trpc4  (Alomone Labs)


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    Name:
    Anti TRPC4 Antibody
    Description:
    Anti TRPC4 Antibody ACC 018 is a highly specific antibody directed against an epitope of the mouse protein The antibody can be used in western blot immunoprecipitation immunohistochemistry and immunocytochemistry applications It has been designed to recognize TRPC4 from human mouse and rat samples
    Catalog Number:
    ACC-018
    Price:
    397.0
    Category:
    Primary Antibody
    Applications:
    Immunocytochemistry, Immunofluorescence, Immunohistochemistry, Immunoprecipitation, Western Blot
    Purity:
    Affinity purified on immobilized antigen.
    Immunogen:
    Synthetic peptide
    Size:
    25 mcl
    Antibody Type:
    Polyclonal Primary Antibodies
    Format:
    Lyophilized Powder
    Host:
    Rabbit
    Isotype:
    Rabbit IgG
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    Structured Review

    Alomone Labs trpc4
    Anti TRPC4 Antibody
    Anti TRPC4 Antibody ACC 018 is a highly specific antibody directed against an epitope of the mouse protein The antibody can be used in western blot immunoprecipitation immunohistochemistry and immunocytochemistry applications It has been designed to recognize TRPC4 from human mouse and rat samples
    https://www.bioz.com/result/trpc4/product/Alomone Labs
    Average 93 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    trpc4 - by Bioz Stars, 2021-09
    93/100 stars

    Images

    1) 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

    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

    2) Product Images from "Calcium Sensing Receptor Modulates Extracellular Calcium Entry and Proliferation via TRPC3/6 Channels in Cultured Human Mesangial Cells"

    Article Title: Calcium Sensing Receptor Modulates Extracellular Calcium Entry and Proliferation via TRPC3/6 Channels in Cultured Human Mesangial Cells

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0098777

    The influence of CaSR activation on TRPC mRNA levels and protein expression. Human MCs were starved for 24-free medium prior to stimulation with 1 mM (Ctl) or 5 mM [Ca 2+ ] o for 24 h. (A) Real-time PCR experiments showed that the administration of 5 mM [Ca 2+ ] o for 24 h significantly increased the TRPC3 and TRPC6 mRNA levels but did not affect the mRNA levels of TRPC1 or TRPC4 (**p
    Figure Legend Snippet: The influence of CaSR activation on TRPC mRNA levels and protein expression. Human MCs were starved for 24-free medium prior to stimulation with 1 mM (Ctl) or 5 mM [Ca 2+ ] o for 24 h. (A) Real-time PCR experiments showed that the administration of 5 mM [Ca 2+ ] o for 24 h significantly increased the TRPC3 and TRPC6 mRNA levels but did not affect the mRNA levels of TRPC1 or TRPC4 (**p

    Techniques Used: Activation Assay, Expressing, CTL Assay, Real-time Polymerase Chain Reaction

    3) Product Images from "Cell-Cell Contact Formation Governs Ca2+ Signaling by TRPC4 in the Vascular Endothelium"

    Article Title: Cell-Cell Contact Formation Governs Ca2+ Signaling by TRPC4 in the Vascular Endothelium

    Journal: The Journal of Biological Chemistry

    doi: 10.1074/jbc.M109.060301

    Membrane presentation of TRPC4 is enhanced by formation of cell-cell contacts and divergently affected by EGF in subconfluent and confluent HMEC-1. A , top , proteins from total cell lysates ( lanes 1 and 4 ), biotinylated fractions ( lanes 2 , 3 , 5 , and 6 ), and non-biotinylated fractions ( lane 7 ) at the indicated conditions were subjected to SDS-PAGE and immunoblotted with TRPC4 antibody. Results are representative of four individual experiments. Bottom , mean values ± S.E. of TRPC4 immunoreactivity detected in biotinylated fractions before and after stimulation with EGF (100 ng/ml, 20 min). *, statistically significant differences between stimulated and unstimulated conditions ( p
    Figure Legend Snippet: Membrane presentation of TRPC4 is enhanced by formation of cell-cell contacts and divergently affected by EGF in subconfluent and confluent HMEC-1. A , top , proteins from total cell lysates ( lanes 1 and 4 ), biotinylated fractions ( lanes 2 , 3 , 5 , and 6 ), and non-biotinylated fractions ( lane 7 ) at the indicated conditions were subjected to SDS-PAGE and immunoblotted with TRPC4 antibody. Results are representative of four individual experiments. Bottom , mean values ± S.E. of TRPC4 immunoreactivity detected in biotinylated fractions before and after stimulation with EGF (100 ng/ml, 20 min). *, statistically significant differences between stimulated and unstimulated conditions ( p

    Techniques Used: SDS Page

    Association of VE-cadherin and β-catenin with TRPC4 depends on cell-cell contact formation. A , top , TRPC4 was detected in total cell lysates (from the left , lanes 1 and 4 ) immunocomplexes precipitated with anti-VE-cadherin ( lanes 2 , 3 , 5 , and 6 ) and lysates precipitated only with beads ( lane 7 ). Results are a representative of four experiments. Bottom , mean values ± S.E. of the TRPC4 immunoreactivity detected in precipitates obtained from cells before and after stimulation with EGF (100 ng/ml, 20 min). B , top , TRPC4 was detected in total cell lysates (from the left , lanes 1 and 4 ), immunocomplexes precipitated with anti-β-catenin ( lanes 2 , 3 , 5 , and 6 ), and lysates precipitated only with beads ( lane 7 ). Results are a representative of four experiments. Bottom , mean values ± S.E. of the TRPC4 immunoreactivity detected in precipitates obtained from cells before and after stimulation with EGF (100 ng/ml, 20 min). * and #, statistically significant differences ( p
    Figure Legend Snippet: Association of VE-cadherin and β-catenin with TRPC4 depends on cell-cell contact formation. A , top , TRPC4 was detected in total cell lysates (from the left , lanes 1 and 4 ) immunocomplexes precipitated with anti-VE-cadherin ( lanes 2 , 3 , 5 , and 6 ) and lysates precipitated only with beads ( lane 7 ). Results are a representative of four experiments. Bottom , mean values ± S.E. of the TRPC4 immunoreactivity detected in precipitates obtained from cells before and after stimulation with EGF (100 ng/ml, 20 min). B , top , TRPC4 was detected in total cell lysates (from the left , lanes 1 and 4 ), immunocomplexes precipitated with anti-β-catenin ( lanes 2 , 3 , 5 , and 6 ), and lysates precipitated only with beads ( lane 7 ). Results are a representative of four experiments. Bottom , mean values ± S.E. of the TRPC4 immunoreactivity detected in precipitates obtained from cells before and after stimulation with EGF (100 ng/ml, 20 min). * and #, statistically significant differences ( p

    Techniques Used:

    Cell-cell contact formation and β-catenin expression determines the cellular targeting of TRPC4. Fluorescence images of single ( top images ) or contact-forming ( bottom images ) HMEC-1 ( A and B ) and HEK293 ( C and D ) cells transfected with either CFP-TRPC4 ( red ) or GFP-β-catenin ( green ) alone ( A and C ) or double transfected with both constructs ( B and D ). The arrows indicate enhanced fluorescence of CFP-TRPC4 or GFP-β-catenin, respectively. A , fluorescence images of single transfected ( single TF ) HMEC-1 cells. B , fluorescence images of double transfected ( double TF ) HMEC-1 cells. C , fluorescence images of single transfected HEK293-cells. D , fluorescence images of double transfected HEK293 cells. Images from cells expressing GFP and CFP have been normalized for channel bleed-through by linear spectral unmixing. Scale bars , 10 μm. For B and D , relative values of fluorescence overlap as a measure for co-localization are shown for defined cellular regions (total cell, nuclear region, and cell-cell contact area). The columns represent the overlap of the two fluorophores (percentage of pixels) within selected cellular areas (mean ± S.E.).
    Figure Legend Snippet: Cell-cell contact formation and β-catenin expression determines the cellular targeting of TRPC4. Fluorescence images of single ( top images ) or contact-forming ( bottom images ) HMEC-1 ( A and B ) and HEK293 ( C and D ) cells transfected with either CFP-TRPC4 ( red ) or GFP-β-catenin ( green ) alone ( A and C ) or double transfected with both constructs ( B and D ). The arrows indicate enhanced fluorescence of CFP-TRPC4 or GFP-β-catenin, respectively. A , fluorescence images of single transfected ( single TF ) HMEC-1 cells. B , fluorescence images of double transfected ( double TF ) HMEC-1 cells. C , fluorescence images of single transfected HEK293-cells. D , fluorescence images of double transfected HEK293 cells. Images from cells expressing GFP and CFP have been normalized for channel bleed-through by linear spectral unmixing. Scale bars , 10 μm. For B and D , relative values of fluorescence overlap as a measure for co-localization are shown for defined cellular regions (total cell, nuclear region, and cell-cell contact area). The columns represent the overlap of the two fluorophores (percentage of pixels) within selected cellular areas (mean ± S.E.).

    Techniques Used: Expressing, Fluorescence, Transfection, Construct

    EGF-induced Ca 2+ entry is altered by endothelial phenotype transitions. A , representative records of fura-2 fluorescence ratio in single (migrating state; open circles ), subconfluent contact-forming (proliferating clusters; closed circles ), and confluent (quiescent, barrier-forming; squares ) cells during a Ca 2+ readdition after stimulation with EGF (100 ng/ml; arrow ). Ca 2+ readdition (elevation of extracellular Ca 2+ from nominally free to 2 m m ) is indicated. Basal Ca 2+ entry into non-stimulated cells is shown for proliferating clusters (unstimulated). The inset shows inhibition of Ca 2+ entry into EGF-stimulated subconfluent cells by TRPC4 siRNA silencing. Responses of a TRPC4-silenced cell ( filled circles ) and a control (scrambled; open circles ) are shown. B , mean peak increases in fura-2 emission ratios (Δ values ± S.E., n ≥ 45) obtained during Ca 2+ readdition. C , mean rates of Ca 2+ -sensitive fluorescence changes during the initial phase of reentry. *, statistically significant difference ( p
    Figure Legend Snippet: EGF-induced Ca 2+ entry is altered by endothelial phenotype transitions. A , representative records of fura-2 fluorescence ratio in single (migrating state; open circles ), subconfluent contact-forming (proliferating clusters; closed circles ), and confluent (quiescent, barrier-forming; squares ) cells during a Ca 2+ readdition after stimulation with EGF (100 ng/ml; arrow ). Ca 2+ readdition (elevation of extracellular Ca 2+ from nominally free to 2 m m ) is indicated. Basal Ca 2+ entry into non-stimulated cells is shown for proliferating clusters (unstimulated). The inset shows inhibition of Ca 2+ entry into EGF-stimulated subconfluent cells by TRPC4 siRNA silencing. Responses of a TRPC4-silenced cell ( filled circles ) and a control (scrambled; open circles ) are shown. B , mean peak increases in fura-2 emission ratios (Δ values ± S.E., n ≥ 45) obtained during Ca 2+ readdition. C , mean rates of Ca 2+ -sensitive fluorescence changes during the initial phase of reentry. *, statistically significant difference ( p

    Techniques Used: Fluorescence, Inhibition

    β-catenin enables cell-cell contact-dependent promotion of Ca 2+ entry into TRPC4-expressing HEK293 cells. Ca 2+ entry into thrombin-stimulated HEK293 cells during a typical Ca 2+ readdition protocol. A , representative recordings of fura-2 fluorescence ratios from single or contact-forming HEK293 cells stably expressing TRPC4 (T4-60), transfected with either pcDNA3 (control) or β-catenin. Ca 2+ readdition (from nominally free to 2 m m ) was performed as indicated after a 5-min stimulation with thrombin (0.5 unit/ml; pretreatment). Shown are mean peak increases in fura-2 fluorescence ratios (Δ values ± S.E.) obtained during Ca 2+ readdition to single or contact-forming T4-60 ( B ) as well as HEK293 cells ( C ). *, statistical significance ( p
    Figure Legend Snippet: β-catenin enables cell-cell contact-dependent promotion of Ca 2+ entry into TRPC4-expressing HEK293 cells. Ca 2+ entry into thrombin-stimulated HEK293 cells during a typical Ca 2+ readdition protocol. A , representative recordings of fura-2 fluorescence ratios from single or contact-forming HEK293 cells stably expressing TRPC4 (T4-60), transfected with either pcDNA3 (control) or β-catenin. Ca 2+ readdition (from nominally free to 2 m m ) was performed as indicated after a 5-min stimulation with thrombin (0.5 unit/ml; pretreatment). Shown are mean peak increases in fura-2 fluorescence ratios (Δ values ± S.E.) obtained during Ca 2+ readdition to single or contact-forming T4-60 ( B ) as well as HEK293 cells ( C ). *, statistical significance ( p

    Techniques Used: Expressing, Fluorescence, Stable Transfection, Transfection

    Proposed model of phenotype-dependent TRPC4 function in growth factor-stimulated endothelial cells via interaction of the channel with β-catenin and targeting into cell-cell contacts. TRPC4 is for a large part sequestered in intracellular compartments and unavailable for Ca 2+ signaling in single cells ( contact deficient ; upper left ). By contrast, formation of immature cell adhesions promotes surface targeting of β-catenin-TRPC4 complexes and enables further recruitment of channels into the plasma membrane and Ca 2+ entry function ( immature contact ; upper right ). Once mature barriers are formed ( mature contact ; lower panel ), TRPC4 resides for a large part in junctional complexes that are rapidly retrieved from the cell surface during growth factor stimulation and are barely available for contribution to global Ca 2+ signaling.
    Figure Legend Snippet: Proposed model of phenotype-dependent TRPC4 function in growth factor-stimulated endothelial cells via interaction of the channel with β-catenin and targeting into cell-cell contacts. TRPC4 is for a large part sequestered in intracellular compartments and unavailable for Ca 2+ signaling in single cells ( contact deficient ; upper left ). By contrast, formation of immature cell adhesions promotes surface targeting of β-catenin-TRPC4 complexes and enables further recruitment of channels into the plasma membrane and Ca 2+ entry function ( immature contact ; upper right ). Once mature barriers are formed ( mature contact ; lower panel ), TRPC4 resides for a large part in junctional complexes that are rapidly retrieved from the cell surface during growth factor stimulation and are barely available for contribution to global Ca 2+ signaling.

    Techniques Used:

    Interaction between TRPC4 and VE-cadherin within cell-cell contacts in the HEK293 expression system. This figure shows donor (CFP), acceptor (YFP), and raw FRET images as well as a color-coded map of FRET intensity (FRET index) after background subtraction. A , FRET analysis of CFP-TRPC4 and GFP-β-catenin transiently expressed in HEK293 cells. B , FRET analysis of CFP-TRPC4 and YFP-VE-cadherin transiently expressed in HEK293 cells. The arrows indicate enhanced FRET signal at cell-cell contact positions and cell-cell junctions. Images are representative of 14–19 individual FRET measurements.
    Figure Legend Snippet: Interaction between TRPC4 and VE-cadherin within cell-cell contacts in the HEK293 expression system. This figure shows donor (CFP), acceptor (YFP), and raw FRET images as well as a color-coded map of FRET intensity (FRET index) after background subtraction. A , FRET analysis of CFP-TRPC4 and GFP-β-catenin transiently expressed in HEK293 cells. B , FRET analysis of CFP-TRPC4 and YFP-VE-cadherin transiently expressed in HEK293 cells. The arrows indicate enhanced FRET signal at cell-cell contact positions and cell-cell junctions. Images are representative of 14–19 individual FRET measurements.

    Techniques Used: Expressing

    Formation of immature cell-cell contacts in proliferating HMEC-1 enables TRPC4-dependent Ca 2+ entry. A , Ca 2+ entry into thrombin or EGF-stimulated single HMEC-1 cells. Left , representative recordings of fluo-4 fluorescence intensity in single cells transfected with pcDNA3-vector (control; open circles ), TRPC4 ( closed circles ), or a functional dominant negative TRPC4 fragment (DNTRPC; squares ). Time courses during acute stimulation with 0.5 unit/ml thrombin ( arrow ) and Ca 2+ readdition (2 m m ) are shown. Center , mean values of peak fluo-4 intensity ( n > 6; open columns , control; hatched columns , TRPC4; filled columns , DNTRPC). Right , mean increases in fura-2 fluorescence ratios ( n ≥ 6) obtained during Ca 2+ readdition after stimulation with EGF (100 ng/ml). B , left , representative recordings of fura-2 fluorescence ratios in the subconfluent, contact-forming state is shown for controls (sham-transfected; open circles ) and DNTRPC-transfected ( closed circles ) cells, HMEC-1 cells were challenged with EGF (100 ng/ml; arrow ), as indicated. Right , increases in fura-2 fluorescence ratios (mean ± S.E.; n ≥ 8). C , left , representative recordings of fura-2 fluorescence ratios in the confluent, barrier-forming state is shown for controls (sham-transfected; open circles ) and DNTRPC-transfected ( closed circles ) cells, and HMEC-1 cells were challenged with EGF (100 ng/ml; arrow ), as indicated. Right , increases in fura-2 fluorescence ratios (mean ± S.E.; n ≥ 8).
    Figure Legend Snippet: Formation of immature cell-cell contacts in proliferating HMEC-1 enables TRPC4-dependent Ca 2+ entry. A , Ca 2+ entry into thrombin or EGF-stimulated single HMEC-1 cells. Left , representative recordings of fluo-4 fluorescence intensity in single cells transfected with pcDNA3-vector (control; open circles ), TRPC4 ( closed circles ), or a functional dominant negative TRPC4 fragment (DNTRPC; squares ). Time courses during acute stimulation with 0.5 unit/ml thrombin ( arrow ) and Ca 2+ readdition (2 m m ) are shown. Center , mean values of peak fluo-4 intensity ( n > 6; open columns , control; hatched columns , TRPC4; filled columns , DNTRPC). Right , mean increases in fura-2 fluorescence ratios ( n ≥ 6) obtained during Ca 2+ readdition after stimulation with EGF (100 ng/ml). B , left , representative recordings of fura-2 fluorescence ratios in the subconfluent, contact-forming state is shown for controls (sham-transfected; open circles ) and DNTRPC-transfected ( closed circles ) cells, HMEC-1 cells were challenged with EGF (100 ng/ml; arrow ), as indicated. Right , increases in fura-2 fluorescence ratios (mean ± S.E.; n ≥ 8). C , left , representative recordings of fura-2 fluorescence ratios in the confluent, barrier-forming state is shown for controls (sham-transfected; open circles ) and DNTRPC-transfected ( closed circles ) cells, and HMEC-1 cells were challenged with EGF (100 ng/ml; arrow ), as indicated. Right , increases in fura-2 fluorescence ratios (mean ± S.E.; n ≥ 8).

    Techniques Used: Fluorescence, Transfection, Plasmid Preparation, Functional Assay, Dominant Negative Mutation

    4) 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

    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

    Related Articles

    Incubation:

    Article Title: TRPC1 mediates slow excitatory synaptic transmission in hippocampal oriens/alveus interneurons
    Article Snippet: .. Sections were incubated 4 days at 4 °C in a mixture of mouse monoclonal anti-mGluR1a (1/100 BD BioSciences, Oakville, ON, Canada) and rabbit polyclonal anti-TRPC1 (1/250) or rabbit anti-TRPC4 (1/100; Alomone Labs, Jerusalem, Israel). ..

    Article Title: Sensory neuron-expressed transient receptor potential channel 4 is a target for the relief of psoriasiform itch and skin inflammation in mice.
    Article Snippet: .. After blocking with 2% BSA, the membranes were incubated overnight at 4 ºC with TRPC4 antibody (1:1000, cat. no. ACC-018, Alomone, Jerusalem, Israel) and glyceraldehyde 3-phosphate dehydrogenase (GAPDH) antibody (1:1000, cat. no. 3683S, Cell Signaling, Danvers, MA), and then incubated with the appropriate HRPconjugated secondary antibody (1:2000, Cell Signaling). ..

    Article Title: Sensory neuron-expressed transient receptor potential channel 4 is a target for the relief of psoriasiform itch and skin inflammation in mice.
    Article Snippet: .. Tissue sections were cut in a cryostat (12 µm), blocked with BlockAidTM blocking solution (cat. no. B10710, Fisher Scientific), and incubated overnight at 4 °C with the following primary antibodies or reagents: TRPC4 (1:1000, cat. no. OST00025W, Fisher Scientific), TRPC4 (1:200, cat. no. ACC-018, Alomone), calcitonin gene-related peptide (CGRP, 1:500, cat. no. ab36001, Abcam, Cambridge, MA), IB4 (1:500, cat. no. I32450, Fisher Scientific) and/or 4’-6-diamidino-2phenylindole dihydrochloride (DAPI; 300 nmol/L, cat. no. D1306, Fisher Scientific). ..

    Blocking Assay:

    Article Title: Sensory neuron-expressed transient receptor potential channel 4 is a target for the relief of psoriasiform itch and skin inflammation in mice.
    Article Snippet: .. After blocking with 2% BSA, the membranes were incubated overnight at 4 ºC with TRPC4 antibody (1:1000, cat. no. ACC-018, Alomone, Jerusalem, Israel) and glyceraldehyde 3-phosphate dehydrogenase (GAPDH) antibody (1:1000, cat. no. 3683S, Cell Signaling, Danvers, MA), and then incubated with the appropriate HRPconjugated secondary antibody (1:2000, Cell Signaling). ..

    Article Title: Sensory neuron-expressed transient receptor potential channel 4 is a target for the relief of psoriasiform itch and skin inflammation in mice.
    Article Snippet: .. Tissue sections were cut in a cryostat (12 µm), blocked with BlockAidTM blocking solution (cat. no. B10710, Fisher Scientific), and incubated overnight at 4 °C with the following primary antibodies or reagents: TRPC4 (1:1000, cat. no. OST00025W, Fisher Scientific), TRPC4 (1:200, cat. no. ACC-018, Alomone), calcitonin gene-related peptide (CGRP, 1:500, cat. no. ab36001, Abcam, Cambridge, MA), IB4 (1:500, cat. no. I32450, Fisher Scientific) and/or 4’-6-diamidino-2phenylindole dihydrochloride (DAPI; 300 nmol/L, cat. no. D1306, Fisher Scientific). ..

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  • 93
    Alomone Labs rabbit anti trpc5 acc 020 anti trpc4
    Knockout of <t>TRPC5</t> protects cerebral neurons from ischemia-reperfusion-induced apoptosis. ( A , B ) Representative images and summarized data showing phosphatidylserine (PS) externalization as determined using an annexin V-FITC assay in neurons of cerebral slices obtained from wild-type ( A , C ) and TRPC5 knockout (TRPC5 KO) ( B , C ) mice subjected to sham surgery (sham control) or cerebral ischemia-reperfusion. ( C ) Summary of average fluorescence intensity (arbitrary units) of PS externalized neurons in cerebral slices obtained from wild-type and TRPC5 KO mice subjected to sham surgery (control) or cerebral ischemia-reperfusion. ( D , E ) Following surgery, TUNEL assays were performed to visualize apoptotic mouse cerebral neurons as red fluorescence signals in the neurons of cerebral slices obtained from wild-type ( D ) or TRPC5 KO ( E ) mice subjected to cerebral ischemia-reperfusion. Cell nuclei were stained blue with DAPI. Merged images show apoptotic signals overlapping with cell nuclei. ( F ) Summary of the percentages of apoptotic neurons in cerebral slices obtained from wild-type and TRPC5 KO mice subjected to sham surgery (control) or cerebral ischemia–reperfusion. Values represent the mean ± SEM (n = 5-8); * p
    Rabbit Anti Trpc5 Acc 020 Anti Trpc4, supplied by Alomone Labs, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/rabbit anti trpc5 acc 020 anti trpc4/product/Alomone Labs
    Average 93 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    rabbit anti trpc5 acc 020 anti trpc4 - by Bioz Stars, 2021-09
    93/100 stars
      Buy from Supplier

    94
    Alomone Labs rabbit anti asic3 serum
    Soma size distribution of knee joint afferents (A) was unimodal with a broad range of size and similar to that of <t>ASIC3-IR</t> knee joint afferents (C). CGRP-IR knee joint afferents consisted of small to medium cells with most cells smaller than 500μm 2 (B). ASIC3 and CGRP double labeled knee joint afferents were small to medium as well (D) whereas ASIC3 without CGRP-IR knee joint afferents were larger (E). There were no significant differences in soma size distribution among each group in knee joint afferents, CGRP, ASIC3 and ASIC3+CGRP. Only significant difference in soma size distribution was observed in ASIC3 without CGRP (E). Carrageenan-induced arthritis resulted in an overall shift in soma size distribution of ASIC3 without CGRP-IR knee joint afferents to the left (E). * p
    Rabbit Anti Asic3 Serum, supplied by Alomone Labs, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/rabbit anti asic3 serum/product/Alomone Labs
    Average 94 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    rabbit anti asic3 serum - by Bioz Stars, 2021-09
    94/100 stars
      Buy from Supplier

    Image Search Results


    Knockout of TRPC5 protects cerebral neurons from ischemia-reperfusion-induced apoptosis. ( A , B ) Representative images and summarized data showing phosphatidylserine (PS) externalization as determined using an annexin V-FITC assay in neurons of cerebral slices obtained from wild-type ( A , C ) and TRPC5 knockout (TRPC5 KO) ( B , C ) mice subjected to sham surgery (sham control) or cerebral ischemia-reperfusion. ( C ) Summary of average fluorescence intensity (arbitrary units) of PS externalized neurons in cerebral slices obtained from wild-type and TRPC5 KO mice subjected to sham surgery (control) or cerebral ischemia-reperfusion. ( D , E ) Following surgery, TUNEL assays were performed to visualize apoptotic mouse cerebral neurons as red fluorescence signals in the neurons of cerebral slices obtained from wild-type ( D ) or TRPC5 KO ( E ) mice subjected to cerebral ischemia-reperfusion. Cell nuclei were stained blue with DAPI. Merged images show apoptotic signals overlapping with cell nuclei. ( F ) Summary of the percentages of apoptotic neurons in cerebral slices obtained from wild-type and TRPC5 KO mice subjected to sham surgery (control) or cerebral ischemia–reperfusion. Values represent the mean ± SEM (n = 5-8); * p

    Journal: Cells

    Article Title: Transient Receptor Potential Canonical 5-Scramblase Signaling Complex Mediates Neuronal Phosphatidylserine Externalization and Apoptosis

    doi: 10.3390/cells9030547

    Figure Lengend Snippet: Knockout of TRPC5 protects cerebral neurons from ischemia-reperfusion-induced apoptosis. ( A , B ) Representative images and summarized data showing phosphatidylserine (PS) externalization as determined using an annexin V-FITC assay in neurons of cerebral slices obtained from wild-type ( A , C ) and TRPC5 knockout (TRPC5 KO) ( B , C ) mice subjected to sham surgery (sham control) or cerebral ischemia-reperfusion. ( C ) Summary of average fluorescence intensity (arbitrary units) of PS externalized neurons in cerebral slices obtained from wild-type and TRPC5 KO mice subjected to sham surgery (control) or cerebral ischemia-reperfusion. ( D , E ) Following surgery, TUNEL assays were performed to visualize apoptotic mouse cerebral neurons as red fluorescence signals in the neurons of cerebral slices obtained from wild-type ( D ) or TRPC5 KO ( E ) mice subjected to cerebral ischemia-reperfusion. Cell nuclei were stained blue with DAPI. Merged images show apoptotic signals overlapping with cell nuclei. ( F ) Summary of the percentages of apoptotic neurons in cerebral slices obtained from wild-type and TRPC5 KO mice subjected to sham surgery (control) or cerebral ischemia–reperfusion. Values represent the mean ± SEM (n = 5-8); * p

    Article Snippet: Rabbit anti-TRPC5 (ACC-020), anti-TRPC4 (ACC-018) antibodies were purchased from Alomone Labs (Jerusalem, Israel).

    Techniques: Knock-Out, Mouse Assay, Fluorescence, TUNEL Assay, Staining

    TRPC5+PLSCR1 stimulates phosphatidylserine (PS) externalization in HEK293 cells. ( A – F ) Representative images showing TRPC5-mCherry expression and PS externalization on the plasma membrane of HEK293 cell transfected with empty vector ( A – C ) or TRPC5-mCherry+PLSCR1 ( D – F ). The cells were treated with saline as a control ( A , D ), a hypotonic solution ( B , E ) or LaCl 3 (100 μmol/L; C and F ). ( G – H ) Summary data showing the PS externalized cells in percentage of total cells (FITC-positive). G : data from A – C ; H : data from FITC channel in D – F . PS externalization was detected as green fluorescence via the annexin V-FITC assay. TRPC5 is detected as red fluorescence. Values are shown as the mean ± SEM (n = 3); * p

    Journal: Cells

    Article Title: Transient Receptor Potential Canonical 5-Scramblase Signaling Complex Mediates Neuronal Phosphatidylserine Externalization and Apoptosis

    doi: 10.3390/cells9030547

    Figure Lengend Snippet: TRPC5+PLSCR1 stimulates phosphatidylserine (PS) externalization in HEK293 cells. ( A – F ) Representative images showing TRPC5-mCherry expression and PS externalization on the plasma membrane of HEK293 cell transfected with empty vector ( A – C ) or TRPC5-mCherry+PLSCR1 ( D – F ). The cells were treated with saline as a control ( A , D ), a hypotonic solution ( B , E ) or LaCl 3 (100 μmol/L; C and F ). ( G – H ) Summary data showing the PS externalized cells in percentage of total cells (FITC-positive). G : data from A – C ; H : data from FITC channel in D – F . PS externalization was detected as green fluorescence via the annexin V-FITC assay. TRPC5 is detected as red fluorescence. Values are shown as the mean ± SEM (n = 3); * p

    Article Snippet: Rabbit anti-TRPC5 (ACC-020), anti-TRPC4 (ACC-018) antibodies were purchased from Alomone Labs (Jerusalem, Israel).

    Techniques: Expressing, Transfection, Plasmid Preparation, Fluorescence

    Effect of PLSCR1 alone or TRPC5 alone on phosphatidylserine (PS) externalization in HEK293 cells. ( A – F ) Representative images showing the expression of PLSCR1-mCherry or TRPC5-mCherry and PS externalization on the plasma membrane of HEK293 cells transfected with PLSCR1-mCherry alone ( A – C ) or TRPC5-mCherry alone ( D – F ). The cells were treated with saline (control) ( A , D ), hypotonic solution ( B , E ) or LaCl 3 (100 μmol/L) ( C , F ). ( G – H ) Summary data showing the PS externalized cells (FITC-positive) in percentage of total cells (%). G : data from FITC channel in A – C ; H : data from FITC channel in D – F . PS externalization was detected as green fluorescence via the annexin V-FITC assay. PLSCR1 ( A – C ) and TRPC5 ( D – F ) are detected as red fluorescence. Values are shown as the mean ± SEM (n = 3); * p

    Journal: Cells

    Article Title: Transient Receptor Potential Canonical 5-Scramblase Signaling Complex Mediates Neuronal Phosphatidylserine Externalization and Apoptosis

    doi: 10.3390/cells9030547

    Figure Lengend Snippet: Effect of PLSCR1 alone or TRPC5 alone on phosphatidylserine (PS) externalization in HEK293 cells. ( A – F ) Representative images showing the expression of PLSCR1-mCherry or TRPC5-mCherry and PS externalization on the plasma membrane of HEK293 cells transfected with PLSCR1-mCherry alone ( A – C ) or TRPC5-mCherry alone ( D – F ). The cells were treated with saline (control) ( A , D ), hypotonic solution ( B , E ) or LaCl 3 (100 μmol/L) ( C , F ). ( G – H ) Summary data showing the PS externalized cells (FITC-positive) in percentage of total cells (%). G : data from FITC channel in A – C ; H : data from FITC channel in D – F . PS externalization was detected as green fluorescence via the annexin V-FITC assay. PLSCR1 ( A – C ) and TRPC5 ( D – F ) are detected as red fluorescence. Values are shown as the mean ± SEM (n = 3); * p

    Article Snippet: Rabbit anti-TRPC5 (ACC-020), anti-TRPC4 (ACC-018) antibodies were purchased from Alomone Labs (Jerusalem, Israel).

    Techniques: Expressing, Transfection, Fluorescence

    Effect of dominant negative TRPC5 (DN-TRPC5) on phosphatidylserine (PS) externalization in HEK293 cells. ( A – C ) Representative images showing DN-TRPC5-mCherry expression and PS externalization on the plasma membrane of HEK293 cell transfected with DN-TRPC5-mCherry+PLSCR1. The cells were treated with saline (control) ( A ), hypotonic solution ( B ), or LaCl 3 (100 μmol/L) ( C ). ( D ) Summary data showing the PS externalized cells (FITC-positive) in percentage of total cells (%) in A – C . ( E ) Combined summary data showing the PS externalized cells (FITC-positive) in percentage of total cells (%) from Figure 2 , Figure 3 and Figure 4 . The data were from FITC channel. PS externalization was detected as green fluorescence via the annexin V-FITC assay. DN-TRPC5-mCherry was detected as red fluorescence. Values are shown as the mean ± SEM (n = 3); * p

    Journal: Cells

    Article Title: Transient Receptor Potential Canonical 5-Scramblase Signaling Complex Mediates Neuronal Phosphatidylserine Externalization and Apoptosis

    doi: 10.3390/cells9030547

    Figure Lengend Snippet: Effect of dominant negative TRPC5 (DN-TRPC5) on phosphatidylserine (PS) externalization in HEK293 cells. ( A – C ) Representative images showing DN-TRPC5-mCherry expression and PS externalization on the plasma membrane of HEK293 cell transfected with DN-TRPC5-mCherry+PLSCR1. The cells were treated with saline (control) ( A ), hypotonic solution ( B ), or LaCl 3 (100 μmol/L) ( C ). ( D ) Summary data showing the PS externalized cells (FITC-positive) in percentage of total cells (%) in A – C . ( E ) Combined summary data showing the PS externalized cells (FITC-positive) in percentage of total cells (%) from Figure 2 , Figure 3 and Figure 4 . The data were from FITC channel. PS externalization was detected as green fluorescence via the annexin V-FITC assay. DN-TRPC5-mCherry was detected as red fluorescence. Values are shown as the mean ± SEM (n = 3); * p

    Article Snippet: Rabbit anti-TRPC5 (ACC-020), anti-TRPC4 (ACC-018) antibodies were purchased from Alomone Labs (Jerusalem, Israel).

    Techniques: Dominant Negative Mutation, Expressing, Transfection, Fluorescence

    Association between TRPC5 and PLSCR1 in HEK293 cells. ( A , B ) Representative images of co-immunoprecipitation experiments in TRPC5-mCherry and PLSCR1-EGFP co-expressing HEK293 cells. ( A ): IP, anti-EGFP antibody or IgG; IB, anti-mCherry antibody. ( B ), IP, anti-mCherry antibody or IgG; IB, anti-EGFP antibody. Immunoblots of cell lysates were also shown on the right. ( C ) Representative images showing the expression of PLSCR1 in wild-type and PLSCR1-overexpressing HEK293 cells. ( D ) Representative images of co-immunoprecipitation experiments in HEK293 cells co-expressed with mCherry plus PLSCR1-EGFP, or with TRPC5-mCherry plus PLSCR1-EGFP. IP, mCherry antibody; IB, anti-EGFP antibody. ( E ) Representative images showing fluorescence signals in a FRET assay. HEK293 cells were overexpressed with different constructs as indicated. TRPC5-mCherry, mCherry tagged at the carboxyl terminus of TRPC5; mCherry-TRPC5, mCherry tagged at the amino terminus of TRPC5; PLSCR1-EGFP, EGFP tagged at the carboxyl terminus of PLSCR1. The EGFP emission signal was detected as green fluorescence (GFP channel), while the mCherry emission signal was detected as red fluorescence (Cherry channel). The FRET efficiency fluorescence was shown in the FRET efficiency channel. ( F ) Summary of data showing the differences in FRET efficiency. Values are shown as the mean ± SEM (n = 15–22); * p

    Journal: Cells

    Article Title: Transient Receptor Potential Canonical 5-Scramblase Signaling Complex Mediates Neuronal Phosphatidylserine Externalization and Apoptosis

    doi: 10.3390/cells9030547

    Figure Lengend Snippet: Association between TRPC5 and PLSCR1 in HEK293 cells. ( A , B ) Representative images of co-immunoprecipitation experiments in TRPC5-mCherry and PLSCR1-EGFP co-expressing HEK293 cells. ( A ): IP, anti-EGFP antibody or IgG; IB, anti-mCherry antibody. ( B ), IP, anti-mCherry antibody or IgG; IB, anti-EGFP antibody. Immunoblots of cell lysates were also shown on the right. ( C ) Representative images showing the expression of PLSCR1 in wild-type and PLSCR1-overexpressing HEK293 cells. ( D ) Representative images of co-immunoprecipitation experiments in HEK293 cells co-expressed with mCherry plus PLSCR1-EGFP, or with TRPC5-mCherry plus PLSCR1-EGFP. IP, mCherry antibody; IB, anti-EGFP antibody. ( E ) Representative images showing fluorescence signals in a FRET assay. HEK293 cells were overexpressed with different constructs as indicated. TRPC5-mCherry, mCherry tagged at the carboxyl terminus of TRPC5; mCherry-TRPC5, mCherry tagged at the amino terminus of TRPC5; PLSCR1-EGFP, EGFP tagged at the carboxyl terminus of PLSCR1. The EGFP emission signal was detected as green fluorescence (GFP channel), while the mCherry emission signal was detected as red fluorescence (Cherry channel). The FRET efficiency fluorescence was shown in the FRET efficiency channel. ( F ) Summary of data showing the differences in FRET efficiency. Values are shown as the mean ± SEM (n = 15–22); * p

    Article Snippet: Rabbit anti-TRPC5 (ACC-020), anti-TRPC4 (ACC-018) antibodies were purchased from Alomone Labs (Jerusalem, Israel).

    Techniques: Immunoprecipitation, Expressing, Western Blot, Fluorescence, Construct

    Activation of TRPC5 stimulates phosphatidylserine (PS) externalization in mouse cerebral neurons. ( A – E ) Representative images showing PS externalization (green fluorescence) as determined by an annexin V-FITC assay in neurons of cerebral slices obtained from wild-type ( A ) or TRPC5 knockout (TRPC5 KO) mice ( C ) or normal C57 mice ( B , D , E ). ( B ) in the absence of extracellular Ca 2+ (Ca 2+ free); ( D ) treated with IgG; ( E ) treated with a functional blocking antibody T5E3. ( F , G ) Summary of average fluorescence intensity (arbitrary units) of PS externalized neurons in cerebral slices obtained from wild-type (wild type, Ca 2+ free) and TRPC5 KO mice ( F ) or normal C57 mice treated with IgG or T5E3 ( G ). In each group, cerebral slices were treated with hypotonic buffer, daidzein (100 μmol/L) or LaCl 3 (100 μmol/L) for 10 min to activate TRPC5, or treated with saline as control. Values represent the mean ± SEM (n = 4-6); * p

    Journal: Cells

    Article Title: Transient Receptor Potential Canonical 5-Scramblase Signaling Complex Mediates Neuronal Phosphatidylserine Externalization and Apoptosis

    doi: 10.3390/cells9030547

    Figure Lengend Snippet: Activation of TRPC5 stimulates phosphatidylserine (PS) externalization in mouse cerebral neurons. ( A – E ) Representative images showing PS externalization (green fluorescence) as determined by an annexin V-FITC assay in neurons of cerebral slices obtained from wild-type ( A ) or TRPC5 knockout (TRPC5 KO) mice ( C ) or normal C57 mice ( B , D , E ). ( B ) in the absence of extracellular Ca 2+ (Ca 2+ free); ( D ) treated with IgG; ( E ) treated with a functional blocking antibody T5E3. ( F , G ) Summary of average fluorescence intensity (arbitrary units) of PS externalized neurons in cerebral slices obtained from wild-type (wild type, Ca 2+ free) and TRPC5 KO mice ( F ) or normal C57 mice treated with IgG or T5E3 ( G ). In each group, cerebral slices were treated with hypotonic buffer, daidzein (100 μmol/L) or LaCl 3 (100 μmol/L) for 10 min to activate TRPC5, or treated with saline as control. Values represent the mean ± SEM (n = 4-6); * p

    Article Snippet: Rabbit anti-TRPC5 (ACC-020), anti-TRPC4 (ACC-018) antibodies were purchased from Alomone Labs (Jerusalem, Israel).

    Techniques: Activation Assay, Fluorescence, Knock-Out, Mouse Assay, Functional Assay, Blocking Assay

    Association between TRPC5 and PLSCR1 in freshly isolated mouse cortical neurons. ( A , B ) Representative images of co-immunoprecipitation experiments. ( A ): IP, anti-PLSCR1 antibody or IgG; IB, anti-TRPC5 antibody. B , IP, anti-TRPC5 antibody or IgG; IB, anti-PLSCR1 antibody. Immunoblots of cell lysates were also shown on the right. ( A ), immunoblot with rabbit anti-TRPC5 antibody; ( B ), immunoblot with goat anti-PLSCR1 antibody). The experiments were repeated for three times. ( C ) Representative images of co-immunoprecipitation experiments. Right: IP, anti-PLSCR1 antibody or IgG; IB, anti-TRPC1, anti-TRPC4, or anti-PLSCR1 antibody. Left, Immunoblots of cell lysates were also shown. ( D , E ) In situ proximity ligation assay (PLA) to identify the association between TRPC5 and PLSCR1 in native neurons. Representative images were in the presence of both anti-TRPC5 and anti-PLSCR1 antibodies ( E ), or in the presence of anti-TRPC5 antibody alone (Control) ( D ). In D and E , gray sign: anti-goat PLA probe Plus, green sign: goat anti-TRPC5 antibody, pink sign: anti-rabbit PLA probe Minus, blue sign: rabbit anti-PLSCR1 antibody, red dot: PLA signal, left oval: TRPC5 protein, right oval: PLSCR1 protein. Nuclei were stained blue by DAPI.

    Journal: Cells

    Article Title: Transient Receptor Potential Canonical 5-Scramblase Signaling Complex Mediates Neuronal Phosphatidylserine Externalization and Apoptosis

    doi: 10.3390/cells9030547

    Figure Lengend Snippet: Association between TRPC5 and PLSCR1 in freshly isolated mouse cortical neurons. ( A , B ) Representative images of co-immunoprecipitation experiments. ( A ): IP, anti-PLSCR1 antibody or IgG; IB, anti-TRPC5 antibody. B , IP, anti-TRPC5 antibody or IgG; IB, anti-PLSCR1 antibody. Immunoblots of cell lysates were also shown on the right. ( A ), immunoblot with rabbit anti-TRPC5 antibody; ( B ), immunoblot with goat anti-PLSCR1 antibody). The experiments were repeated for three times. ( C ) Representative images of co-immunoprecipitation experiments. Right: IP, anti-PLSCR1 antibody or IgG; IB, anti-TRPC1, anti-TRPC4, or anti-PLSCR1 antibody. Left, Immunoblots of cell lysates were also shown. ( D , E ) In situ proximity ligation assay (PLA) to identify the association between TRPC5 and PLSCR1 in native neurons. Representative images were in the presence of both anti-TRPC5 and anti-PLSCR1 antibodies ( E ), or in the presence of anti-TRPC5 antibody alone (Control) ( D ). In D and E , gray sign: anti-goat PLA probe Plus, green sign: goat anti-TRPC5 antibody, pink sign: anti-rabbit PLA probe Minus, blue sign: rabbit anti-PLSCR1 antibody, red dot: PLA signal, left oval: TRPC5 protein, right oval: PLSCR1 protein. Nuclei were stained blue by DAPI.

    Article Snippet: Rabbit anti-TRPC5 (ACC-020), anti-TRPC4 (ACC-018) antibodies were purchased from Alomone Labs (Jerusalem, Israel).

    Techniques: Isolation, Immunoprecipitation, Western Blot, In Situ, Proximity Ligation Assay, Staining

    Expression of C-type transient receptor potential channels (TRPCs), STIM1, and Orai1 in primary cultured WT and APP KO astrocytes. A – L : Western blot analysis of TRPC1 ( A and B ), TRPC4 ( C and D ), TRPC5 ( E and F ), TRPC3 ( G and H ), STIM1 ( I and

    Journal: American Journal of Physiology - Cell Physiology

    Article Title: Dysregulation of Ca2+ signaling in astrocytes from mice lacking amyloid precursor protein

    doi: 10.1152/ajpcell.00379.2010

    Figure Lengend Snippet: Expression of C-type transient receptor potential channels (TRPCs), STIM1, and Orai1 in primary cultured WT and APP KO astrocytes. A – L : Western blot analysis of TRPC1 ( A and B ), TRPC4 ( C and D ), TRPC5 ( E and F ), TRPC3 ( G and H ), STIM1 ( I and

    Article Snippet: The following antibodies were used: mouse anti-Alzheimer precursor protein A4 monoclonal antibody (APP; clone 22C11) (Millipore, Billerica MA); rabbit polyclonal anti-TRPC1, anti-TRPC3, anti-TRPC4, anti-TRPC5, and anti-TRPC6 (Alomone Laboratories, Jerusalem, Israel); mouse monoclonal anti-Na+ /Ca2+ exchanger 1 (NCX1; clone R3 F1 ) (Swant, Bellinzona, Switzerland); rabbit polyclonal anti-Orai1 (ProSci, Poway, CA); monoclonal anti-plasma membrane Ca2+ pump (PMCA) (Affinity Bioreagents, Rockford, IL); rabbit polyclonal anti-STIM1 (a gift from Dr. J. Roos, Torrey Pines Therapeutics, La Jolla, CA).

    Techniques: Expressing, Cell Culture, Western Blot

    The role of TRPC4 in regulating calcium homeostasis in astrocytes. ( A ) Extracellular Ca 2+ -induced Fluo4 fluorescence changes after TG-induced ER Ca 2+ depletion in Mecp2 -/y astrocytes in the absence (control) or presence of ML204. ( B ) Average traces of TG-induced Fluo4 fluorescence changes in Mecp2 -/y astrocytes with or without 24 hr of ML204 treatment. ( C ) Quantification of percentage (left) of astrocytes showing spontaneous Ca 2+ oscillation and the frequency (middle) and amplitude (right) of such oscillation with or without 24 hr of ML204 treatment. ( D ) TRPC4 Western blot result showing decreased expression level of TRPC4 in mutant RTT human astrocytes after infected with lentivirus-shTrpc4-GFP, compared with astrocytes infected with lentivirus-GFP. ( E ) Quantification of the TRPC4 Western blot result. ( F ) Average traces of extracellular Ca 2+ (2 mM)-induced Rhod-2 fluorescence changes from mutant RTT astrocytes infected with lentivirus expressing either shTRPC4/GFP or GFP alone, after depletion of ER Ca 2+ store by TG pre-treatment. ( G ) Average traces of TG-induced Rhod-2 fluorescence changes from mutant RTT astrocytes infected with lentivirus expressing either shTRPC4/GFP or GFP alone. ( H ) Quantification of the frequency of spontaneous Ca 2+ elevations from mutant RTT astrocytes infected with lentivirus expressing either shTRPC4/GFP or GFP alone. ( I ) Representative images of wild type human RTT astrocytes infected with lentivirus co-expressing TRPC4 and GFP (left) or with lentivirus expressing GFP alone (right). Note anti-TRPC4 immunoreactivity is higher in astrocytes infected with lenti-GFP/TRPC4 than in those infected with lenti-GFP. Scale bar = 50 μm. ( J ) Average traces of extracellular Ca 2+ (2 mM)-induced Rhod-2 fluorescence changes from wild type astrocytes infected with lentivirus expressing either GFP/TRPC4 or GFP alone, after depletion of ER Ca 2+ store by TG pre-treatment. ( K ) Average traces of TG-induced Rhod-2 fluorescence changes from wild type astrocytes infected with lentivirus expressing either GFP/TRPC4 or GFP alone. ( L ) Quantification of percentage (left) of astrocytes infected with lentivirus expressing either GFP/TRPC4 or GFP alone that showed spontaneous Ca 2+ oscillation and the frequency (middle) and amplitude (right) of such oscillations. The bar graphs in this figure show the mean ±s.e.m. *p

    Journal: eLife

    Article Title: Mechanism and consequence of abnormal calcium homeostasis in Rett syndrome astrocytes

    doi: 10.7554/eLife.33417

    Figure Lengend Snippet: The role of TRPC4 in regulating calcium homeostasis in astrocytes. ( A ) Extracellular Ca 2+ -induced Fluo4 fluorescence changes after TG-induced ER Ca 2+ depletion in Mecp2 -/y astrocytes in the absence (control) or presence of ML204. ( B ) Average traces of TG-induced Fluo4 fluorescence changes in Mecp2 -/y astrocytes with or without 24 hr of ML204 treatment. ( C ) Quantification of percentage (left) of astrocytes showing spontaneous Ca 2+ oscillation and the frequency (middle) and amplitude (right) of such oscillation with or without 24 hr of ML204 treatment. ( D ) TRPC4 Western blot result showing decreased expression level of TRPC4 in mutant RTT human astrocytes after infected with lentivirus-shTrpc4-GFP, compared with astrocytes infected with lentivirus-GFP. ( E ) Quantification of the TRPC4 Western blot result. ( F ) Average traces of extracellular Ca 2+ (2 mM)-induced Rhod-2 fluorescence changes from mutant RTT astrocytes infected with lentivirus expressing either shTRPC4/GFP or GFP alone, after depletion of ER Ca 2+ store by TG pre-treatment. ( G ) Average traces of TG-induced Rhod-2 fluorescence changes from mutant RTT astrocytes infected with lentivirus expressing either shTRPC4/GFP or GFP alone. ( H ) Quantification of the frequency of spontaneous Ca 2+ elevations from mutant RTT astrocytes infected with lentivirus expressing either shTRPC4/GFP or GFP alone. ( I ) Representative images of wild type human RTT astrocytes infected with lentivirus co-expressing TRPC4 and GFP (left) or with lentivirus expressing GFP alone (right). Note anti-TRPC4 immunoreactivity is higher in astrocytes infected with lenti-GFP/TRPC4 than in those infected with lenti-GFP. Scale bar = 50 μm. ( J ) Average traces of extracellular Ca 2+ (2 mM)-induced Rhod-2 fluorescence changes from wild type astrocytes infected with lentivirus expressing either GFP/TRPC4 or GFP alone, after depletion of ER Ca 2+ store by TG pre-treatment. ( K ) Average traces of TG-induced Rhod-2 fluorescence changes from wild type astrocytes infected with lentivirus expressing either GFP/TRPC4 or GFP alone. ( L ) Quantification of percentage (left) of astrocytes infected with lentivirus expressing either GFP/TRPC4 or GFP alone that showed spontaneous Ca 2+ oscillation and the frequency (middle) and amplitude (right) of such oscillations. The bar graphs in this figure show the mean ±s.e.m. *p

    Article Snippet: Primary antibody dilutions were as follows: anti-GFAP (Millipore MAB3402 RRID: AB_94844 , 1:500; and Dako, Z0334 RRID: AB_10013382 , 1:500), anti-MeCP2 (Abcam ab50005 RRID: AB_881466 , 1:500 and ab2828 RRID: AB_2143853 , 1:500; and Cell Signaling Technology Cat# 3456S, RRID: AB_10828482 , 1:500), anti-NeuN (Millipore MAB377 RRID: AB_2298772 , 1:100), anti-S100β (Abcam Cat# ab868, RRID: AB_306716 , 1:500) anti-TRPC4 (Alomone labs ACC-018 RRID: AB_2040239 , 1:100; and Abcam ab84813 RRID: AB_1860087 , 1:500).

    Techniques: Fluorescence, Western Blot, Expressing, Mutagenesis, Infection

    Co-staining of TRPC4 and astrocyte marker S100β on hippocampal sections from wild type (WT) and Mecp2 knockout ( Mecp2 -/y ) mice. ( A ) Strong TRPC4 immunoreactivity from Mecp2 -/y (top) but not WT (bottom) mice. Arrows indicate TRPC4 fluorescence co-localized with S100β positive cells. White inserts provide zoomed-in views of the yellow boxed regions. Scale bar = 50 μm. ( B ) Quantification of relative TRPC4 immunoreactivity in S100β positive cells in the hippocampus of WT and Mecp2 -/y mice (WT = 1). ***p

    Journal: eLife

    Article Title: Mechanism and consequence of abnormal calcium homeostasis in Rett syndrome astrocytes

    doi: 10.7554/eLife.33417

    Figure Lengend Snippet: Co-staining of TRPC4 and astrocyte marker S100β on hippocampal sections from wild type (WT) and Mecp2 knockout ( Mecp2 -/y ) mice. ( A ) Strong TRPC4 immunoreactivity from Mecp2 -/y (top) but not WT (bottom) mice. Arrows indicate TRPC4 fluorescence co-localized with S100β positive cells. White inserts provide zoomed-in views of the yellow boxed regions. Scale bar = 50 μm. ( B ) Quantification of relative TRPC4 immunoreactivity in S100β positive cells in the hippocampus of WT and Mecp2 -/y mice (WT = 1). ***p

    Article Snippet: Primary antibody dilutions were as follows: anti-GFAP (Millipore MAB3402 RRID: AB_94844 , 1:500; and Dako, Z0334 RRID: AB_10013382 , 1:500), anti-MeCP2 (Abcam ab50005 RRID: AB_881466 , 1:500 and ab2828 RRID: AB_2143853 , 1:500; and Cell Signaling Technology Cat# 3456S, RRID: AB_10828482 , 1:500), anti-NeuN (Millipore MAB377 RRID: AB_2298772 , 1:100), anti-S100β (Abcam Cat# ab868, RRID: AB_306716 , 1:500) anti-TRPC4 (Alomone labs ACC-018 RRID: AB_2040239 , 1:100; and Abcam ab84813 RRID: AB_1860087 , 1:500).

    Techniques: Staining, Marker, Knock-Out, Mouse Assay, Fluorescence

    Mecp2 -/y astrocytes display abnormal ER Ca 2+ load, Ca 2+ leakage, store operated Ca 2+ entry, and TRPC4 expression. ( A ) Left, average traces of Fluo4 fluorescence changes in wild type and Mecp2 -/y astrocytes treated with 1 μM TG to release ER calcium. Ca 2+ -free aCSF (0 mM Ca 2+ plus 2 mM EGTA) was used; right, quantification of the peak amplitude of F/F 0 in response to TG. p=0.001. ( B ) Left, the rise phase (dots) shown in ( a ) is fitted with a single exponential curve; right, the time constant of the rise phase from Mecp2 -/y astrocytes in response to TG was smaller than that from wild type cells. p=0.004. ( C ) The average trace of Fluo4 fluorescence changes in response to 2 mM aCSF after depletion of ER Ca 2+ store by TG. ( D ) The quantification of ML204 sensitive current from wild type and Mecp2 -/y astrocytes at −70 mV, illustrating higher TRPC4 mediated current in Mecp2 -/y astrocytes. p=0.04. 9–38 WT and 10–39 Mecp2 null astrocytes were included in these analyses. ( E ) Western blot analysis showing increased level of TRPC4 protein in both mouse and human mutant astrocytes. The bar graphs in this figure show the mean ±s.e.m. *p

    Journal: eLife

    Article Title: Mechanism and consequence of abnormal calcium homeostasis in Rett syndrome astrocytes

    doi: 10.7554/eLife.33417

    Figure Lengend Snippet: Mecp2 -/y astrocytes display abnormal ER Ca 2+ load, Ca 2+ leakage, store operated Ca 2+ entry, and TRPC4 expression. ( A ) Left, average traces of Fluo4 fluorescence changes in wild type and Mecp2 -/y astrocytes treated with 1 μM TG to release ER calcium. Ca 2+ -free aCSF (0 mM Ca 2+ plus 2 mM EGTA) was used; right, quantification of the peak amplitude of F/F 0 in response to TG. p=0.001. ( B ) Left, the rise phase (dots) shown in ( a ) is fitted with a single exponential curve; right, the time constant of the rise phase from Mecp2 -/y astrocytes in response to TG was smaller than that from wild type cells. p=0.004. ( C ) The average trace of Fluo4 fluorescence changes in response to 2 mM aCSF after depletion of ER Ca 2+ store by TG. ( D ) The quantification of ML204 sensitive current from wild type and Mecp2 -/y astrocytes at −70 mV, illustrating higher TRPC4 mediated current in Mecp2 -/y astrocytes. p=0.04. 9–38 WT and 10–39 Mecp2 null astrocytes were included in these analyses. ( E ) Western blot analysis showing increased level of TRPC4 protein in both mouse and human mutant astrocytes. The bar graphs in this figure show the mean ±s.e.m. *p

    Article Snippet: Primary antibody dilutions were as follows: anti-GFAP (Millipore MAB3402 RRID: AB_94844 , 1:500; and Dako, Z0334 RRID: AB_10013382 , 1:500), anti-MeCP2 (Abcam ab50005 RRID: AB_881466 , 1:500 and ab2828 RRID: AB_2143853 , 1:500; and Cell Signaling Technology Cat# 3456S, RRID: AB_10828482 , 1:500), anti-NeuN (Millipore MAB377 RRID: AB_2298772 , 1:100), anti-S100β (Abcam Cat# ab868, RRID: AB_306716 , 1:500) anti-TRPC4 (Alomone labs ACC-018 RRID: AB_2040239 , 1:100; and Abcam ab84813 RRID: AB_1860087 , 1:500).

    Techniques: Expressing, Fluorescence, Western Blot, Mutagenesis

    ChIP-qPCR analysis of MeCP2 occupancy on the promoter of the TrpC4 gene in primary mouse astrocytes isolated from either Mecp2-Flag mice (Flag) or control mice (ctrl). 4 pairs of primers (TRPC4-P-1 to −4) were used to cover the TrpC4 promoter. MeCP2 binding signal from the ChIP samples was normalize to signal from the input samples. The bar graphs in this figure show the mean ±s.e.m.

    Journal: eLife

    Article Title: Mechanism and consequence of abnormal calcium homeostasis in Rett syndrome astrocytes

    doi: 10.7554/eLife.33417

    Figure Lengend Snippet: ChIP-qPCR analysis of MeCP2 occupancy on the promoter of the TrpC4 gene in primary mouse astrocytes isolated from either Mecp2-Flag mice (Flag) or control mice (ctrl). 4 pairs of primers (TRPC4-P-1 to −4) were used to cover the TrpC4 promoter. MeCP2 binding signal from the ChIP samples was normalize to signal from the input samples. The bar graphs in this figure show the mean ±s.e.m.

    Article Snippet: Primary antibody dilutions were as follows: anti-GFAP (Millipore MAB3402 RRID: AB_94844 , 1:500; and Dako, Z0334 RRID: AB_10013382 , 1:500), anti-MeCP2 (Abcam ab50005 RRID: AB_881466 , 1:500 and ab2828 RRID: AB_2143853 , 1:500; and Cell Signaling Technology Cat# 3456S, RRID: AB_10828482 , 1:500), anti-NeuN (Millipore MAB377 RRID: AB_2298772 , 1:100), anti-S100β (Abcam Cat# ab868, RRID: AB_306716 , 1:500) anti-TRPC4 (Alomone labs ACC-018 RRID: AB_2040239 , 1:100; and Abcam ab84813 RRID: AB_1860087 , 1:500).

    Techniques: Chromatin Immunoprecipitation, Real-time Polymerase Chain Reaction, Isolation, Mouse Assay, Binding Assay

    Soma size distribution of knee joint afferents (A) was unimodal with a broad range of size and similar to that of ASIC3-IR knee joint afferents (C). CGRP-IR knee joint afferents consisted of small to medium cells with most cells smaller than 500μm 2 (B). ASIC3 and CGRP double labeled knee joint afferents were small to medium as well (D) whereas ASIC3 without CGRP-IR knee joint afferents were larger (E). There were no significant differences in soma size distribution among each group in knee joint afferents, CGRP, ASIC3 and ASIC3+CGRP. Only significant difference in soma size distribution was observed in ASIC3 without CGRP (E). Carrageenan-induced arthritis resulted in an overall shift in soma size distribution of ASIC3 without CGRP-IR knee joint afferents to the left (E). * p

    Journal: The journal of pain : official journal of the American Pain Society

    Article Title: Acid sensing ion channel 3 expression in mice knee joint afferents and effects of carrageenan-induced arthritis

    doi: 10.1016/j.jpain.2008.10.010

    Figure Lengend Snippet: Soma size distribution of knee joint afferents (A) was unimodal with a broad range of size and similar to that of ASIC3-IR knee joint afferents (C). CGRP-IR knee joint afferents consisted of small to medium cells with most cells smaller than 500μm 2 (B). ASIC3 and CGRP double labeled knee joint afferents were small to medium as well (D) whereas ASIC3 without CGRP-IR knee joint afferents were larger (E). There were no significant differences in soma size distribution among each group in knee joint afferents, CGRP, ASIC3 and ASIC3+CGRP. Only significant difference in soma size distribution was observed in ASIC3 without CGRP (E). Carrageenan-induced arthritis resulted in an overall shift in soma size distribution of ASIC3 without CGRP-IR knee joint afferents to the left (E). * p

    Article Snippet: The primary antisera used in this study were rabbit anti-ASIC3 serum (1:500; Alomone Labs, Jerusalem, Israel) and rabbit anti-CGRP serum (1:1000; Peninsula Laboratories, SanCalros, CA).

    Techniques: Labeling

    The number of knee joint afferents (A) and the percentage of knee joint afferents immunoreactive for CGRP (B), ASIC3 (C), ASIC3+CGRP (D) and ASIC3 without CGRP (E). Values are represented as mean±SEM. Knee joint afferents immunoreactive for CGRP, ASIC3 and ASIC3+CGRP were significantly increased 24h after carrageenan-induced arthritis. * p

    Journal: The journal of pain : official journal of the American Pain Society

    Article Title: Acid sensing ion channel 3 expression in mice knee joint afferents and effects of carrageenan-induced arthritis

    doi: 10.1016/j.jpain.2008.10.010

    Figure Lengend Snippet: The number of knee joint afferents (A) and the percentage of knee joint afferents immunoreactive for CGRP (B), ASIC3 (C), ASIC3+CGRP (D) and ASIC3 without CGRP (E). Values are represented as mean±SEM. Knee joint afferents immunoreactive for CGRP, ASIC3 and ASIC3+CGRP were significantly increased 24h after carrageenan-induced arthritis. * p

    Article Snippet: The primary antisera used in this study were rabbit anti-ASIC3 serum (1:500; Alomone Labs, Jerusalem, Israel) and rabbit anti-CGRP serum (1:1000; Peninsula Laboratories, SanCalros, CA).

    Techniques:

    Fast Blue labeling (A,B,C and D) and immnohistochemistry staining for ASIC3 (E,F,G and H) and CGRP (I,J,K and L) showing DRG from ASIC3 +/+ (first and second rows) and ASIC3 -/- (third and fourth rows) mice without joint inflammation (first and third rows) and those 24h after joint inflammation (second and fourth rows). Photos in each row are the same DRG. Arrows indicate Fast Blue labeled DRG neurons. Asterisks indicate ASIC3-IR neurons. No immunoreactivity for ASIC3 was observed in ASIC3 -/-mice(G and H), while CGRP was observed in all groups. Some DRG neurons showing both ASIC3 and CGRP immunoreactivity were observed as yellow in ASIC3+CGRP merged images (M and N).

    Journal: The journal of pain : official journal of the American Pain Society

    Article Title: Acid sensing ion channel 3 expression in mice knee joint afferents and effects of carrageenan-induced arthritis

    doi: 10.1016/j.jpain.2008.10.010

    Figure Lengend Snippet: Fast Blue labeling (A,B,C and D) and immnohistochemistry staining for ASIC3 (E,F,G and H) and CGRP (I,J,K and L) showing DRG from ASIC3 +/+ (first and second rows) and ASIC3 -/- (third and fourth rows) mice without joint inflammation (first and third rows) and those 24h after joint inflammation (second and fourth rows). Photos in each row are the same DRG. Arrows indicate Fast Blue labeled DRG neurons. Asterisks indicate ASIC3-IR neurons. No immunoreactivity for ASIC3 was observed in ASIC3 -/-mice(G and H), while CGRP was observed in all groups. Some DRG neurons showing both ASIC3 and CGRP immunoreactivity were observed as yellow in ASIC3+CGRP merged images (M and N).

    Article Snippet: The primary antisera used in this study were rabbit anti-ASIC3 serum (1:500; Alomone Labs, Jerusalem, Israel) and rabbit anti-CGRP serum (1:1000; Peninsula Laboratories, SanCalros, CA).

    Techniques: Labeling, Staining, Mouse Assay