dra I- Pvu II restriction fragment from the transgene is shown below the drawing. (B) Southern blot analysis of genomic DNA isolated from tail biopsies of hemizygous L7-mGluR1a-P/E transgenic (Tg) mice. Genomic integration of the transgene was confirmed by the presence of a 2.6-kb Dra I- Pvu II fragment. Among four founder mice, #28 line was intercrossed to generate homozygous Tg mice for further experiments. (C) Immunofluorescence analysis of mGluR1 protein expression. Parasagittal sections from WT (left), mGluR1-knock out (KO; middle) and mGluR1a-P/E-rescue (right) mice were stained with an antibody against the extracellular domain of mGluR1. Scale bar, 1 mm. (D) Double immunofluorescence analysis of mGluR1α expression in cerebellar Purkinje cells (PCs; somata indicated by *) of 10 weeks-old WT (upper) and mGluR1a-P/E-rescue (lower) mice. Sections were stained with antibodies against mGluR1α (RED), calbindin (green) and counterstained with DAPI (blue). Mo, molecular layer. Scale bars, 20 μm (left panels) and 5 μm (right panels). (E) Immunoprecipitation analysis of the mGluR1 protein complex. GluRδ2 and transient receptor potential channel 3 (TRPC3), but not long Homers, were immunoprecipitated (IP) by the mGluR1 antibody from mouse cerebellar protein extracts, confirming that the P/E mutation indeed prevents in vivo the interaction between the C-terminal tail of mGluR1α and long Homers. (F) Rotarod task in WT ( n = 4), mGluR1a-P/E-rescue ( n = 6) and mGluR1-KO ( n = 6) mice. Each mouse was subjected to three training sessions per day for 5 days on the accelerating rotarod (4–40 rpm over 300 s). No significant differences were detected between WT and mGluR1a-P/E-rescue mice (Two-way ANOVA, interaction F (8,65) = 1.389, p = 0.2181), whereas mGluR1-KO mice were unable to remain on the rod (genotype F (2,65) = 122.8, p

    Techniques Used: Mouse Assay, Construct, Binding Assay, Plasmid Preparation, Sequencing, Southern Blot, Isolation, Transgenic Assay, Immunofluorescence, Expressing, Knock-Out, Staining, Immunoprecipitation, Mutagenesis, In Vivo

    30) Product Images from "Overexpression of TRPC3 increases apoptosis but not necrosis in response to ischemia/reperfusion in adult mouse cardiomyocytes"

    Article Title: Overexpression of TRPC3 increases apoptosis but not necrosis in response to ischemia/reperfusion in adult mouse cardiomyocytes

    Journal: American journal of physiology. Cell physiology

    doi: 10.1152/ajpcell.00313.2007

    After 1h of isolation, cardiomyocytes from WT and TRPC3 transgenic mice were treated with 10 ng/ml TNF-α for 2h or 18h, viability and apoptosis were measured by Annexin V-propidium iodide staining. Data presented as mean ± SEM of six individual experiments, i.e., six separate mouse cardiomyocytes isolations; with at least 300 cells counted per experiment under each condition. * = p
    Figure Legend Snippet: After 1h of isolation, cardiomyocytes from WT and TRPC3 transgenic mice were treated with 10 ng/ml TNF-α for 2h or 18h, viability and apoptosis were measured by Annexin V-propidium iodide staining. Data presented as mean ± SEM of six individual experiments, i.e., six separate mouse cardiomyocytes isolations; with at least 300 cells counted per experiment under each condition. * = p

    Techniques Used: Isolation, Transgenic Assay, Mouse Assay, Staining

    A) Bright field phase contrast images of cardiomyocytes from wild-type (WT) and TRPC3 transgenic mice at baseline, at the end of 90 min ischemia and after 90 min ischemia and 3 hours reperfusion; B) cardiomyocytes from wild-type (WT) and TRPC3 transgenic mice after 90 min ischemia and 3 hours reperfusion showing Annexin V and propidium iodide staining. Arrows in merged image indicate necrotic cells, staining positive for both Annexin V and propidium iodide.
    Figure Legend Snippet: A) Bright field phase contrast images of cardiomyocytes from wild-type (WT) and TRPC3 transgenic mice at baseline, at the end of 90 min ischemia and after 90 min ischemia and 3 hours reperfusion; B) cardiomyocytes from wild-type (WT) and TRPC3 transgenic mice after 90 min ischemia and 3 hours reperfusion showing Annexin V and propidium iodide staining. Arrows in merged image indicate necrotic cells, staining positive for both Annexin V and propidium iodide.

    Techniques Used: Transgenic Assay, Mouse Assay, Staining

    A) Cell viability assessed by % rod shaped cells; B) % apoptotic cells indicated by Annexin V positive and propidium iodide negative staining; C) % necrotic cells indicated by Annexin V and propidium iodide positive staining in cardiomyocytes from wild-type (WT) and TRPC3 transgenic mice at baseline, at the end of 90 min ischemia and after 90 min ischemia and 3 hours reperfusion (I/R). Data presented as mean ± SEM of six individual experiments (i.e., six separate mouse cardiomyocytes isolations; with at least 300 cells counted per experiment under each condition). # = p
    Figure Legend Snippet: A) Cell viability assessed by % rod shaped cells; B) % apoptotic cells indicated by Annexin V positive and propidium iodide negative staining; C) % necrotic cells indicated by Annexin V and propidium iodide positive staining in cardiomyocytes from wild-type (WT) and TRPC3 transgenic mice at baseline, at the end of 90 min ischemia and after 90 min ischemia and 3 hours reperfusion (I/R). Data presented as mean ± SEM of six individual experiments (i.e., six separate mouse cardiomyocytes isolations; with at least 300 cells counted per experiment under each condition). # = p

    Techniques Used: Negative Staining, Staining, Transgenic Assay, Mouse Assay

    Cardiomyocytes from wild-type (WT) and TRPC3 transgenic mice were treated with 5 μM thapsigargin for 5 mins in the absence of extracellular Ca 2+ and were then exposed to 2.5 mM extracellular calcium. Upper panels show a typical time course of myocyte rounding following addition of 2.5 mM Ca 2+ in WT and TRPC3 cardiomyocytes. Data are mean ± SEM from 15 cells of at least 3 individual experiments. # = p
    Figure Legend Snippet: Cardiomyocytes from wild-type (WT) and TRPC3 transgenic mice were treated with 5 μM thapsigargin for 5 mins in the absence of extracellular Ca 2+ and were then exposed to 2.5 mM extracellular calcium. Upper panels show a typical time course of myocyte rounding following addition of 2.5 mM Ca 2+ in WT and TRPC3 cardiomyocytes. Data are mean ± SEM from 15 cells of at least 3 individual experiments. # = p

    Techniques Used: Transgenic Assay, Mouse Assay

    A) TRPC3 and B) TRPC1 protein expression in the hearts from wild-type (WT) and TRPC3 transgenic mice. Upper panels are representative immunoblots and the lower panels are mean densitometric data from 6 individual experiments normalized to β-actin. # = p
    Figure Legend Snippet: A) TRPC3 and B) TRPC1 protein expression in the hearts from wild-type (WT) and TRPC3 transgenic mice. Upper panels are representative immunoblots and the lower panels are mean densitometric data from 6 individual experiments normalized to β-actin. # = p

    Techniques Used: Expressing, Transgenic Assay, Mouse Assay, Western Blot

    31) Product Images from "The involvement of TRPC3 channels in sinoatrial arrhythmias"

    Article Title: The involvement of TRPC3 channels in sinoatrial arrhythmias

    Journal: Frontiers in Physiology

    doi: 10.3389/fphys.2015.00086

    The effect of angiotension II (Ang II) and a selective TRPC3 channel blocker Pyr10 on pacemaker action potential after application of Ang II. (A–D) Intracellular recordings from WT mice; (E–H ) Intracellular recordings from TRPC3 −/− mice with conditions as indicated in each panel.
    Figure Legend Snippet: The effect of angiotension II (Ang II) and a selective TRPC3 channel blocker Pyr10 on pacemaker action potential after application of Ang II. (A–D) Intracellular recordings from WT mice; (E–H ) Intracellular recordings from TRPC3 −/− mice with conditions as indicated in each panel.

    Techniques Used: Mouse Assay

    Atrial arrhythmias induced by Ang II and pacing . Reconstructed excitation maps of data from multi-electrode array recordings (images below) from a Langendorff-perfused isolated heart. (A) sinus rhythm and (B) pacing induced atrial tachycardia. Note in AT, the ectopic beat starts from a different site to sinus rhythms. (C) Representative ECG recordings from isolated heart from WT and TRPC3 −/− mice under control conditions. (D) Atrial tachycardia and atrial fibrillation induced by burst pacing in WT mice and atrial-ventricular conduction block recorded from a TRPC −/− mouse heart respectively. (E) On average, the arrhythmia index was significant reduced in TRPC3KO mice ( n = 8) compared to WT mice ( n = 11, P = 0.004).
    Figure Legend Snippet: Atrial arrhythmias induced by Ang II and pacing . Reconstructed excitation maps of data from multi-electrode array recordings (images below) from a Langendorff-perfused isolated heart. (A) sinus rhythm and (B) pacing induced atrial tachycardia. Note in AT, the ectopic beat starts from a different site to sinus rhythms. (C) Representative ECG recordings from isolated heart from WT and TRPC3 −/− mice under control conditions. (D) Atrial tachycardia and atrial fibrillation induced by burst pacing in WT mice and atrial-ventricular conduction block recorded from a TRPC −/− mouse heart respectively. (E) On average, the arrhythmia index was significant reduced in TRPC3KO mice ( n = 8) compared to WT mice ( n = 11, P = 0.004).

    Techniques Used: Isolation, Mouse Assay, Blocking Assay

    Isolated single SAN cells express TRPC3, STIM1, and IP 3 R2 . Confocal immunofluorenscence images of isolated single pacemaker cells. Isolated group (A) and single (B) pacemaker cells were positively stained with anti HCN4 (in green). (C) Anti-TRPC3 in red, anti-STIM1 in green. (D) Anti-IP3R2 in red, anti-STIM1 in green. (E) 3D reconstruction image using N-SIM microscopy. Anti-IP 3 R2 in red, anti-STIM1 in green. Areas of co-localisation appear yellow due to color mixing.
    Figure Legend Snippet: Isolated single SAN cells express TRPC3, STIM1, and IP 3 R2 . Confocal immunofluorenscence images of isolated single pacemaker cells. Isolated group (A) and single (B) pacemaker cells were positively stained with anti HCN4 (in green). (C) Anti-TRPC3 in red, anti-STIM1 in green. (D) Anti-IP3R2 in red, anti-STIM1 in green. (E) 3D reconstruction image using N-SIM microscopy. Anti-IP 3 R2 in red, anti-STIM1 in green. Areas of co-localisation appear yellow due to color mixing.

    Techniques Used: Isolation, Staining, Microscopy

    The effect of Pyr10 on intracellular Ca 2+ after application of 1-oleoy1-2-acyl-sn-glycerol (OAG) 100 μM . The intact SANs were loaded with Ca 2+ indicator indo-1. (A,B) show intracellular Ca 2+ recordings from a WT mouse. (B,C) The statistics pool data shows percentage changes in resting Ca 2+ , Ca 2+ transient and firing rate in WT and TRPC3 KO mice respectively. (B) OAG treatment against control. (C) 2 μM Pyr10 treatment against OAG treatment. * P
    Figure Legend Snippet: The effect of Pyr10 on intracellular Ca 2+ after application of 1-oleoy1-2-acyl-sn-glycerol (OAG) 100 μM . The intact SANs were loaded with Ca 2+ indicator indo-1. (A,B) show intracellular Ca 2+ recordings from a WT mouse. (B,C) The statistics pool data shows percentage changes in resting Ca 2+ , Ca 2+ transient and firing rate in WT and TRPC3 KO mice respectively. (B) OAG treatment against control. (C) 2 μM Pyr10 treatment against OAG treatment. * P

    Techniques Used: Mouse Assay

    Signaling pathways involved in the activation of TRPC3 . The G-protein coupled receptor (GPCRs) activates phospholiase C (PLC), resulting in generation of IP3 and diacylglycerol (DAG). IP3 activates its receptor that leads to Ca 2+ release from SR/ER and the depletion of SR/ER Ca 2+ store. The Ca 2+ content change in the store can be sensed by STIM1, the ER Ca 2+ sensor and cause store-operated Ca 2+ entry (SOCE) through TRPC3 channels. An additional or alternate possibility is that DAG can directly activate TRPC3, and produce receptor-operated Ca 2+ entry (ROCE). GPCRs, G-protein coupled receptors; IP3R, inositol 1,4,5-trisphosphate receptors; PLC, Phospholipase C; ROCE, Receptor–operated Ca 2+ entry; SOCE, store-operated Ca 2+ entry; SR/ER, sarco- endo-plasmic reticulum; STIM1, stromal interacting molecule 1; TRPC3,4,6 canonical transient receptor potential channel types 3,4,6; NCX Na/Ca exchange.
    Figure Legend Snippet: Signaling pathways involved in the activation of TRPC3 . The G-protein coupled receptor (GPCRs) activates phospholiase C (PLC), resulting in generation of IP3 and diacylglycerol (DAG). IP3 activates its receptor that leads to Ca 2+ release from SR/ER and the depletion of SR/ER Ca 2+ store. The Ca 2+ content change in the store can be sensed by STIM1, the ER Ca 2+ sensor and cause store-operated Ca 2+ entry (SOCE) through TRPC3 channels. An additional or alternate possibility is that DAG can directly activate TRPC3, and produce receptor-operated Ca 2+ entry (ROCE). GPCRs, G-protein coupled receptors; IP3R, inositol 1,4,5-trisphosphate receptors; PLC, Phospholipase C; ROCE, Receptor–operated Ca 2+ entry; SOCE, store-operated Ca 2+ entry; SR/ER, sarco- endo-plasmic reticulum; STIM1, stromal interacting molecule 1; TRPC3,4,6 canonical transient receptor potential channel types 3,4,6; NCX Na/Ca exchange.

    Techniques Used: Activation Assay, Planar Chromatography

    32) Product Images from "Store-operated Ca2+ entry in first trimester and term human placenta"

    Article Title: Store-operated Ca2+ entry in first trimester and term human placenta

    Journal: The Journal of Physiology

    doi: 10.1113/jphysiol.2003.044149

    RT-PCR detection of TRPC mRNA RT-PCR was performed using gene-specific primers for A , TRPC1, B , TRPC3, C , TRPC4, D , TRPC5, E , TRPC6 and F , β-actin. PCR product sizes are indicated. L, 100 bp ladder; −, negative control (H 2 O); SI, small intestine (Clontech); HB, human brain (Clontech); I, first trimester placenta ( n = 7 pooled); II, second trimester placentas ( n = 5 pooled); III, term placentas ( n = 8 pooled).
    Figure Legend Snippet: RT-PCR detection of TRPC mRNA RT-PCR was performed using gene-specific primers for A , TRPC1, B , TRPC3, C , TRPC4, D , TRPC5, E , TRPC6 and F , β-actin. PCR product sizes are indicated. L, 100 bp ladder; −, negative control (H 2 O); SI, small intestine (Clontech); HB, human brain (Clontech); I, first trimester placenta ( n = 7 pooled); II, second trimester placentas ( n = 5 pooled); III, term placentas ( n = 8 pooled).

    Techniques Used: Reverse Transcription Polymerase Chain Reaction, Polymerase Chain Reaction, Negative Control

    Immunocytochemistry showing localisation for TRPC3, TRPC4 and TRPC6 in first trimester ( A , C and E ) and term placenta ( B , D and F ) Negative controls for first trimester ( G ) and term placenta ( H ) are also shown. Samples were magnified × 1000; a scale bar is given in H . S, syncytiotrophoblast; C, cytotrophoblast; VC, villous core; M, microvillous membrane; B, basal membrane.
    Figure Legend Snippet: Immunocytochemistry showing localisation for TRPC3, TRPC4 and TRPC6 in first trimester ( A , C and E ) and term placenta ( B , D and F ) Negative controls for first trimester ( G ) and term placenta ( H ) are also shown. Samples were magnified × 1000; a scale bar is given in H . S, syncytiotrophoblast; C, cytotrophoblast; VC, villous core; M, microvillous membrane; B, basal membrane.

    Techniques Used: Immunocytochemistry

    Western blotting for TRPC1 ( A , B ), TRPC3 ( C , D ) and TRPC6 ( E , F ) Protein sizes are given. Preabsorption of each primary antibody with antigen (Ag) shows removal of specific bands ( A , C , E ). ECL detection was for 30 min for preabsorption studies and 1 h for comparison of first trimester and term tissue. RB, rat brain (positive control tissue); T, term placenta; F, first trimester placenta. In B , D , F , and G each lane corresponds to samples prepared from four different first trimester and term placentas.
    Figure Legend Snippet: Western blotting for TRPC1 ( A , B ), TRPC3 ( C , D ) and TRPC6 ( E , F ) Protein sizes are given. Preabsorption of each primary antibody with antigen (Ag) shows removal of specific bands ( A , C , E ). ECL detection was for 30 min for preabsorption studies and 1 h for comparison of first trimester and term tissue. RB, rat brain (positive control tissue); T, term placenta; F, first trimester placenta. In B , D , F , and G each lane corresponds to samples prepared from four different first trimester and term placentas.

    Techniques Used: Western Blot, Positive Control

    33) Product Images from "Resistance to Store Depletion-induced Endothelial Injury in Rat Lung after Chronic Heart Failure"

    Article Title: Resistance to Store Depletion-induced Endothelial Injury in Rat Lung after Chronic Heart Failure

    Journal:

    doi: 10.1164/rccm.200506-847OC

    Immunostaining for transient receptor potential proteins TRPC1, TRPC3, TRPC4, and TRPC6/7 in rat lungs. Expression of each TRPC isoform in endothelium was confirmed in small muscular or partially muscularized extraalveolar vessels ( A ) and conduit artery
    Figure Legend Snippet: Immunostaining for transient receptor potential proteins TRPC1, TRPC3, TRPC4, and TRPC6/7 in rat lungs. Expression of each TRPC isoform in endothelium was confirmed in small muscular or partially muscularized extraalveolar vessels ( A ) and conduit artery

    Techniques Used: Immunostaining, Expressing

    34) Product Images from "The involvement of TRPC3 channels in sinoatrial arrhythmias"

    Article Title: The involvement of TRPC3 channels in sinoatrial arrhythmias

    Journal: Frontiers in Physiology

    doi: 10.3389/fphys.2015.00086

    The effect of angiotension II (Ang II) and a selective TRPC3 channel blocker Pyr10 on pacemaker action potential after application of Ang II. (A–D) Intracellular recordings from WT mice; (E–H ) Intracellular recordings from TRPC3 −/− mice with conditions as indicated in each panel.
    Figure Legend Snippet: The effect of angiotension II (Ang II) and a selective TRPC3 channel blocker Pyr10 on pacemaker action potential after application of Ang II. (A–D) Intracellular recordings from WT mice; (E–H ) Intracellular recordings from TRPC3 −/− mice with conditions as indicated in each panel.

    Techniques Used: Mouse Assay

    Atrial arrhythmias induced by Ang II and pacing . Reconstructed excitation maps of data from multi-electrode array recordings (images below) from a Langendorff-perfused isolated heart. (A) sinus rhythm and (B) pacing induced atrial tachycardia. Note in AT, the ectopic beat starts from a different site to sinus rhythms. (C) Representative ECG recordings from isolated heart from WT and TRPC3 −/− mice under control conditions. (D) Atrial tachycardia and atrial fibrillation induced by burst pacing in WT mice and atrial-ventricular conduction block recorded from a TRPC −/− mouse heart respectively. (E) On average, the arrhythmia index was significant reduced in TRPC3KO mice ( n = 8) compared to WT mice ( n = 11, P = 0.004).
    Figure Legend Snippet: Atrial arrhythmias induced by Ang II and pacing . Reconstructed excitation maps of data from multi-electrode array recordings (images below) from a Langendorff-perfused isolated heart. (A) sinus rhythm and (B) pacing induced atrial tachycardia. Note in AT, the ectopic beat starts from a different site to sinus rhythms. (C) Representative ECG recordings from isolated heart from WT and TRPC3 −/− mice under control conditions. (D) Atrial tachycardia and atrial fibrillation induced by burst pacing in WT mice and atrial-ventricular conduction block recorded from a TRPC −/− mouse heart respectively. (E) On average, the arrhythmia index was significant reduced in TRPC3KO mice ( n = 8) compared to WT mice ( n = 11, P = 0.004).

    Techniques Used: Isolation, Mouse Assay, Blocking Assay

    Isolated single SAN cells express TRPC3, STIM1, and IP 3 R2 . Confocal immunofluorenscence images of isolated single pacemaker cells. Isolated group (A) and single (B) pacemaker cells were positively stained with anti HCN4 (in green). (C) Anti-TRPC3 in red, anti-STIM1 in green. (D) Anti-IP3R2 in red, anti-STIM1 in green. (E) 3D reconstruction image using N-SIM microscopy. Anti-IP 3 R2 in red, anti-STIM1 in green. Areas of co-localisation appear yellow due to color mixing.
    Figure Legend Snippet: Isolated single SAN cells express TRPC3, STIM1, and IP 3 R2 . Confocal immunofluorenscence images of isolated single pacemaker cells. Isolated group (A) and single (B) pacemaker cells were positively stained with anti HCN4 (in green). (C) Anti-TRPC3 in red, anti-STIM1 in green. (D) Anti-IP3R2 in red, anti-STIM1 in green. (E) 3D reconstruction image using N-SIM microscopy. Anti-IP 3 R2 in red, anti-STIM1 in green. Areas of co-localisation appear yellow due to color mixing.

    Techniques Used: Isolation, Staining, Microscopy

    The effect of Pyr10 on intracellular Ca 2+ after application of 1-oleoy1-2-acyl-sn-glycerol (OAG) 100 μM . The intact SANs were loaded with Ca 2+ indicator indo-1. (A,B) show intracellular Ca 2+ recordings from a WT mouse. (B,C) The statistics pool data shows percentage changes in resting Ca 2+ , Ca 2+ transient and firing rate in WT and TRPC3 KO mice respectively. (B) OAG treatment against control. (C) 2 μM Pyr10 treatment against OAG treatment. * P
    Figure Legend Snippet: The effect of Pyr10 on intracellular Ca 2+ after application of 1-oleoy1-2-acyl-sn-glycerol (OAG) 100 μM . The intact SANs were loaded with Ca 2+ indicator indo-1. (A,B) show intracellular Ca 2+ recordings from a WT mouse. (B,C) The statistics pool data shows percentage changes in resting Ca 2+ , Ca 2+ transient and firing rate in WT and TRPC3 KO mice respectively. (B) OAG treatment against control. (C) 2 μM Pyr10 treatment against OAG treatment. * P

    Techniques Used: Mouse Assay

    Signaling pathways involved in the activation of TRPC3 . The G-protein coupled receptor (GPCRs) activates phospholiase C (PLC), resulting in generation of IP3 and diacylglycerol (DAG). IP3 activates its receptor that leads to Ca 2+ release from SR/ER and the depletion of SR/ER Ca 2+ store. The Ca 2+ content change in the store can be sensed by STIM1, the ER Ca 2+ sensor and cause store-operated Ca 2+ entry (SOCE) through TRPC3 channels. An additional or alternate possibility is that DAG can directly activate TRPC3, and produce receptor-operated Ca 2+ entry (ROCE). GPCRs, G-protein coupled receptors; IP3R, inositol 1,4,5-trisphosphate receptors; PLC, Phospholipase C; ROCE, Receptor–operated Ca 2+ entry; SOCE, store-operated Ca 2+ entry; SR/ER, sarco- endo-plasmic reticulum; STIM1, stromal interacting molecule 1; TRPC3,4,6 canonical transient receptor potential channel types 3,4,6; NCX Na/Ca exchange.
    Figure Legend Snippet: Signaling pathways involved in the activation of TRPC3 . The G-protein coupled receptor (GPCRs) activates phospholiase C (PLC), resulting in generation of IP3 and diacylglycerol (DAG). IP3 activates its receptor that leads to Ca 2+ release from SR/ER and the depletion of SR/ER Ca 2+ store. The Ca 2+ content change in the store can be sensed by STIM1, the ER Ca 2+ sensor and cause store-operated Ca 2+ entry (SOCE) through TRPC3 channels. An additional or alternate possibility is that DAG can directly activate TRPC3, and produce receptor-operated Ca 2+ entry (ROCE). GPCRs, G-protein coupled receptors; IP3R, inositol 1,4,5-trisphosphate receptors; PLC, Phospholipase C; ROCE, Receptor–operated Ca 2+ entry; SOCE, store-operated Ca 2+ entry; SR/ER, sarco- endo-plasmic reticulum; STIM1, stromal interacting molecule 1; TRPC3,4,6 canonical transient receptor potential channel types 3,4,6; NCX Na/Ca exchange.

    Techniques Used: Activation Assay, Planar Chromatography

    35) Product Images from "The involvement of TRPC3 channels in sinoatrial arrhythmias"

    Article Title: The involvement of TRPC3 channels in sinoatrial arrhythmias

    Journal: Frontiers in Physiology

    doi: 10.3389/fphys.2015.00086

    The effect of angiotension II (Ang II) and a selective TRPC3 channel blocker Pyr10 on pacemaker action potential after application of Ang II. (A–D) Intracellular recordings from WT mice; (E–H ) Intracellular recordings from TRPC3 −/− mice with conditions as indicated in each panel.
    Figure Legend Snippet: The effect of angiotension II (Ang II) and a selective TRPC3 channel blocker Pyr10 on pacemaker action potential after application of Ang II. (A–D) Intracellular recordings from WT mice; (E–H ) Intracellular recordings from TRPC3 −/− mice with conditions as indicated in each panel.

    Techniques Used: Mouse Assay

    Atrial arrhythmias induced by Ang II and pacing . Reconstructed excitation maps of data from multi-electrode array recordings (images below) from a Langendorff-perfused isolated heart. (A) sinus rhythm and (B) pacing induced atrial tachycardia. Note in AT, the ectopic beat starts from a different site to sinus rhythms. (C) Representative ECG recordings from isolated heart from WT and TRPC3 −/− mice under control conditions. (D) Atrial tachycardia and atrial fibrillation induced by burst pacing in WT mice and atrial-ventricular conduction block recorded from a TRPC −/− mouse heart respectively. (E) On average, the arrhythmia index was significant reduced in TRPC3KO mice ( n = 8) compared to WT mice ( n = 11, P = 0.004).
    Figure Legend Snippet: Atrial arrhythmias induced by Ang II and pacing . Reconstructed excitation maps of data from multi-electrode array recordings (images below) from a Langendorff-perfused isolated heart. (A) sinus rhythm and (B) pacing induced atrial tachycardia. Note in AT, the ectopic beat starts from a different site to sinus rhythms. (C) Representative ECG recordings from isolated heart from WT and TRPC3 −/− mice under control conditions. (D) Atrial tachycardia and atrial fibrillation induced by burst pacing in WT mice and atrial-ventricular conduction block recorded from a TRPC −/− mouse heart respectively. (E) On average, the arrhythmia index was significant reduced in TRPC3KO mice ( n = 8) compared to WT mice ( n = 11, P = 0.004).

    Techniques Used: Isolation, Mouse Assay, Blocking Assay

    Isolated single SAN cells express TRPC3, STIM1, and IP 3 R2 . Confocal immunofluorenscence images of isolated single pacemaker cells. Isolated group (A) and single (B) pacemaker cells were positively stained with anti HCN4 (in green). (C) Anti-TRPC3 in red, anti-STIM1 in green. (D) Anti-IP3R2 in red, anti-STIM1 in green. (E) 3D reconstruction image using N-SIM microscopy. Anti-IP 3 R2 in red, anti-STIM1 in green. Areas of co-localisation appear yellow due to color mixing.
    Figure Legend Snippet: Isolated single SAN cells express TRPC3, STIM1, and IP 3 R2 . Confocal immunofluorenscence images of isolated single pacemaker cells. Isolated group (A) and single (B) pacemaker cells were positively stained with anti HCN4 (in green). (C) Anti-TRPC3 in red, anti-STIM1 in green. (D) Anti-IP3R2 in red, anti-STIM1 in green. (E) 3D reconstruction image using N-SIM microscopy. Anti-IP 3 R2 in red, anti-STIM1 in green. Areas of co-localisation appear yellow due to color mixing.

    Techniques Used: Isolation, Staining, Microscopy

    The effect of Pyr10 on intracellular Ca 2+ after application of 1-oleoy1-2-acyl-sn-glycerol (OAG) 100 μM . The intact SANs were loaded with Ca 2+ indicator indo-1. (A,B) show intracellular Ca 2+ recordings from a WT mouse. (B,C) The statistics pool data shows percentage changes in resting Ca 2+ , Ca 2+ transient and firing rate in WT and TRPC3 KO mice respectively. (B) OAG treatment against control. (C) 2 μM Pyr10 treatment against OAG treatment. * P
    Figure Legend Snippet: The effect of Pyr10 on intracellular Ca 2+ after application of 1-oleoy1-2-acyl-sn-glycerol (OAG) 100 μM . The intact SANs were loaded with Ca 2+ indicator indo-1. (A,B) show intracellular Ca 2+ recordings from a WT mouse. (B,C) The statistics pool data shows percentage changes in resting Ca 2+ , Ca 2+ transient and firing rate in WT and TRPC3 KO mice respectively. (B) OAG treatment against control. (C) 2 μM Pyr10 treatment against OAG treatment. * P

    Techniques Used: Mouse Assay

    Signaling pathways involved in the activation of TRPC3 . The G-protein coupled receptor (GPCRs) activates phospholiase C (PLC), resulting in generation of IP3 and diacylglycerol (DAG). IP3 activates its receptor that leads to Ca 2+ release from SR/ER and the depletion of SR/ER Ca 2+ store. The Ca 2+ content change in the store can be sensed by STIM1, the ER Ca 2+ sensor and cause store-operated Ca 2+ entry (SOCE) through TRPC3 channels. An additional or alternate possibility is that DAG can directly activate TRPC3, and produce receptor-operated Ca 2+ entry (ROCE). GPCRs, G-protein coupled receptors; IP3R, inositol 1,4,5-trisphosphate receptors; PLC, Phospholipase C; ROCE, Receptor–operated Ca 2+ entry; SOCE, store-operated Ca 2+ entry; SR/ER, sarco- endo-plasmic reticulum; STIM1, stromal interacting molecule 1; TRPC3,4,6 canonical transient receptor potential channel types 3,4,6; NCX Na/Ca exchange.
    Figure Legend Snippet: Signaling pathways involved in the activation of TRPC3 . The G-protein coupled receptor (GPCRs) activates phospholiase C (PLC), resulting in generation of IP3 and diacylglycerol (DAG). IP3 activates its receptor that leads to Ca 2+ release from SR/ER and the depletion of SR/ER Ca 2+ store. The Ca 2+ content change in the store can be sensed by STIM1, the ER Ca 2+ sensor and cause store-operated Ca 2+ entry (SOCE) through TRPC3 channels. An additional or alternate possibility is that DAG can directly activate TRPC3, and produce receptor-operated Ca 2+ entry (ROCE). GPCRs, G-protein coupled receptors; IP3R, inositol 1,4,5-trisphosphate receptors; PLC, Phospholipase C; ROCE, Receptor–operated Ca 2+ entry; SOCE, store-operated Ca 2+ entry; SR/ER, sarco- endo-plasmic reticulum; STIM1, stromal interacting molecule 1; TRPC3,4,6 canonical transient receptor potential channel types 3,4,6; NCX Na/Ca exchange.

    Techniques Used: Activation Assay, Planar Chromatography

    36) Product Images from "Increased rhythmicity in hypertensive arterial smooth muscle is linked to transient receptor potential canonical channels"

    Article Title: Increased rhythmicity in hypertensive arterial smooth muscle is linked to transient receptor potential canonical channels

    Journal: Journal of Cellular and Molecular Medicine

    doi: 10.1111/j.1582-4934.2009.00890.x

    Effect of candesartan or telmisartan but not amlodipine on TRPC expression in mesenteric arterioles. Long-term administration of angiotensin AT1 receptor antagonist telmisartan or candesartan, but not of calcium channel blocker amlodipine reduces TRPC1, TRPC3 and TRPC5 channel protein expression in vivo. The angiotensin AT1 receptor antagonist telmisartan (5 mg/kg per day) or candesartan (4 mg/kg per day), calcium channel blocker amlodipine (10 mg/kg per day), or placebo were administered to SHR by gavage for 16 weeks. Representative immunoblottings of TRPC channel protein expressions in mesenteric arterioles from treated SHR (Fig. 7A) and summary data are shown (Fig. 7B). * P
    Figure Legend Snippet: Effect of candesartan or telmisartan but not amlodipine on TRPC expression in mesenteric arterioles. Long-term administration of angiotensin AT1 receptor antagonist telmisartan or candesartan, but not of calcium channel blocker amlodipine reduces TRPC1, TRPC3 and TRPC5 channel protein expression in vivo. The angiotensin AT1 receptor antagonist telmisartan (5 mg/kg per day) or candesartan (4 mg/kg per day), calcium channel blocker amlodipine (10 mg/kg per day), or placebo were administered to SHR by gavage for 16 weeks. Representative immunoblottings of TRPC channel protein expressions in mesenteric arterioles from treated SHR (Fig. 7A) and summary data are shown (Fig. 7B). * P

    Techniques Used: Expressing, In Vivo

    Inhibition of norepinephrine-induced vasomotion in mesenteric arterioles from SHR by specific anti-TRPC antibodies Representative tracings of norepinephrine-induced vasomotion in mesenteric arterioles from SHR under control conditions (A), in the presence of anti-TRPC1 antibodies (B), anti-TRPC3 antibodies (C), anti-TRPC3 antibodies with antigenic peptide (D), anti-TRPC5 antibodies (E), anti-TRPC1 plus anti-TRPC3 antibodies (F) or in the presence of anti-β-actin antibodies (G). Representative tracings of norepinephrine-induced vasomotion in mesenteric arterioles from SHR under control conditions after short-term exposure to hypotonic 0.45% NaCl without antibody (H), or after short-term exposure to hypotonic 0.45% NaCl plus TRPC3 antibody (I). Summary data (J) show that inhibition of TRPC channels by specific anti-TRPC antibodies significantly attenuates norepinephrine-induced vasomotion in SHR. Data are mean ± S.E.M. of n = 6 independent experiments. * P
    Figure Legend Snippet: Inhibition of norepinephrine-induced vasomotion in mesenteric arterioles from SHR by specific anti-TRPC antibodies Representative tracings of norepinephrine-induced vasomotion in mesenteric arterioles from SHR under control conditions (A), in the presence of anti-TRPC1 antibodies (B), anti-TRPC3 antibodies (C), anti-TRPC3 antibodies with antigenic peptide (D), anti-TRPC5 antibodies (E), anti-TRPC1 plus anti-TRPC3 antibodies (F) or in the presence of anti-β-actin antibodies (G). Representative tracings of norepinephrine-induced vasomotion in mesenteric arterioles from SHR under control conditions after short-term exposure to hypotonic 0.45% NaCl without antibody (H), or after short-term exposure to hypotonic 0.45% NaCl plus TRPC3 antibody (I). Summary data (J) show that inhibition of TRPC channels by specific anti-TRPC antibodies significantly attenuates norepinephrine-induced vasomotion in SHR. Data are mean ± S.E.M. of n = 6 independent experiments. * P

    Techniques Used: Inhibition

    Expression of TRPC1, TRPC3 and TRPC5 channels in mesenteric arterioles from SHR. Representative immunoblottings (A) and summary data (B) of TRPC1, TRPC3, TRPC4, TRPC5,TRPC6 channels and TRPC3 antibody with its respective antigenic peptide the protein expression in mesenteric arterioles from WKY (open bars) and SHR (filled bars). Data are mean ± S.E.M. of n = 4 independent experiments. * P
    Figure Legend Snippet: Expression of TRPC1, TRPC3 and TRPC5 channels in mesenteric arterioles from SHR. Representative immunoblottings (A) and summary data (B) of TRPC1, TRPC3, TRPC4, TRPC5,TRPC6 channels and TRPC3 antibody with its respective antigenic peptide the protein expression in mesenteric arterioles from WKY (open bars) and SHR (filled bars). Data are mean ± S.E.M. of n = 4 independent experiments. * P

    Techniques Used: Expressing

    Inhibition of norepinephrine-induced calcium increase in mesenteric arterioles from SHR by specific anti-TRPC antibodies. Representative tracings of norepinephrine-induced calcium increase in mesenteric arterioles from SHR under control conditions and in the presence of anti-TRPC1 antibodies (A), anti-TRPC3 antibodies (B), anti-TRPC5 antibodies (C), anti-TRPC1 plus anti-TRPC3 plus anti-TRPC5 antibodies (D), anti-TRPC3 antibodies plus TRPC3 antigenic peptide (E) or anti-β-actin antibodies (F). Summary data (G) shows the effects of specific anti-TRPC antibodies on norepinephrine-induced calcium influx in mesenteric arterioles from SHR. Data are mean ± S.E.M. of n = 8 independent experiments. * P
    Figure Legend Snippet: Inhibition of norepinephrine-induced calcium increase in mesenteric arterioles from SHR by specific anti-TRPC antibodies. Representative tracings of norepinephrine-induced calcium increase in mesenteric arterioles from SHR under control conditions and in the presence of anti-TRPC1 antibodies (A), anti-TRPC3 antibodies (B), anti-TRPC5 antibodies (C), anti-TRPC1 plus anti-TRPC3 plus anti-TRPC5 antibodies (D), anti-TRPC3 antibodies plus TRPC3 antigenic peptide (E) or anti-β-actin antibodies (F). Summary data (G) shows the effects of specific anti-TRPC antibodies on norepinephrine-induced calcium influx in mesenteric arterioles from SHR. Data are mean ± S.E.M. of n = 8 independent experiments. * P

    Techniques Used: Inhibition

    37) Product Images from "Resistance to Store Depletion-induced Endothelial Injury in Rat Lung after Chronic Heart Failure"

    Article Title: Resistance to Store Depletion-induced Endothelial Injury in Rat Lung after Chronic Heart Failure

    Journal:

    doi: 10.1164/rccm.200506-847OC

    Immunostaining for transient receptor potential proteins TRPC1, TRPC3, TRPC4, and TRPC6/7 in rat lungs. Expression of each TRPC isoform in endothelium was confirmed in small muscular or partially muscularized extraalveolar vessels ( A ) and conduit artery
    Figure Legend Snippet: Immunostaining for transient receptor potential proteins TRPC1, TRPC3, TRPC4, and TRPC6/7 in rat lungs. Expression of each TRPC isoform in endothelium was confirmed in small muscular or partially muscularized extraalveolar vessels ( A ) and conduit artery

    Techniques Used: Immunostaining, Expressing

    38) Product Images from "Resistance to Store Depletion-induced Endothelial Injury in Rat Lung after Chronic Heart Failure"

    Article Title: Resistance to Store Depletion-induced Endothelial Injury in Rat Lung after Chronic Heart Failure

    Journal:

    doi: 10.1164/rccm.200506-847OC

    Immunostaining for transient receptor potential proteins TRPC1, TRPC3, TRPC4, and TRPC6/7 in rat lungs. Expression of each TRPC isoform in endothelium was confirmed in small muscular or partially muscularized extraalveolar vessels ( A ) and conduit artery
    Figure Legend Snippet: Immunostaining for transient receptor potential proteins TRPC1, TRPC3, TRPC4, and TRPC6/7 in rat lungs. Expression of each TRPC isoform in endothelium was confirmed in small muscular or partially muscularized extraalveolar vessels ( A ) and conduit artery

    Techniques Used: Immunostaining, Expressing

    39) Product Images from "The involvement of TRPC3 channels in sinoatrial arrhythmias"

    Article Title: The involvement of TRPC3 channels in sinoatrial arrhythmias

    Journal: Frontiers in Physiology

    doi: 10.3389/fphys.2015.00086

    The effect of angiotension II (Ang II) and a selective TRPC3 channel blocker Pyr10 on pacemaker action potential after application of Ang II. (A–D) Intracellular recordings from WT mice; (E–H ) Intracellular recordings from TRPC3 −/− mice with conditions as indicated in each panel.
    Figure Legend Snippet: The effect of angiotension II (Ang II) and a selective TRPC3 channel blocker Pyr10 on pacemaker action potential after application of Ang II. (A–D) Intracellular recordings from WT mice; (E–H ) Intracellular recordings from TRPC3 −/− mice with conditions as indicated in each panel.

    Techniques Used: Mouse Assay

    Atrial arrhythmias induced by Ang II and pacing . Reconstructed excitation maps of data from multi-electrode array recordings (images below) from a Langendorff-perfused isolated heart. (A) sinus rhythm and (B) pacing induced atrial tachycardia. Note in AT, the ectopic beat starts from a different site to sinus rhythms. (C) Representative ECG recordings from isolated heart from WT and TRPC3 −/− mice under control conditions. (D) Atrial tachycardia and atrial fibrillation induced by burst pacing in WT mice and atrial-ventricular conduction block recorded from a TRPC −/− mouse heart respectively. (E) On average, the arrhythmia index was significant reduced in TRPC3KO mice ( n = 8) compared to WT mice ( n = 11, P = 0.004).
    Figure Legend Snippet: Atrial arrhythmias induced by Ang II and pacing . Reconstructed excitation maps of data from multi-electrode array recordings (images below) from a Langendorff-perfused isolated heart. (A) sinus rhythm and (B) pacing induced atrial tachycardia. Note in AT, the ectopic beat starts from a different site to sinus rhythms. (C) Representative ECG recordings from isolated heart from WT and TRPC3 −/− mice under control conditions. (D) Atrial tachycardia and atrial fibrillation induced by burst pacing in WT mice and atrial-ventricular conduction block recorded from a TRPC −/− mouse heart respectively. (E) On average, the arrhythmia index was significant reduced in TRPC3KO mice ( n = 8) compared to WT mice ( n = 11, P = 0.004).

    Techniques Used: Isolation, Mouse Assay, Blocking Assay

    Isolated single SAN cells express TRPC3, STIM1, and IP 3 R2 . Confocal immunofluorenscence images of isolated single pacemaker cells. Isolated group (A) and single (B) pacemaker cells were positively stained with anti HCN4 (in green). (C) Anti-TRPC3 in red, anti-STIM1 in green. (D) Anti-IP3R2 in red, anti-STIM1 in green. (E) 3D reconstruction image using N-SIM microscopy. Anti-IP 3 R2 in red, anti-STIM1 in green. Areas of co-localisation appear yellow due to color mixing.
    Figure Legend Snippet: Isolated single SAN cells express TRPC3, STIM1, and IP 3 R2 . Confocal immunofluorenscence images of isolated single pacemaker cells. Isolated group (A) and single (B) pacemaker cells were positively stained with anti HCN4 (in green). (C) Anti-TRPC3 in red, anti-STIM1 in green. (D) Anti-IP3R2 in red, anti-STIM1 in green. (E) 3D reconstruction image using N-SIM microscopy. Anti-IP 3 R2 in red, anti-STIM1 in green. Areas of co-localisation appear yellow due to color mixing.

    Techniques Used: Isolation, Staining, Microscopy

    The effect of Pyr10 on intracellular Ca 2+ after application of 1-oleoy1-2-acyl-sn-glycerol (OAG) 100 μM . The intact SANs were loaded with Ca 2+ indicator indo-1. (A,B) show intracellular Ca 2+ recordings from a WT mouse. (B,C) The statistics pool data shows percentage changes in resting Ca 2+ , Ca 2+ transient and firing rate in WT and TRPC3 KO mice respectively. (B) OAG treatment against control. (C) 2 μM Pyr10 treatment against OAG treatment. * P
    Figure Legend Snippet: The effect of Pyr10 on intracellular Ca 2+ after application of 1-oleoy1-2-acyl-sn-glycerol (OAG) 100 μM . The intact SANs were loaded with Ca 2+ indicator indo-1. (A,B) show intracellular Ca 2+ recordings from a WT mouse. (B,C) The statistics pool data shows percentage changes in resting Ca 2+ , Ca 2+ transient and firing rate in WT and TRPC3 KO mice respectively. (B) OAG treatment against control. (C) 2 μM Pyr10 treatment against OAG treatment. * P

    Techniques Used: Mouse Assay

    Signaling pathways involved in the activation of TRPC3 . The G-protein coupled receptor (GPCRs) activates phospholiase C (PLC), resulting in generation of IP3 and diacylglycerol (DAG). IP3 activates its receptor that leads to Ca 2+ release from SR/ER and the depletion of SR/ER Ca 2+ store. The Ca 2+ content change in the store can be sensed by STIM1, the ER Ca 2+ sensor and cause store-operated Ca 2+ entry (SOCE) through TRPC3 channels. An additional or alternate possibility is that DAG can directly activate TRPC3, and produce receptor-operated Ca 2+ entry (ROCE). GPCRs, G-protein coupled receptors; IP3R, inositol 1,4,5-trisphosphate receptors; PLC, Phospholipase C; ROCE, Receptor–operated Ca 2+ entry; SOCE, store-operated Ca 2+ entry; SR/ER, sarco- endo-plasmic reticulum; STIM1, stromal interacting molecule 1; TRPC3,4,6 canonical transient receptor potential channel types 3,4,6; NCX Na/Ca exchange.
    Figure Legend Snippet: Signaling pathways involved in the activation of TRPC3 . The G-protein coupled receptor (GPCRs) activates phospholiase C (PLC), resulting in generation of IP3 and diacylglycerol (DAG). IP3 activates its receptor that leads to Ca 2+ release from SR/ER and the depletion of SR/ER Ca 2+ store. The Ca 2+ content change in the store can be sensed by STIM1, the ER Ca 2+ sensor and cause store-operated Ca 2+ entry (SOCE) through TRPC3 channels. An additional or alternate possibility is that DAG can directly activate TRPC3, and produce receptor-operated Ca 2+ entry (ROCE). GPCRs, G-protein coupled receptors; IP3R, inositol 1,4,5-trisphosphate receptors; PLC, Phospholipase C; ROCE, Receptor–operated Ca 2+ entry; SOCE, store-operated Ca 2+ entry; SR/ER, sarco- endo-plasmic reticulum; STIM1, stromal interacting molecule 1; TRPC3,4,6 canonical transient receptor potential channel types 3,4,6; NCX Na/Ca exchange.

    Techniques Used: Activation Assay, Planar Chromatography

    40) Product Images from "The involvement of TRPC3 channels in sinoatrial arrhythmias"

    Article Title: The involvement of TRPC3 channels in sinoatrial arrhythmias

    Journal: Frontiers in Physiology

    doi: 10.3389/fphys.2015.00086

    The effect of angiotension II (Ang II) and a selective TRPC3 channel blocker Pyr10 on pacemaker action potential after application of Ang II. (A–D) Intracellular recordings from WT mice; (E–H ) Intracellular recordings from TRPC3 −/− mice with conditions as indicated in each panel.
    Figure Legend Snippet: The effect of angiotension II (Ang II) and a selective TRPC3 channel blocker Pyr10 on pacemaker action potential after application of Ang II. (A–D) Intracellular recordings from WT mice; (E–H ) Intracellular recordings from TRPC3 −/− mice with conditions as indicated in each panel.

    Techniques Used: Mouse Assay

    Atrial arrhythmias induced by Ang II and pacing . Reconstructed excitation maps of data from multi-electrode array recordings (images below) from a Langendorff-perfused isolated heart. (A) sinus rhythm and (B) pacing induced atrial tachycardia. Note in AT, the ectopic beat starts from a different site to sinus rhythms. (C) Representative ECG recordings from isolated heart from WT and TRPC3 −/− mice under control conditions. (D) Atrial tachycardia and atrial fibrillation induced by burst pacing in WT mice and atrial-ventricular conduction block recorded from a TRPC −/− mouse heart respectively. (E) On average, the arrhythmia index was significant reduced in TRPC3KO mice ( n = 8) compared to WT mice ( n = 11, P = 0.004).
    Figure Legend Snippet: Atrial arrhythmias induced by Ang II and pacing . Reconstructed excitation maps of data from multi-electrode array recordings (images below) from a Langendorff-perfused isolated heart. (A) sinus rhythm and (B) pacing induced atrial tachycardia. Note in AT, the ectopic beat starts from a different site to sinus rhythms. (C) Representative ECG recordings from isolated heart from WT and TRPC3 −/− mice under control conditions. (D) Atrial tachycardia and atrial fibrillation induced by burst pacing in WT mice and atrial-ventricular conduction block recorded from a TRPC −/− mouse heart respectively. (E) On average, the arrhythmia index was significant reduced in TRPC3KO mice ( n = 8) compared to WT mice ( n = 11, P = 0.004).

    Techniques Used: Isolation, Mouse Assay, Blocking Assay

    Isolated single SAN cells express TRPC3, STIM1, and IP 3 R2 . Confocal immunofluorenscence images of isolated single pacemaker cells. Isolated group (A) and single (B) pacemaker cells were positively stained with anti HCN4 (in green). (C) Anti-TRPC3 in red, anti-STIM1 in green. (D) Anti-IP3R2 in red, anti-STIM1 in green. (E) 3D reconstruction image using N-SIM microscopy. Anti-IP 3 R2 in red, anti-STIM1 in green. Areas of co-localisation appear yellow due to color mixing.
    Figure Legend Snippet: Isolated single SAN cells express TRPC3, STIM1, and IP 3 R2 . Confocal immunofluorenscence images of isolated single pacemaker cells. Isolated group (A) and single (B) pacemaker cells were positively stained with anti HCN4 (in green). (C) Anti-TRPC3 in red, anti-STIM1 in green. (D) Anti-IP3R2 in red, anti-STIM1 in green. (E) 3D reconstruction image using N-SIM microscopy. Anti-IP 3 R2 in red, anti-STIM1 in green. Areas of co-localisation appear yellow due to color mixing.

    Techniques Used: Isolation, Staining, Microscopy

    The effect of Pyr10 on intracellular Ca 2+ after application of 1-oleoy1-2-acyl-sn-glycerol (OAG) 100 μM . The intact SANs were loaded with Ca 2+ indicator indo-1. (A,B) show intracellular Ca 2+ recordings from a WT mouse. (B,C) The statistics pool data shows percentage changes in resting Ca 2+ , Ca 2+ transient and firing rate in WT and TRPC3 KO mice respectively. (B) OAG treatment against control. (C) 2 μM Pyr10 treatment against OAG treatment. * P
    Figure Legend Snippet: The effect of Pyr10 on intracellular Ca 2+ after application of 1-oleoy1-2-acyl-sn-glycerol (OAG) 100 μM . The intact SANs were loaded with Ca 2+ indicator indo-1. (A,B) show intracellular Ca 2+ recordings from a WT mouse. (B,C) The statistics pool data shows percentage changes in resting Ca 2+ , Ca 2+ transient and firing rate in WT and TRPC3 KO mice respectively. (B) OAG treatment against control. (C) 2 μM Pyr10 treatment against OAG treatment. * P

    Techniques Used: Mouse Assay

    Signaling pathways involved in the activation of TRPC3 . The G-protein coupled receptor (GPCRs) activates phospholiase C (PLC), resulting in generation of IP3 and diacylglycerol (DAG). IP3 activates its receptor that leads to Ca 2+ release from SR/ER and the depletion of SR/ER Ca 2+ store. The Ca 2+ content change in the store can be sensed by STIM1, the ER Ca 2+ sensor and cause store-operated Ca 2+ entry (SOCE) through TRPC3 channels. An additional or alternate possibility is that DAG can directly activate TRPC3, and produce receptor-operated Ca 2+ entry (ROCE). GPCRs, G-protein coupled receptors; IP3R, inositol 1,4,5-trisphosphate receptors; PLC, Phospholipase C; ROCE, Receptor–operated Ca 2+ entry; SOCE, store-operated Ca 2+ entry; SR/ER, sarco- endo-plasmic reticulum; STIM1, stromal interacting molecule 1; TRPC3,4,6 canonical transient receptor potential channel types 3,4,6; NCX Na/Ca exchange.
    Figure Legend Snippet: Signaling pathways involved in the activation of TRPC3 . The G-protein coupled receptor (GPCRs) activates phospholiase C (PLC), resulting in generation of IP3 and diacylglycerol (DAG). IP3 activates its receptor that leads to Ca 2+ release from SR/ER and the depletion of SR/ER Ca 2+ store. The Ca 2+ content change in the store can be sensed by STIM1, the ER Ca 2+ sensor and cause store-operated Ca 2+ entry (SOCE) through TRPC3 channels. An additional or alternate possibility is that DAG can directly activate TRPC3, and produce receptor-operated Ca 2+ entry (ROCE). GPCRs, G-protein coupled receptors; IP3R, inositol 1,4,5-trisphosphate receptors; PLC, Phospholipase C; ROCE, Receptor–operated Ca 2+ entry; SOCE, store-operated Ca 2+ entry; SR/ER, sarco- endo-plasmic reticulum; STIM1, stromal interacting molecule 1; TRPC3,4,6 canonical transient receptor potential channel types 3,4,6; NCX Na/Ca exchange.

    Techniques Used: Activation Assay, Planar Chromatography

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    Alomone Labs trpc3
    The effect of angiotension II (Ang II) and a selective <t>TRPC3</t> channel blocker Pyr10 on pacemaker action potential after application of Ang II. (A–D) Intracellular recordings from WT mice; (E–H ) Intracellular recordings from TRPC3 −/− mice with conditions as indicated in each panel.
    Trpc3, supplied by Alomone Labs, used in various techniques. Bioz Stars score: 91/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/trpc3/product/Alomone Labs
    Average 91 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    trpc3 - by Bioz Stars, 2022-09
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    Alomone Labs anti trpc3
    Schematic model of <t>TRPC3</t> transmembrane domains and protein binding sites. Predicted transmembrane domains, calcium entry pore, IP 3 R, and PLCγ SH2 binding sites and deleted and substituted sites in TRPC3-DEL and TRPC3-SUB are shown. aa , amino
    Anti Trpc3, supplied by Alomone Labs, used in various techniques. Bioz Stars score: 95/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/anti trpc3/product/Alomone Labs
    Average 95 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    anti trpc3 - by Bioz Stars, 2022-09
    95/100 stars
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    The effect of angiotension II (Ang II) and a selective TRPC3 channel blocker Pyr10 on pacemaker action potential after application of Ang II. (A–D) Intracellular recordings from WT mice; (E–H ) Intracellular recordings from TRPC3 −/− mice with conditions as indicated in each panel.

    Journal: Frontiers in Physiology

    Article Title: The involvement of TRPC3 channels in sinoatrial arrhythmias

    doi: 10.3389/fphys.2015.00086

    Figure Lengend Snippet: The effect of angiotension II (Ang II) and a selective TRPC3 channel blocker Pyr10 on pacemaker action potential after application of Ang II. (A–D) Intracellular recordings from WT mice; (E–H ) Intracellular recordings from TRPC3 −/− mice with conditions as indicated in each panel.

    Article Snippet: To investigate this possibility, we examined expression and molecular localisation of STIM1, and the possibility of molecular interaction between TRPC3, STIM1, and IP3R2 in isolated cardiac pacemaker cells.

    Techniques: Mouse Assay

    Atrial arrhythmias induced by Ang II and pacing . Reconstructed excitation maps of data from multi-electrode array recordings (images below) from a Langendorff-perfused isolated heart. (A) sinus rhythm and (B) pacing induced atrial tachycardia. Note in AT, the ectopic beat starts from a different site to sinus rhythms. (C) Representative ECG recordings from isolated heart from WT and TRPC3 −/− mice under control conditions. (D) Atrial tachycardia and atrial fibrillation induced by burst pacing in WT mice and atrial-ventricular conduction block recorded from a TRPC −/− mouse heart respectively. (E) On average, the arrhythmia index was significant reduced in TRPC3KO mice ( n = 8) compared to WT mice ( n = 11, P = 0.004).

    Journal: Frontiers in Physiology

    Article Title: The involvement of TRPC3 channels in sinoatrial arrhythmias

    doi: 10.3389/fphys.2015.00086

    Figure Lengend Snippet: Atrial arrhythmias induced by Ang II and pacing . Reconstructed excitation maps of data from multi-electrode array recordings (images below) from a Langendorff-perfused isolated heart. (A) sinus rhythm and (B) pacing induced atrial tachycardia. Note in AT, the ectopic beat starts from a different site to sinus rhythms. (C) Representative ECG recordings from isolated heart from WT and TRPC3 −/− mice under control conditions. (D) Atrial tachycardia and atrial fibrillation induced by burst pacing in WT mice and atrial-ventricular conduction block recorded from a TRPC −/− mouse heart respectively. (E) On average, the arrhythmia index was significant reduced in TRPC3KO mice ( n = 8) compared to WT mice ( n = 11, P = 0.004).

    Article Snippet: To investigate this possibility, we examined expression and molecular localisation of STIM1, and the possibility of molecular interaction between TRPC3, STIM1, and IP3R2 in isolated cardiac pacemaker cells.

    Techniques: Isolation, Mouse Assay, Blocking Assay

    Isolated single SAN cells express TRPC3, STIM1, and IP 3 R2 . Confocal immunofluorenscence images of isolated single pacemaker cells. Isolated group (A) and single (B) pacemaker cells were positively stained with anti HCN4 (in green). (C) Anti-TRPC3 in red, anti-STIM1 in green. (D) Anti-IP3R2 in red, anti-STIM1 in green. (E) 3D reconstruction image using N-SIM microscopy. Anti-IP 3 R2 in red, anti-STIM1 in green. Areas of co-localisation appear yellow due to color mixing.

    Journal: Frontiers in Physiology

    Article Title: The involvement of TRPC3 channels in sinoatrial arrhythmias

    doi: 10.3389/fphys.2015.00086

    Figure Lengend Snippet: Isolated single SAN cells express TRPC3, STIM1, and IP 3 R2 . Confocal immunofluorenscence images of isolated single pacemaker cells. Isolated group (A) and single (B) pacemaker cells were positively stained with anti HCN4 (in green). (C) Anti-TRPC3 in red, anti-STIM1 in green. (D) Anti-IP3R2 in red, anti-STIM1 in green. (E) 3D reconstruction image using N-SIM microscopy. Anti-IP 3 R2 in red, anti-STIM1 in green. Areas of co-localisation appear yellow due to color mixing.

    Article Snippet: To investigate this possibility, we examined expression and molecular localisation of STIM1, and the possibility of molecular interaction between TRPC3, STIM1, and IP3R2 in isolated cardiac pacemaker cells.

    Techniques: Isolation, Staining, Microscopy

    The effect of Pyr10 on intracellular Ca 2+ after application of 1-oleoy1-2-acyl-sn-glycerol (OAG) 100 μM . The intact SANs were loaded with Ca 2+ indicator indo-1. (A,B) show intracellular Ca 2+ recordings from a WT mouse. (B,C) The statistics pool data shows percentage changes in resting Ca 2+ , Ca 2+ transient and firing rate in WT and TRPC3 KO mice respectively. (B) OAG treatment against control. (C) 2 μM Pyr10 treatment against OAG treatment. * P

    Journal: Frontiers in Physiology

    Article Title: The involvement of TRPC3 channels in sinoatrial arrhythmias

    doi: 10.3389/fphys.2015.00086

    Figure Lengend Snippet: The effect of Pyr10 on intracellular Ca 2+ after application of 1-oleoy1-2-acyl-sn-glycerol (OAG) 100 μM . The intact SANs were loaded with Ca 2+ indicator indo-1. (A,B) show intracellular Ca 2+ recordings from a WT mouse. (B,C) The statistics pool data shows percentage changes in resting Ca 2+ , Ca 2+ transient and firing rate in WT and TRPC3 KO mice respectively. (B) OAG treatment against control. (C) 2 μM Pyr10 treatment against OAG treatment. * P

    Article Snippet: To investigate this possibility, we examined expression and molecular localisation of STIM1, and the possibility of molecular interaction between TRPC3, STIM1, and IP3R2 in isolated cardiac pacemaker cells.

    Techniques: Mouse Assay

    Signaling pathways involved in the activation of TRPC3 . The G-protein coupled receptor (GPCRs) activates phospholiase C (PLC), resulting in generation of IP3 and diacylglycerol (DAG). IP3 activates its receptor that leads to Ca 2+ release from SR/ER and the depletion of SR/ER Ca 2+ store. The Ca 2+ content change in the store can be sensed by STIM1, the ER Ca 2+ sensor and cause store-operated Ca 2+ entry (SOCE) through TRPC3 channels. An additional or alternate possibility is that DAG can directly activate TRPC3, and produce receptor-operated Ca 2+ entry (ROCE). GPCRs, G-protein coupled receptors; IP3R, inositol 1,4,5-trisphosphate receptors; PLC, Phospholipase C; ROCE, Receptor–operated Ca 2+ entry; SOCE, store-operated Ca 2+ entry; SR/ER, sarco- endo-plasmic reticulum; STIM1, stromal interacting molecule 1; TRPC3,4,6 canonical transient receptor potential channel types 3,4,6; NCX Na/Ca exchange.

    Journal: Frontiers in Physiology

    Article Title: The involvement of TRPC3 channels in sinoatrial arrhythmias

    doi: 10.3389/fphys.2015.00086

    Figure Lengend Snippet: Signaling pathways involved in the activation of TRPC3 . The G-protein coupled receptor (GPCRs) activates phospholiase C (PLC), resulting in generation of IP3 and diacylglycerol (DAG). IP3 activates its receptor that leads to Ca 2+ release from SR/ER and the depletion of SR/ER Ca 2+ store. The Ca 2+ content change in the store can be sensed by STIM1, the ER Ca 2+ sensor and cause store-operated Ca 2+ entry (SOCE) through TRPC3 channels. An additional or alternate possibility is that DAG can directly activate TRPC3, and produce receptor-operated Ca 2+ entry (ROCE). GPCRs, G-protein coupled receptors; IP3R, inositol 1,4,5-trisphosphate receptors; PLC, Phospholipase C; ROCE, Receptor–operated Ca 2+ entry; SOCE, store-operated Ca 2+ entry; SR/ER, sarco- endo-plasmic reticulum; STIM1, stromal interacting molecule 1; TRPC3,4,6 canonical transient receptor potential channel types 3,4,6; NCX Na/Ca exchange.

    Article Snippet: To investigate this possibility, we examined expression and molecular localisation of STIM1, and the possibility of molecular interaction between TRPC3, STIM1, and IP3R2 in isolated cardiac pacemaker cells.

    Techniques: Activation Assay, Planar Chromatography

    Schematic model of TRPC3 transmembrane domains and protein binding sites. Predicted transmembrane domains, calcium entry pore, IP 3 R, and PLCγ SH2 binding sites and deleted and substituted sites in TRPC3-DEL and TRPC3-SUB are shown. aa , amino

    Journal: The Journal of Biological Chemistry

    Article Title: TRPC3 Is the Erythropoietin-regulated Calcium Channel in Human Erythroid Cells *

    doi: 10.1074/jbc.M710231200

    Figure Lengend Snippet: Schematic model of TRPC3 transmembrane domains and protein binding sites. Predicted transmembrane domains, calcium entry pore, IP 3 R, and PLCγ SH2 binding sites and deleted and substituted sites in TRPC3-DEL and TRPC3-SUB are shown. aa , amino

    Article Snippet: Sample buffer (3×) was added to the pellets, and the samples were heated at 60 °C for 30 min. Western blotting was performed as described above, and blots were probed with anti-V5-HRP or anti-Epo-R, anti-PLCγ1, anti-IP3 R type II, anti-TRPC3, or anti-actin antibodies, followed by the appropriate HRP-conjugated secondary antibodies and ECL.

    Techniques: Protein Binding, Binding Assay

    Association of TRPC3 IP 3 R binding mutants with IP 3 R. IP 3 R type II and V5-TRPC3, V5-TRPC3-DEL, or V5-TRPC3-SUB were expressed in HEK 293T cells. Immunoprecipitation ( IP ) was performed on lysates with anti-V5 or anti-IP 3 R antibodies or normal rabbit

    Journal: The Journal of Biological Chemistry

    Article Title: TRPC3 Is the Erythropoietin-regulated Calcium Channel in Human Erythroid Cells *

    doi: 10.1074/jbc.M710231200

    Figure Lengend Snippet: Association of TRPC3 IP 3 R binding mutants with IP 3 R. IP 3 R type II and V5-TRPC3, V5-TRPC3-DEL, or V5-TRPC3-SUB were expressed in HEK 293T cells. Immunoprecipitation ( IP ) was performed on lysates with anti-V5 or anti-IP 3 R antibodies or normal rabbit

    Article Snippet: Sample buffer (3×) was added to the pellets, and the samples were heated at 60 °C for 30 min. Western blotting was performed as described above, and blots were probed with anti-V5-HRP or anti-Epo-R, anti-PLCγ1, anti-IP3 R type II, anti-TRPC3, or anti-actin antibodies, followed by the appropriate HRP-conjugated secondary antibodies and ECL.

    Techniques: Binding Assay, Immunoprecipitation

    Dose response and time course of [Ca 2+ ] i after Epo stimulation of HEK 293T cells transfected with TRPC3 and Epo-R. A , Epo dose response. HEK 293T cells transfected with TRPC3 and Epo-R were stimulated with 0–40 units/ml Epo. [Ca 2+ ] i was measured

    Journal: The Journal of Biological Chemistry

    Article Title: TRPC3 Is the Erythropoietin-regulated Calcium Channel in Human Erythroid Cells *

    doi: 10.1074/jbc.M710231200

    Figure Lengend Snippet: Dose response and time course of [Ca 2+ ] i after Epo stimulation of HEK 293T cells transfected with TRPC3 and Epo-R. A , Epo dose response. HEK 293T cells transfected with TRPC3 and Epo-R were stimulated with 0–40 units/ml Epo. [Ca 2+ ] i was measured

    Article Snippet: Sample buffer (3×) was added to the pellets, and the samples were heated at 60 °C for 30 min. Western blotting was performed as described above, and blots were probed with anti-V5-HRP or anti-Epo-R, anti-PLCγ1, anti-IP3 R type II, anti-TRPC3, or anti-actin antibodies, followed by the appropriate HRP-conjugated secondary antibodies and ECL.

    Techniques: Transfection

    Requirement for external calcium in the Epo-stimulated calcium increase in HEK 293T cells. Fura Red-loaded HEK 293T cells were transfected with BFP-TRPC3 and Epo-R. A , cells were treated with 40 units/ml Epo in the presence (0.68 m m ) or absence (2

    Journal: The Journal of Biological Chemistry

    Article Title: TRPC3 Is the Erythropoietin-regulated Calcium Channel in Human Erythroid Cells *

    doi: 10.1074/jbc.M710231200

    Figure Lengend Snippet: Requirement for external calcium in the Epo-stimulated calcium increase in HEK 293T cells. Fura Red-loaded HEK 293T cells were transfected with BFP-TRPC3 and Epo-R. A , cells were treated with 40 units/ml Epo in the presence (0.68 m m ) or absence (2

    Article Snippet: Sample buffer (3×) was added to the pellets, and the samples were heated at 60 °C for 30 min. Western blotting was performed as described above, and blots were probed with anti-V5-HRP or anti-Epo-R, anti-PLCγ1, anti-IP3 R type II, anti-TRPC3, or anti-actin antibodies, followed by the appropriate HRP-conjugated secondary antibodies and ECL.

    Techniques: Transfection

    Interaction of TRPC3 with PLCγ SH2 binding site substitutions with PLCγ. PLCγ and V5-TRPC3, V5-TRPC3-F4, or V5-TRPC3-Y226F were expressed in HEK 293T cells. Immunoprecipitation ( IP ) was performed on lysates with anti-PLCγ

    Journal: The Journal of Biological Chemistry

    Article Title: TRPC3 Is the Erythropoietin-regulated Calcium Channel in Human Erythroid Cells *

    doi: 10.1074/jbc.M710231200

    Figure Lengend Snippet: Interaction of TRPC3 with PLCγ SH2 binding site substitutions with PLCγ. PLCγ and V5-TRPC3, V5-TRPC3-F4, or V5-TRPC3-Y226F were expressed in HEK 293T cells. Immunoprecipitation ( IP ) was performed on lysates with anti-PLCγ

    Article Snippet: Sample buffer (3×) was added to the pellets, and the samples were heated at 60 °C for 30 min. Western blotting was performed as described above, and blots were probed with anti-V5-HRP or anti-Epo-R, anti-PLCγ1, anti-IP3 R type II, anti-TRPC3, or anti-actin antibodies, followed by the appropriate HRP-conjugated secondary antibodies and ECL.

    Techniques: Binding Assay, Immunoprecipitation

    Western blot of HEK 293T cells transfected with siRNA targeted to PLC γ. Lysates were prepared from HEK 293T cells transfected ( Tx ' d ) with or without BFP-TRPC3 and Epo-R, and siRNA was targeted to PLCγ or control siRNA. Blots were probed

    Journal: The Journal of Biological Chemistry

    Article Title: TRPC3 Is the Erythropoietin-regulated Calcium Channel in Human Erythroid Cells *

    doi: 10.1074/jbc.M710231200

    Figure Lengend Snippet: Western blot of HEK 293T cells transfected with siRNA targeted to PLC γ. Lysates were prepared from HEK 293T cells transfected ( Tx ' d ) with or without BFP-TRPC3 and Epo-R, and siRNA was targeted to PLCγ or control siRNA. Blots were probed

    Article Snippet: Sample buffer (3×) was added to the pellets, and the samples were heated at 60 °C for 30 min. Western blotting was performed as described above, and blots were probed with anti-V5-HRP or anti-Epo-R, anti-PLCγ1, anti-IP3 R type II, anti-TRPC3, or anti-actin antibodies, followed by the appropriate HRP-conjugated secondary antibodies and ECL.

    Techniques: Western Blot, Transfection, Planar Chromatography

    Endogenous expression of TRPC3 in human hematopoietic cells. Western blotting was performed on lysates from UT-7 and TF-1 Epo-responsive cell lines, from CD34 + cells and from day 10 and 14 BFU-E-derived erythroblasts. Equivalent amounts of protein

    Journal: The Journal of Biological Chemistry

    Article Title: TRPC3 Is the Erythropoietin-regulated Calcium Channel in Human Erythroid Cells *

    doi: 10.1074/jbc.M710231200

    Figure Lengend Snippet: Endogenous expression of TRPC3 in human hematopoietic cells. Western blotting was performed on lysates from UT-7 and TF-1 Epo-responsive cell lines, from CD34 + cells and from day 10 and 14 BFU-E-derived erythroblasts. Equivalent amounts of protein

    Article Snippet: Sample buffer (3×) was added to the pellets, and the samples were heated at 60 °C for 30 min. Western blotting was performed as described above, and blots were probed with anti-V5-HRP or anti-Epo-R, anti-PLCγ1, anti-IP3 R type II, anti-TRPC3, or anti-actin antibodies, followed by the appropriate HRP-conjugated secondary antibodies and ECL.

    Techniques: Expressing, Western Blot, Derivative Assay

    Plasma membrane externalization of TRPC3 detected by cell surface biotinylation. Cell surface biotinylation was performed with HEK 293T cells expressing V5-TRPC3, V5-TRPC3-DEL, V5-TRPC3-SUB, or V5-TRPC3-F4 and Epo-R. Lysates were prepared, and immunoprecipitation

    Journal: The Journal of Biological Chemistry

    Article Title: TRPC3 Is the Erythropoietin-regulated Calcium Channel in Human Erythroid Cells *

    doi: 10.1074/jbc.M710231200

    Figure Lengend Snippet: Plasma membrane externalization of TRPC3 detected by cell surface biotinylation. Cell surface biotinylation was performed with HEK 293T cells expressing V5-TRPC3, V5-TRPC3-DEL, V5-TRPC3-SUB, or V5-TRPC3-F4 and Epo-R. Lysates were prepared, and immunoprecipitation

    Article Snippet: Sample buffer (3×) was added to the pellets, and the samples were heated at 60 °C for 30 min. Western blotting was performed as described above, and blots were probed with anti-V5-HRP or anti-Epo-R, anti-PLCγ1, anti-IP3 R type II, anti-TRPC3, or anti-actin antibodies, followed by the appropriate HRP-conjugated secondary antibodies and ECL.

    Techniques: Expressing, Immunoprecipitation

    Association of TRPC3 and TRPC3-F4 with PLCγ or Epo-R. A , PLCγ and V5-TRPC3 or V5-TRPC3-F4 were expressed in HEK 293T cells. Immunoprecipitation ( IP ) was performed on lysates with anti-PLCγ or anti-V5 antibodies or normal rabbit

    Journal: The Journal of Biological Chemistry

    Article Title: TRPC3 Is the Erythropoietin-regulated Calcium Channel in Human Erythroid Cells *

    doi: 10.1074/jbc.M710231200

    Figure Lengend Snippet: Association of TRPC3 and TRPC3-F4 with PLCγ or Epo-R. A , PLCγ and V5-TRPC3 or V5-TRPC3-F4 were expressed in HEK 293T cells. Immunoprecipitation ( IP ) was performed on lysates with anti-PLCγ or anti-V5 antibodies or normal rabbit

    Article Snippet: Sample buffer (3×) was added to the pellets, and the samples were heated at 60 °C for 30 min. Western blotting was performed as described above, and blots were probed with anti-V5-HRP or anti-Epo-R, anti-PLCγ1, anti-IP3 R type II, anti-TRPC3, or anti-actin antibodies, followed by the appropriate HRP-conjugated secondary antibodies and ECL.

    Techniques: Immunoprecipitation

    A reduction in Ca 2+ transients in response to membrane depolarization, and the disruption of the binding between endogenous MG29 and TRPC3 in mouse primary skeletal myotubes expressing Δ116-MG29

    Journal: Biochemical and biophysical research communications

    Article Title: Interaction between mitsugumin 29 and TRPC3 participates in regulating Ca2+ transients in skeletal muscle

    doi: 10.1016/j.bbrc.2015.06.096

    Figure Lengend Snippet: A reduction in Ca 2+ transients in response to membrane depolarization, and the disruption of the binding between endogenous MG29 and TRPC3 in mouse primary skeletal myotubes expressing Δ116-MG29

    Article Snippet: For immunoblot assays, various antibodies were used: anti-RyR1, anti-DHPR, anti-SERCA1a, anti-MG29, anti-JP1, anti-JP2, and anti-GST antibodies (1:1,000) from Thermo Scientific Inc. (Rockford, IL, USA), anti-TRPC3 and anti-TRPC4 antibodies (1:800) from Alomone Laboratories (Jerusalem 9104201, Israel), and anti-Orai1, anti-STIM1, and anti-α-actin antibodies (1:1,000) from Abcam (Cambridge, MA, USA).

    Techniques: Binding Assay, Expressing

    Co-immunoprecipitation of TRPC3 with each MG29 portion

    Journal: Biochemical and biophysical research communications

    Article Title: Interaction between mitsugumin 29 and TRPC3 participates in regulating Ca2+ transients in skeletal muscle

    doi: 10.1016/j.bbrc.2015.06.096

    Figure Lengend Snippet: Co-immunoprecipitation of TRPC3 with each MG29 portion

    Article Snippet: For immunoblot assays, various antibodies were used: anti-RyR1, anti-DHPR, anti-SERCA1a, anti-MG29, anti-JP1, anti-JP2, and anti-GST antibodies (1:1,000) from Thermo Scientific Inc. (Rockford, IL, USA), anti-TRPC3 and anti-TRPC4 antibodies (1:800) from Alomone Laboratories (Jerusalem 9104201, Israel), and anti-Orai1, anti-STIM1, and anti-α-actin antibodies (1:1,000) from Abcam (Cambridge, MA, USA).

    Techniques: Immunoprecipitation

    TRPC3 forms a stable ternary complex with Nox2 and p22 phox . ( a , b ) Expression of Nox2 and p22 phox proteins in HEK293 cells that express a different combination of TRPC3-GFP and GFP. Results of a quantitative analysis are shown in ( b ) (n = 3). ( c ) Nox2 mRNA amounts in HEK293 cells co-expressing Nox2 with GFP or TRPC3-GFP (n = 3). ( d ) Increased Nox2 and p22 phox protein in HEK293 cells co-expressing pore-dead mutant of TRPC3 (n = 3). ( e ) Interaction of TRPC3 with Nox2 in HEK293 cells. Immunoprecipitation was performed using an anti-flag antibody. ( f ) Nox2 protein expression in HEK293 cells expressing TRPC3 alone or co-expressing TRPC3 and TRPC6 (n = 3). Error bars, s.e.m. *P

    Journal: Scientific Reports

    Article Title: TRPC3 positively regulates reactive oxygen species driving maladaptive cardiac remodeling

    doi: 10.1038/srep37001

    Figure Lengend Snippet: TRPC3 forms a stable ternary complex with Nox2 and p22 phox . ( a , b ) Expression of Nox2 and p22 phox proteins in HEK293 cells that express a different combination of TRPC3-GFP and GFP. Results of a quantitative analysis are shown in ( b ) (n = 3). ( c ) Nox2 mRNA amounts in HEK293 cells co-expressing Nox2 with GFP or TRPC3-GFP (n = 3). ( d ) Increased Nox2 and p22 phox protein in HEK293 cells co-expressing pore-dead mutant of TRPC3 (n = 3). ( e ) Interaction of TRPC3 with Nox2 in HEK293 cells. Immunoprecipitation was performed using an anti-flag antibody. ( f ) Nox2 protein expression in HEK293 cells expressing TRPC3 alone or co-expressing TRPC3 and TRPC6 (n = 3). Error bars, s.e.m. *P

    Article Snippet: The following primary antibodies were used: GAPDH (sc-25778), gp91phox (sc-130543), p47phox (sc-17845) and p22phox (sc-20781) from Santa Cruz Biotechnology, flag M2-HRP (A8592) from Sigma Aldrich, GFP (CHIP grade, ab290) from Abcam, myc-tag (05–742) from Merck, phospho p47phox (p-Ser370) (A1171) from Assay Bio Tech, and TRPC3 (ACC-016) from Alomone Labs.

    Techniques: Expressing, Mutagenesis, Immunoprecipitation

    TRPC3 plays a critical role in Mechanical stretch-induced ROS production. ( a , b ) Effects of siRNA targeting TRPC1, C3 or C6 on mechanical stretch (MS)-induced ROS production (n = 3). ( c ) mRNA expression of either TRPC1 or TRPC6 in NRCM transfected with siRNAs against either TRPC1 or TRPC6, respectively (n = 3). ( d,e ) Time courses of MS-induced ROS production in NRCMs treated with GsMTx-4 (1 μM; ( d ) or TRPV4 inhibitor (RN1734, 50 μM; ( e ) Reagents were added to cells 5 min before MS (n = 3). ( f ) MS-induced ROS production in TRPC(1–7)-deficient MEF cells expressing TRPC3, TRPC6, TRPC7, or LacZ (n = 30). Data are representative of three independent experiments. ( g–i ) Effect of TRPC3 siRNA on the protein abundances of TRPC3 ( h ) and Nox2 ( i ) protein expressions in NRCMs (n = 3). Error bars, s.e.m. *P

    Journal: Scientific Reports

    Article Title: TRPC3 positively regulates reactive oxygen species driving maladaptive cardiac remodeling

    doi: 10.1038/srep37001

    Figure Lengend Snippet: TRPC3 plays a critical role in Mechanical stretch-induced ROS production. ( a , b ) Effects of siRNA targeting TRPC1, C3 or C6 on mechanical stretch (MS)-induced ROS production (n = 3). ( c ) mRNA expression of either TRPC1 or TRPC6 in NRCM transfected with siRNAs against either TRPC1 or TRPC6, respectively (n = 3). ( d,e ) Time courses of MS-induced ROS production in NRCMs treated with GsMTx-4 (1 μM; ( d ) or TRPV4 inhibitor (RN1734, 50 μM; ( e ) Reagents were added to cells 5 min before MS (n = 3). ( f ) MS-induced ROS production in TRPC(1–7)-deficient MEF cells expressing TRPC3, TRPC6, TRPC7, or LacZ (n = 30). Data are representative of three independent experiments. ( g–i ) Effect of TRPC3 siRNA on the protein abundances of TRPC3 ( h ) and Nox2 ( i ) protein expressions in NRCMs (n = 3). Error bars, s.e.m. *P

    Article Snippet: The following primary antibodies were used: GAPDH (sc-25778), gp91phox (sc-130543), p47phox (sc-17845) and p22phox (sc-20781) from Santa Cruz Biotechnology, flag M2-HRP (A8592) from Sigma Aldrich, GFP (CHIP grade, ab290) from Abcam, myc-tag (05–742) from Merck, phospho p47phox (p-Ser370) (A1171) from Assay Bio Tech, and TRPC3 (ACC-016) from Alomone Labs.

    Techniques: Mass Spectrometry, Expressing, Transfection

    TRPC3 prevents Nox2 protein from proteasomal degradation. ( a–e ) Abundances of Nox2 protein ( a , b ) and mRNAs of TRPC3 ( c ), Nox2 ( d ), and p22 phox ( e ) in NRCM transfected with siRNAs targeting TRPC3 with or without MG132. Cells were treated with siRNAs and MG132 (1 μM) simultaneously (n = 3). ( f , g ) Effect of siRNA targeting TRPC3 on Nox2 protein abundance in cell surface (Surface) and total lysates (Total) from NRCMs (n = 3). GAPDH was used as an internal control. Error bars, s.e.m. *P

    Journal: Scientific Reports

    Article Title: TRPC3 positively regulates reactive oxygen species driving maladaptive cardiac remodeling

    doi: 10.1038/srep37001

    Figure Lengend Snippet: TRPC3 prevents Nox2 protein from proteasomal degradation. ( a–e ) Abundances of Nox2 protein ( a , b ) and mRNAs of TRPC3 ( c ), Nox2 ( d ), and p22 phox ( e ) in NRCM transfected with siRNAs targeting TRPC3 with or without MG132. Cells were treated with siRNAs and MG132 (1 μM) simultaneously (n = 3). ( f , g ) Effect of siRNA targeting TRPC3 on Nox2 protein abundance in cell surface (Surface) and total lysates (Total) from NRCMs (n = 3). GAPDH was used as an internal control. Error bars, s.e.m. *P

    Article Snippet: The following primary antibodies were used: GAPDH (sc-25778), gp91phox (sc-130543), p47phox (sc-17845) and p22phox (sc-20781) from Santa Cruz Biotechnology, flag M2-HRP (A8592) from Sigma Aldrich, GFP (CHIP grade, ab290) from Abcam, myc-tag (05–742) from Merck, phospho p47phox (p-Ser370) (A1171) from Assay Bio Tech, and TRPC3 (ACC-016) from Alomone Labs.

    Techniques: Transfection

    TRPC3 forms a stable ternary complex with Nox2 and p22 phox proteins in endogenously p22 phox -absent CHO cells. ( a ) Expression of Nox2 and p22 phox proteins in CHO cells that express a different combination of TRPC3-GFP and GFP. ( b ) Results of quantitative analysis (n = 3). ( c ) Expression of Nox2 and p22 phox co-expressed with either GFP or TRPC3-GFP in MG132 (10 μM)-treated CHO cells. ( d ) Graphs depict the relative expression of either Nox2 or p22 phox protein to that in non-treated cells. Band intensities were normalized by GAPDH. ( e–g ) Interaction of TRPC3 with p22 phox and Nox2 in CHO cells. ( h ) Localization of Nox2 in CHO cells co-expressing Nox2 with TRPC3-GFP (or GFP-F). Error bars, s.e.m. *P

    Journal: Scientific Reports

    Article Title: TRPC3 positively regulates reactive oxygen species driving maladaptive cardiac remodeling

    doi: 10.1038/srep37001

    Figure Lengend Snippet: TRPC3 forms a stable ternary complex with Nox2 and p22 phox proteins in endogenously p22 phox -absent CHO cells. ( a ) Expression of Nox2 and p22 phox proteins in CHO cells that express a different combination of TRPC3-GFP and GFP. ( b ) Results of quantitative analysis (n = 3). ( c ) Expression of Nox2 and p22 phox co-expressed with either GFP or TRPC3-GFP in MG132 (10 μM)-treated CHO cells. ( d ) Graphs depict the relative expression of either Nox2 or p22 phox protein to that in non-treated cells. Band intensities were normalized by GAPDH. ( e–g ) Interaction of TRPC3 with p22 phox and Nox2 in CHO cells. ( h ) Localization of Nox2 in CHO cells co-expressing Nox2 with TRPC3-GFP (or GFP-F). Error bars, s.e.m. *P

    Article Snippet: The following primary antibodies were used: GAPDH (sc-25778), gp91phox (sc-130543), p47phox (sc-17845) and p22phox (sc-20781) from Santa Cruz Biotechnology, flag M2-HRP (A8592) from Sigma Aldrich, GFP (CHIP grade, ab290) from Abcam, myc-tag (05–742) from Merck, phospho p47phox (p-Ser370) (A1171) from Assay Bio Tech, and TRPC3 (ACC-016) from Alomone Labs.

    Techniques: Expressing

    Formation of a TRPC3/Nox2 complex promotes TRPC3 channel activity through stabilization at the plasma membrane. ( a ) Effect of Nox2 siRNA on expression of TRPC3 in NRCMs (n = 3). ( b ) Representative images showing the levels of TRPC3-GFP and GFP expression in HEK293 cells co-expressing p22 phox or Nox2 (n = 3). ( c ) Expression of TRPC3-GFP mRNA in HEK293 cells co-expressing p22 phox or Nox2 (n = 3). ( d–f ) Representative time courses of TRPC3 currents ( d ) and the current-voltage (I-V) relationships ( e ) and peak TRPC3 current densities at −60 mV ( f ) induced by 100 μM carbachol (CCh) in HEK293 cells expressing TRPC3-mCherry alone or with p22 phox , Nox2, both p22 phox and Nox2, or Nox2 treated with DPI. DPI (0.3 μM) was treated 1 min before CCh stimulation. ( g , h ) Representative Ca 2+ responses in the presence ( g ) or absence ( h ) of pyrazole-3 (Pyr3, 1 μM) upon mechanical stretch (MS) application. ( i ) Peak Ca 2+ increases after MS in NRCMs treated with (n = 61) or without Pyr3 (n = 78). ( j ) Changes of minimal [Ca 2+ ] i before and after MS application. Minimal [Ca 2+ ] i from Ca 2+ responses in every 1 min were analyzed and represented as diastolic [Ca 2+ ] i . ( k ) Schematic images showing phosphorylation of p47 phox via TRPC3-PKCβ activation induced by MS in the heart. ( l–n ) Effects of TRPC3 ( l , m ) or PKCβ ( n) ; 10 μM Gö6976) inhibitors on p47 phox phosphorylation induced by MS in NRCMs (n = 3). ( o ) MS-induced ROS generation in NRCMs treated with a PKCβ inhibitor (n = 3). ( p ) Co-immunoprecipitation of TRPC3 with PKCβ, Nox2 and p22 phox in mouse hearts 1week after TAC operation (n = 3). Error bars, s.e.m.*P

    Journal: Scientific Reports

    Article Title: TRPC3 positively regulates reactive oxygen species driving maladaptive cardiac remodeling

    doi: 10.1038/srep37001

    Figure Lengend Snippet: Formation of a TRPC3/Nox2 complex promotes TRPC3 channel activity through stabilization at the plasma membrane. ( a ) Effect of Nox2 siRNA on expression of TRPC3 in NRCMs (n = 3). ( b ) Representative images showing the levels of TRPC3-GFP and GFP expression in HEK293 cells co-expressing p22 phox or Nox2 (n = 3). ( c ) Expression of TRPC3-GFP mRNA in HEK293 cells co-expressing p22 phox or Nox2 (n = 3). ( d–f ) Representative time courses of TRPC3 currents ( d ) and the current-voltage (I-V) relationships ( e ) and peak TRPC3 current densities at −60 mV ( f ) induced by 100 μM carbachol (CCh) in HEK293 cells expressing TRPC3-mCherry alone or with p22 phox , Nox2, both p22 phox and Nox2, or Nox2 treated with DPI. DPI (0.3 μM) was treated 1 min before CCh stimulation. ( g , h ) Representative Ca 2+ responses in the presence ( g ) or absence ( h ) of pyrazole-3 (Pyr3, 1 μM) upon mechanical stretch (MS) application. ( i ) Peak Ca 2+ increases after MS in NRCMs treated with (n = 61) or without Pyr3 (n = 78). ( j ) Changes of minimal [Ca 2+ ] i before and after MS application. Minimal [Ca 2+ ] i from Ca 2+ responses in every 1 min were analyzed and represented as diastolic [Ca 2+ ] i . ( k ) Schematic images showing phosphorylation of p47 phox via TRPC3-PKCβ activation induced by MS in the heart. ( l–n ) Effects of TRPC3 ( l , m ) or PKCβ ( n) ; 10 μM Gö6976) inhibitors on p47 phox phosphorylation induced by MS in NRCMs (n = 3). ( o ) MS-induced ROS generation in NRCMs treated with a PKCβ inhibitor (n = 3). ( p ) Co-immunoprecipitation of TRPC3 with PKCβ, Nox2 and p22 phox in mouse hearts 1week after TAC operation (n = 3). Error bars, s.e.m.*P

    Article Snippet: The following primary antibodies were used: GAPDH (sc-25778), gp91phox (sc-130543), p47phox (sc-17845) and p22phox (sc-20781) from Santa Cruz Biotechnology, flag M2-HRP (A8592) from Sigma Aldrich, GFP (CHIP grade, ab290) from Abcam, myc-tag (05–742) from Merck, phospho p47phox (p-Ser370) (A1171) from Assay Bio Tech, and TRPC3 (ACC-016) from Alomone Labs.

    Techniques: Activity Assay, Expressing, Mass Spectrometry, Activation Assay, Immunoprecipitation

    TRPC3 deletion suppresses TAC-induced LV dysfunction and dilation through Nox2 inhibition. ( a ) Left ventricular end-diastolic pressure (LVEDP; left) and dP/dT max (right) in TAC-operated TRPC3 (+/+) (n = 13) and TRPC3 (−/−) (n = 12) mice 6 week post-operation. ( b ) Myocardial malondialdehyde concentrations 1 week after TAC (n = 4). ( c ) Abundance of Nox2 protein in TRPC3 (+/+) and TRPC3 (−/−) hearts 1 week after TAC (n = 3). ( d ) Representative immunofluorescence images of TRPC3, p22 phox , and caveolin-3 (Cav-3) in adult mouse cardiomyocytes isolated from muscle LIM protein-deficient hearts. ( e ) Representative immunofluorescence images of p22 phox in adult mouse cardiomyocytes: green, anti-p22 phox ; blue, DAPI. ( f ) Relative abundances of p22 phox and Nox2 mRNA in mouse hearts 1 week after TAC (n = 4). ( g ) Abundance of Nox2 protein in TRPC6 (+/+) and TRPC6 (−/−) hearts 1 week after TAC (n = 3). Error bars, s.e.m. *P

    Journal: Scientific Reports

    Article Title: TRPC3 positively regulates reactive oxygen species driving maladaptive cardiac remodeling

    doi: 10.1038/srep37001

    Figure Lengend Snippet: TRPC3 deletion suppresses TAC-induced LV dysfunction and dilation through Nox2 inhibition. ( a ) Left ventricular end-diastolic pressure (LVEDP; left) and dP/dT max (right) in TAC-operated TRPC3 (+/+) (n = 13) and TRPC3 (−/−) (n = 12) mice 6 week post-operation. ( b ) Myocardial malondialdehyde concentrations 1 week after TAC (n = 4). ( c ) Abundance of Nox2 protein in TRPC3 (+/+) and TRPC3 (−/−) hearts 1 week after TAC (n = 3). ( d ) Representative immunofluorescence images of TRPC3, p22 phox , and caveolin-3 (Cav-3) in adult mouse cardiomyocytes isolated from muscle LIM protein-deficient hearts. ( e ) Representative immunofluorescence images of p22 phox in adult mouse cardiomyocytes: green, anti-p22 phox ; blue, DAPI. ( f ) Relative abundances of p22 phox and Nox2 mRNA in mouse hearts 1 week after TAC (n = 4). ( g ) Abundance of Nox2 protein in TRPC6 (+/+) and TRPC6 (−/−) hearts 1 week after TAC (n = 3). Error bars, s.e.m. *P

    Article Snippet: The following primary antibodies were used: GAPDH (sc-25778), gp91phox (sc-130543), p47phox (sc-17845) and p22phox (sc-20781) from Santa Cruz Biotechnology, flag M2-HRP (A8592) from Sigma Aldrich, GFP (CHIP grade, ab290) from Abcam, myc-tag (05–742) from Merck, phospho p47phox (p-Ser370) (A1171) from Assay Bio Tech, and TRPC3 (ACC-016) from Alomone Labs.

    Techniques: Inhibition, Mouse Assay, Immunofluorescence, Isolation

    Physical interaction between TRPC3 and Nox2 is critical for stabilization of Nox2. ( a ) Schematic illustration of TRPC3 terminal deletion mutants. ( b , c ) Expression of Nox2 and p22 phox co-expressed with TRPC3 deletion mutants in HEK293 cells (n = 3). ( d ) OAG-induced ROS production in NRCMs expressing Nox2-interacting TRPC3 C-terminal fragment (C3-C fragment) (n = 20–28). ( e ) Co-immunoprecipitation of TRPC3 with Nox2 in the presence or absence of C3-C fragment. Representative blot from three independent experiments was shown. ( f ) ATP (100 μM)-induced Ca 2+ responses in HEK293 cells expressing TRPC3 with or without C3-C fragment (n = 35–51). Timing of solution exchanges were indicated by horizontal bars above the graph. ( g ) Model of the regulation of TRPC3-Nox2 stability and induction of LV dysfunction induced by diastolic stretch of cardiomyocytes. Error bars, s.e.m.*P

    Journal: Scientific Reports

    Article Title: TRPC3 positively regulates reactive oxygen species driving maladaptive cardiac remodeling

    doi: 10.1038/srep37001

    Figure Lengend Snippet: Physical interaction between TRPC3 and Nox2 is critical for stabilization of Nox2. ( a ) Schematic illustration of TRPC3 terminal deletion mutants. ( b , c ) Expression of Nox2 and p22 phox co-expressed with TRPC3 deletion mutants in HEK293 cells (n = 3). ( d ) OAG-induced ROS production in NRCMs expressing Nox2-interacting TRPC3 C-terminal fragment (C3-C fragment) (n = 20–28). ( e ) Co-immunoprecipitation of TRPC3 with Nox2 in the presence or absence of C3-C fragment. Representative blot from three independent experiments was shown. ( f ) ATP (100 μM)-induced Ca 2+ responses in HEK293 cells expressing TRPC3 with or without C3-C fragment (n = 35–51). Timing of solution exchanges were indicated by horizontal bars above the graph. ( g ) Model of the regulation of TRPC3-Nox2 stability and induction of LV dysfunction induced by diastolic stretch of cardiomyocytes. Error bars, s.e.m.*P

    Article Snippet: The following primary antibodies were used: GAPDH (sc-25778), gp91phox (sc-130543), p47phox (sc-17845) and p22phox (sc-20781) from Santa Cruz Biotechnology, flag M2-HRP (A8592) from Sigma Aldrich, GFP (CHIP grade, ab290) from Abcam, myc-tag (05–742) from Merck, phospho p47phox (p-Ser370) (A1171) from Assay Bio Tech, and TRPC3 (ACC-016) from Alomone Labs.

    Techniques: Expressing, Immunoprecipitation