hcn4  (Alomone Labs)


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    Structured Review

    Alomone Labs hcn4
    Identification of ICC-specific <t>Hcn4</t> . (A) A genomic map view of Hcn4 variants expressed in JICC and CICC. Three alternative initial exons (V1-3) are circled in red and the differential last exon (E8) is boxed in blue. (B) Expression levels of total Hcn isoform genes in JICC and CICC. (C) Expression levels of Hcn4 transcriptional vaiants in JICC and CICC. (D) A topological map of HCN4 variants. Each circle denotes a single amino acid. Colors on amino acid sequence show distinct regions and domains. Green represents start codons found in alternatively initiated variants (V1-3). Six transmembrane domains (S1-6) and a pore region are shown. Red represe nts voltage sensor residues in S4. Two cAMP binding sites are in purple. (E) Western blot analysis showing HCN4 protein expressed in multiple tissues including stomach, jejunum, and colon muscularis. (F) Cryosection images of HCN4 protein in jejunum and colon. HCN4 was detected in the KIT + serosal layer, myenteric plexus, and deep muscular plexus in jejunum. It was also detected in the serosal layer and submuscular plexus in colon with much lower levels. (G) Whole mount images of HCN4 protein in jejunum (top panels) and colon (bottom panels) showing the protein found in KIT + ICC-SS, ICC-MY, and ICC-DMP in jejunum, and KIT + ICC-SS AND ICC-SMP in colon. All scale bars are 50 μm.
    Hcn4, supplied by Alomone Labs, used in various techniques. Bioz Stars score: 92/100, based on 30 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Images

    1) Product Images from "Transcriptome of interstitial cells of Cajal reveals unique and selective gene signatures"

    Article Title: Transcriptome of interstitial cells of Cajal reveals unique and selective gene signatures

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0176031

    Identification of ICC-specific Hcn4 . (A) A genomic map view of Hcn4 variants expressed in JICC and CICC. Three alternative initial exons (V1-3) are circled in red and the differential last exon (E8) is boxed in blue. (B) Expression levels of total Hcn isoform genes in JICC and CICC. (C) Expression levels of Hcn4 transcriptional vaiants in JICC and CICC. (D) A topological map of HCN4 variants. Each circle denotes a single amino acid. Colors on amino acid sequence show distinct regions and domains. Green represents start codons found in alternatively initiated variants (V1-3). Six transmembrane domains (S1-6) and a pore region are shown. Red represe nts voltage sensor residues in S4. Two cAMP binding sites are in purple. (E) Western blot analysis showing HCN4 protein expressed in multiple tissues including stomach, jejunum, and colon muscularis. (F) Cryosection images of HCN4 protein in jejunum and colon. HCN4 was detected in the KIT + serosal layer, myenteric plexus, and deep muscular plexus in jejunum. It was also detected in the serosal layer and submuscular plexus in colon with much lower levels. (G) Whole mount images of HCN4 protein in jejunum (top panels) and colon (bottom panels) showing the protein found in KIT + ICC-SS, ICC-MY, and ICC-DMP in jejunum, and KIT + ICC-SS AND ICC-SMP in colon. All scale bars are 50 μm.
    Figure Legend Snippet: Identification of ICC-specific Hcn4 . (A) A genomic map view of Hcn4 variants expressed in JICC and CICC. Three alternative initial exons (V1-3) are circled in red and the differential last exon (E8) is boxed in blue. (B) Expression levels of total Hcn isoform genes in JICC and CICC. (C) Expression levels of Hcn4 transcriptional vaiants in JICC and CICC. (D) A topological map of HCN4 variants. Each circle denotes a single amino acid. Colors on amino acid sequence show distinct regions and domains. Green represents start codons found in alternatively initiated variants (V1-3). Six transmembrane domains (S1-6) and a pore region are shown. Red represe nts voltage sensor residues in S4. Two cAMP binding sites are in purple. (E) Western blot analysis showing HCN4 protein expressed in multiple tissues including stomach, jejunum, and colon muscularis. (F) Cryosection images of HCN4 protein in jejunum and colon. HCN4 was detected in the KIT + serosal layer, myenteric plexus, and deep muscular plexus in jejunum. It was also detected in the serosal layer and submuscular plexus in colon with much lower levels. (G) Whole mount images of HCN4 protein in jejunum (top panels) and colon (bottom panels) showing the protein found in KIT + ICC-SS, ICC-MY, and ICC-DMP in jejunum, and KIT + ICC-SS AND ICC-SMP in colon. All scale bars are 50 μm.

    Techniques Used: Immunocytochemistry, Expressing, Sequencing, Binding Assay, Western Blot

    2) Product Images from "Spatiotemporal Regulation of an Hcn4 Enhancer Defines a Role for Mef2c and HDACs in Cardiac Electrical Patterning"

    Article Title: Spatiotemporal Regulation of an Hcn4 Enhancer Defines a Role for Mef2c and HDACs in Cardiac Electrical Patterning

    Journal: Developmental biology

    doi: 10.1016/j.ydbio.2012.10.017

    Identification of minimal enhancer regions within R2R3 that direct reporter activity in the Hcn4 expression domain
    Figure Legend Snippet: Identification of minimal enhancer regions within R2R3 that direct reporter activity in the Hcn4 expression domain

    Techniques Used: Activity Assay, Expressing

    HDAC Activity Regulates Hcn4 R2R3
    Figure Legend Snippet: HDAC Activity Regulates Hcn4 R2R3

    Techniques Used: Activity Assay

    Mef2C regulates R2R3-LacZ and Hcn4 expression in vivo
    Figure Legend Snippet: Mef2C regulates R2R3-LacZ and Hcn4 expression in vivo

    Techniques Used: Expressing, In Vivo

    Identification and in vivo testing of Hcn4 regulatory elements
    Figure Legend Snippet: Identification and in vivo testing of Hcn4 regulatory elements

    Techniques Used: In Vivo

    In vivo temporal reporter activity directed by the combined R2-R3 Hcn4 Enhancers
    Figure Legend Snippet: In vivo temporal reporter activity directed by the combined R2-R3 Hcn4 Enhancers

    Techniques Used: In Vivo, Activity Assay

    In vivo expression directed by the R2R3 Hcn4 enhancer in the post-natal AV bundle and its interface with the AV Node
    Figure Legend Snippet: In vivo expression directed by the R2R3 Hcn4 enhancer in the post-natal AV bundle and its interface with the AV Node

    Techniques Used: In Vivo, Expressing

    Mef2C regulates R2R3 Hcn4 enhancer activity via a conserved Mef2 Site
    Figure Legend Snippet: Mef2C regulates R2R3 Hcn4 enhancer activity via a conserved Mef2 Site

    Techniques Used: Activity Assay

    Expression pattern of Hcn4 mRNA by whole-mount in-situ hybridization
    Figure Legend Snippet: Expression pattern of Hcn4 mRNA by whole-mount in-situ hybridization

    Techniques Used: Expressing, In Situ Hybridization

    3) Product Images from "Spatiotemporal Regulation of an Hcn4 Enhancer Defines a Role for Mef2c and HDACs in Cardiac Electrical Patterning"

    Article Title: Spatiotemporal Regulation of an Hcn4 Enhancer Defines a Role for Mef2c and HDACs in Cardiac Electrical Patterning

    Journal: Developmental biology

    doi: 10.1016/j.ydbio.2012.10.017

    Identification of minimal enhancer regions within R2R3 that direct reporter activity in the Hcn4 expression domain
    Figure Legend Snippet: Identification of minimal enhancer regions within R2R3 that direct reporter activity in the Hcn4 expression domain

    Techniques Used: Activity Assay, Expressing

    HDAC Activity Regulates Hcn4 R2R3
    Figure Legend Snippet: HDAC Activity Regulates Hcn4 R2R3

    Techniques Used: Activity Assay

    Mef2C regulates R2R3-LacZ and Hcn4 expression in vivo
    Figure Legend Snippet: Mef2C regulates R2R3-LacZ and Hcn4 expression in vivo

    Techniques Used: Expressing, In Vivo

    Identification and in vivo testing of Hcn4 regulatory elements
    Figure Legend Snippet: Identification and in vivo testing of Hcn4 regulatory elements

    Techniques Used: In Vivo

    In vivo temporal reporter activity directed by the combined R2-R3 Hcn4 Enhancers
    Figure Legend Snippet: In vivo temporal reporter activity directed by the combined R2-R3 Hcn4 Enhancers

    Techniques Used: In Vivo, Activity Assay

    In vivo expression directed by the R2R3 Hcn4 enhancer in the post-natal AV bundle and its interface with the AV Node
    Figure Legend Snippet: In vivo expression directed by the R2R3 Hcn4 enhancer in the post-natal AV bundle and its interface with the AV Node

    Techniques Used: In Vivo, Expressing

    Mef2C regulates R2R3 Hcn4 enhancer activity via a conserved Mef2 Site
    Figure Legend Snippet: Mef2C regulates R2R3 Hcn4 enhancer activity via a conserved Mef2 Site

    Techniques Used: Activity Assay

    Expression pattern of Hcn4 mRNA by whole-mount in-situ hybridization
    Figure Legend Snippet: Expression pattern of Hcn4 mRNA by whole-mount in-situ hybridization

    Techniques Used: Expressing, In Situ Hybridization

    4) Product Images from "The sinus venosus myocardium contributes to the atrioventricular canal: potential role during atrioventricular node development?"

    Article Title: The sinus venosus myocardium contributes to the atrioventricular canal: potential role during atrioventricular node development?

    Journal: Journal of Cellular and Molecular Medicine

    doi: 10.1111/jcmm.12525

    Characterization of the myocardial continuity between the sinus venosus and AV canal. (A) Whole mount ISL1 staining, posterior view of the heart. Arrows indicate ISL1+ cells in continuity sinus venosus and AV canal. Rectangle 1 shows region from which SV-AVC continuity cells were collected. Rectangle 2 shows origin of right ventricular cells for patch clamp. (B–E) mRNA expression after laser capture microscopy (LCM) of TNNT2, NKX2-5, HCN4 and ISL1 . (F–M) Results of single cell patch-clamp experiments. (F and J) Representative tracing of SV-AVC continuity cells (F, pacemaker-like tracing) and RV cells (J, ventricular-like phenotype). (G–I, K–M) Graphs showing the difference in maximum upstroke velocity (G), frequency (H), amplitude (I), APD50 (K), APD90 (L) and maximum diastolic potential (M). * Indicates P
    Figure Legend Snippet: Characterization of the myocardial continuity between the sinus venosus and AV canal. (A) Whole mount ISL1 staining, posterior view of the heart. Arrows indicate ISL1+ cells in continuity sinus venosus and AV canal. Rectangle 1 shows region from which SV-AVC continuity cells were collected. Rectangle 2 shows origin of right ventricular cells for patch clamp. (B–E) mRNA expression after laser capture microscopy (LCM) of TNNT2, NKX2-5, HCN4 and ISL1 . (F–M) Results of single cell patch-clamp experiments. (F and J) Representative tracing of SV-AVC continuity cells (F, pacemaker-like tracing) and RV cells (J, ventricular-like phenotype). (G–I, K–M) Graphs showing the difference in maximum upstroke velocity (G), frequency (H), amplitude (I), APD50 (K), APD90 (L) and maximum diastolic potential (M). * Indicates P

    Techniques Used: Staining, Patch Clamp, Expressing, Microscopy, Laser Capture Microdissection

    HCN4 expression in myocardial continuity between sinus venosus and posterior AV canal. (A) TNNI2 stain of HH19 chick embryo. Arrow indicates continuity between sinus venosus myocardium and posterior AV canal. (B) ISH shows HCN4 mRNA expression in continuity (arrow). (C) Merge of ISL1, TNNI2, HCN4 and DAPI staining of E11.5 mouse embryo, showing the myocardial continuity between the sinus venosus and posterior AV canal. The ISL1+/HCN4+/TNNI2+ SAN (arrowhead) and RVV (arrow) are shown. Note the thick endocardial cushions (asterisks) in the AVC. (D–F) Higher magnifications of boxed area in C and D. ISL1+/TNNI2+/HCN4+ cells are seen in the continuity between sinus venosus myocardium and posterior AV canal myocardium. (E and F) Grey values of ISL1 (E) and HCN4 (F). AVC: atrioventricular canal; LA: left atrium; LB: long bud; LCV: left cardinal vein; LV: left ventricle; OFT: outflow tract; RA: right atrium; RCV: right cardinal vein; SV: sinus venosus; V: ventricle. (C–F) White: TNNI2, green: ISL1, red: HCN4, blue: DAPI; scale bars: 50 μm.
    Figure Legend Snippet: HCN4 expression in myocardial continuity between sinus venosus and posterior AV canal. (A) TNNI2 stain of HH19 chick embryo. Arrow indicates continuity between sinus venosus myocardium and posterior AV canal. (B) ISH shows HCN4 mRNA expression in continuity (arrow). (C) Merge of ISL1, TNNI2, HCN4 and DAPI staining of E11.5 mouse embryo, showing the myocardial continuity between the sinus venosus and posterior AV canal. The ISL1+/HCN4+/TNNI2+ SAN (arrowhead) and RVV (arrow) are shown. Note the thick endocardial cushions (asterisks) in the AVC. (D–F) Higher magnifications of boxed area in C and D. ISL1+/TNNI2+/HCN4+ cells are seen in the continuity between sinus venosus myocardium and posterior AV canal myocardium. (E and F) Grey values of ISL1 (E) and HCN4 (F). AVC: atrioventricular canal; LA: left atrium; LB: long bud; LCV: left cardinal vein; LV: left ventricle; OFT: outflow tract; RA: right atrium; RCV: right cardinal vein; SV: sinus venosus; V: ventricle. (C–F) White: TNNI2, green: ISL1, red: HCN4, blue: DAPI; scale bars: 50 μm.

    Techniques Used: Expressing, Staining, In Situ Hybridization

    5) Product Images from "Spatiotemporal Regulation of an Hcn4 Enhancer Defines a Role for Mef2c and HDACs in Cardiac Electrical Patterning"

    Article Title: Spatiotemporal Regulation of an Hcn4 Enhancer Defines a Role for Mef2c and HDACs in Cardiac Electrical Patterning

    Journal: Developmental biology

    doi: 10.1016/j.ydbio.2012.10.017

    Identification of minimal enhancer regions within R2R3 that direct reporter activity in the Hcn4 expression domain
    Figure Legend Snippet: Identification of minimal enhancer regions within R2R3 that direct reporter activity in the Hcn4 expression domain

    Techniques Used: Activity Assay, Expressing

    HDAC Activity Regulates Hcn4 R2R3
    Figure Legend Snippet: HDAC Activity Regulates Hcn4 R2R3

    Techniques Used: Activity Assay

    Mef2C regulates R2R3-LacZ and Hcn4 expression in vivo
    Figure Legend Snippet: Mef2C regulates R2R3-LacZ and Hcn4 expression in vivo

    Techniques Used: Expressing, In Vivo

    Identification and in vivo testing of Hcn4 regulatory elements
    Figure Legend Snippet: Identification and in vivo testing of Hcn4 regulatory elements

    Techniques Used: In Vivo

    In vivo temporal reporter activity directed by the combined R2-R3 Hcn4 Enhancers
    Figure Legend Snippet: In vivo temporal reporter activity directed by the combined R2-R3 Hcn4 Enhancers

    Techniques Used: In Vivo, Activity Assay

    In vivo expression directed by the R2R3 Hcn4 enhancer in the post-natal AV bundle and its interface with the AV Node
    Figure Legend Snippet: In vivo expression directed by the R2R3 Hcn4 enhancer in the post-natal AV bundle and its interface with the AV Node

    Techniques Used: In Vivo, Expressing

    Mef2C regulates R2R3 Hcn4 enhancer activity via a conserved Mef2 Site
    Figure Legend Snippet: Mef2C regulates R2R3 Hcn4 enhancer activity via a conserved Mef2 Site

    Techniques Used: Activity Assay

    Expression pattern of Hcn4 mRNA by whole-mount in-situ hybridization
    Figure Legend Snippet: Expression pattern of Hcn4 mRNA by whole-mount in-situ hybridization

    Techniques Used: Expressing, In Situ Hybridization

    6) Product Images from "Transcriptome of interstitial cells of Cajal reveals unique and selective gene signatures"

    Article Title: Transcriptome of interstitial cells of Cajal reveals unique and selective gene signatures

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0176031

    Identification of ICC-specific Hcn4 . (A) A genomic map view of Hcn4 variants expressed in JICC and CICC. Three alternative initial exons (V1-3) are circled in red and the differential last exon (E8) is boxed in blue. (B) Expression levels of total Hcn isoform genes in JICC and CICC. (C) Expression levels of Hcn4 transcriptional vaiants in JICC and CICC. (D) A topological map of HCN4 variants. Each circle denotes a single amino acid. Colors on amino acid sequence show distinct regions and domains. Green represents start codons found in alternatively initiated variants (V1-3). Six transmembrane domains (S1-6) and a pore region are shown. Red represe nts voltage sensor residues in S4. Two cAMP binding sites are in purple. (E) Western blot analysis showing HCN4 protein expressed in multiple tissues including stomach, jejunum, and colon muscularis. (F) Cryosection images of HCN4 protein in jejunum and colon. HCN4 was detected in the KIT + serosal layer, myenteric plexus, and deep muscular plexus in jejunum. It was also detected in the serosal layer and submuscular plexus in colon with much lower levels. (G) Whole mount images of HCN4 protein in jejunum (top panels) and colon (bottom panels) showing the protein found in KIT + ICC-SS, ICC-MY, and ICC-DMP in jejunum, and KIT + ICC-SS AND ICC-SMP in colon. All scale bars are 50 μm.
    Figure Legend Snippet: Identification of ICC-specific Hcn4 . (A) A genomic map view of Hcn4 variants expressed in JICC and CICC. Three alternative initial exons (V1-3) are circled in red and the differential last exon (E8) is boxed in blue. (B) Expression levels of total Hcn isoform genes in JICC and CICC. (C) Expression levels of Hcn4 transcriptional vaiants in JICC and CICC. (D) A topological map of HCN4 variants. Each circle denotes a single amino acid. Colors on amino acid sequence show distinct regions and domains. Green represents start codons found in alternatively initiated variants (V1-3). Six transmembrane domains (S1-6) and a pore region are shown. Red represe nts voltage sensor residues in S4. Two cAMP binding sites are in purple. (E) Western blot analysis showing HCN4 protein expressed in multiple tissues including stomach, jejunum, and colon muscularis. (F) Cryosection images of HCN4 protein in jejunum and colon. HCN4 was detected in the KIT + serosal layer, myenteric plexus, and deep muscular plexus in jejunum. It was also detected in the serosal layer and submuscular plexus in colon with much lower levels. (G) Whole mount images of HCN4 protein in jejunum (top panels) and colon (bottom panels) showing the protein found in KIT + ICC-SS, ICC-MY, and ICC-DMP in jejunum, and KIT + ICC-SS AND ICC-SMP in colon. All scale bars are 50 μm.

    Techniques Used: Immunocytochemistry, Expressing, Sequencing, Binding Assay, Western Blot

    7) Product Images from "Distinct perinatal features of the hyperpolarization-activated non-selective cation current Ih in the rat cortical plate"

    Article Title: Distinct perinatal features of the hyperpolarization-activated non-selective cation current Ih in the rat cortical plate

    Journal: Neural Development

    doi: 10.1186/1749-8104-7-21

    Developmental expression change of HCN subunits. (A) Quantitative real-time PCR of rat neocortical tissue for developmental stages E20, P0 and P30 revealed age-dependent changes in HCN3 and HCN4 mRNA expression levels, whereas the expression of HCN1 and HCN2 remained stable. Data were normalized to respective HPRT expression. Experiments were repeated three times and error bars represent SEM. (B) A family of pharmacologically isolated I h at E20 recorded from a holding voltage of −40 mV and activation potentials between −40 and −130 mV confirmed a functional expression of I h already at E20. (C) Western blot analysis from neocortical lysates for HCN1-HCN4 proteins at P0 and P30. For all blots we used anti-beta-actin as loading control and the corresponding subunits (HCN1-HCN4) over-expressed in HEK293 cells as positive controls (ctrl, right lanes ). HCN1, HCN2 and HCN4 membrane protein fraction probes were treated with peptide:N-glycosidase F (PNGase F) in order to remove N-linked glyocsylation. HCN1 was detected in both, P0 and P30 protein extracts, but at higher molecular weights at P30. Deglycosylation at P30 shifted the band to lower molecular weights. However, no shift was observed at P0. Note that the bands at P0 still do not match the bands of deglyocsylated P30 protein. HCN2 was not detectable at P0 but at P30. Deglyocosylation of protein extracts shifted the top band to a lower molecular weight in the P30 samples. HCN4 was only detectable at P0 and the upper band was partially reduced by PNGase F treatment. HCN3 was present in membrane extracts of P0 neocortex tissue whereas no protein was detectable in P30 membrane fractions.
    Figure Legend Snippet: Developmental expression change of HCN subunits. (A) Quantitative real-time PCR of rat neocortical tissue for developmental stages E20, P0 and P30 revealed age-dependent changes in HCN3 and HCN4 mRNA expression levels, whereas the expression of HCN1 and HCN2 remained stable. Data were normalized to respective HPRT expression. Experiments were repeated three times and error bars represent SEM. (B) A family of pharmacologically isolated I h at E20 recorded from a holding voltage of −40 mV and activation potentials between −40 and −130 mV confirmed a functional expression of I h already at E20. (C) Western blot analysis from neocortical lysates for HCN1-HCN4 proteins at P0 and P30. For all blots we used anti-beta-actin as loading control and the corresponding subunits (HCN1-HCN4) over-expressed in HEK293 cells as positive controls (ctrl, right lanes ). HCN1, HCN2 and HCN4 membrane protein fraction probes were treated with peptide:N-glycosidase F (PNGase F) in order to remove N-linked glyocsylation. HCN1 was detected in both, P0 and P30 protein extracts, but at higher molecular weights at P30. Deglycosylation at P30 shifted the band to lower molecular weights. However, no shift was observed at P0. Note that the bands at P0 still do not match the bands of deglyocsylated P30 protein. HCN2 was not detectable at P0 but at P30. Deglyocosylation of protein extracts shifted the top band to a lower molecular weight in the P30 samples. HCN4 was only detectable at P0 and the upper band was partially reduced by PNGase F treatment. HCN3 was present in membrane extracts of P0 neocortex tissue whereas no protein was detectable in P30 membrane fractions.

    Techniques Used: Expressing, Real-time Polymerase Chain Reaction, Isolation, Activation Assay, Functional Assay, Western Blot, Molecular Weight

    Simulation of I h properties and co-expression of HCN1 and HCN4 subunits in HEK293 cells. (A) The simulated combination of a fast- and a slow-activating HCN subunit replicates the characteristic kinetic properties (fast activation and slow deactivation) at P0. Simulation of I h conducted by a fast- and slow-activating HCN subunit ( black trace ) elicited by a −130 mV step for 2 seconds ( bottom ) superimposed onto a trace obtained by the same voltage protocol in a neocortical neuron at P0 ( grey trace ). Inset: the kinetic behavior is explained by the time-dependent contribution of a fast ( blue line ) and a slow ( black line ) component. (B) Families of HCN1- and HCN4-mediated I h recorded from HEK293 cells co-expressing these subunits. Currents in the absence ( top ) and presence ( bottom ) of cAMP were activated from a holding potential of −40 mV to command potentials between −40 and −130 mV of 2 seconds length ( inset ). Tail currents were recorded after stepping back to −40 mV and showed a slow deactivation and tail current amplitudes were measured at the time point indicated by the dotted red line. (C) Voltage sensitivity of HCN1 and HCN4 subunits co-expressed in HEK293 cells. Activation curves in the presence ( closed box ) and absence (open box) of cAMP are depicted as mean ± SEM. For illustration purposes the population means were fitted using a Boltzmann function. For evaluation of the voltage sensitivity individual neurons were considered (see D). Intracellular cAMP clearly shifted the half-maximal activation voltage by approximately 16 mV. (D) Population data on half-maximal activation voltage V 1/2 for the HCN1 and HCN4 mediated I h showed the marked difference ( P
    Figure Legend Snippet: Simulation of I h properties and co-expression of HCN1 and HCN4 subunits in HEK293 cells. (A) The simulated combination of a fast- and a slow-activating HCN subunit replicates the characteristic kinetic properties (fast activation and slow deactivation) at P0. Simulation of I h conducted by a fast- and slow-activating HCN subunit ( black trace ) elicited by a −130 mV step for 2 seconds ( bottom ) superimposed onto a trace obtained by the same voltage protocol in a neocortical neuron at P0 ( grey trace ). Inset: the kinetic behavior is explained by the time-dependent contribution of a fast ( blue line ) and a slow ( black line ) component. (B) Families of HCN1- and HCN4-mediated I h recorded from HEK293 cells co-expressing these subunits. Currents in the absence ( top ) and presence ( bottom ) of cAMP were activated from a holding potential of −40 mV to command potentials between −40 and −130 mV of 2 seconds length ( inset ). Tail currents were recorded after stepping back to −40 mV and showed a slow deactivation and tail current amplitudes were measured at the time point indicated by the dotted red line. (C) Voltage sensitivity of HCN1 and HCN4 subunits co-expressed in HEK293 cells. Activation curves in the presence ( closed box ) and absence (open box) of cAMP are depicted as mean ± SEM. For illustration purposes the population means were fitted using a Boltzmann function. For evaluation of the voltage sensitivity individual neurons were considered (see D). Intracellular cAMP clearly shifted the half-maximal activation voltage by approximately 16 mV. (D) Population data on half-maximal activation voltage V 1/2 for the HCN1 and HCN4 mediated I h showed the marked difference ( P

    Techniques Used: Expressing, Activation Assay

    8) Product Images from "Spatiotemporal Regulation of an Hcn4 Enhancer Defines a Role for Mef2c and HDACs in Cardiac Electrical Patterning"

    Article Title: Spatiotemporal Regulation of an Hcn4 Enhancer Defines a Role for Mef2c and HDACs in Cardiac Electrical Patterning

    Journal: Developmental biology

    doi: 10.1016/j.ydbio.2012.10.017

    Identification of minimal enhancer regions within R2R3 that direct reporter activity in the Hcn4 expression domain
    Figure Legend Snippet: Identification of minimal enhancer regions within R2R3 that direct reporter activity in the Hcn4 expression domain

    Techniques Used: Activity Assay, Expressing

    HDAC Activity Regulates Hcn4 R2R3
    Figure Legend Snippet: HDAC Activity Regulates Hcn4 R2R3

    Techniques Used: Activity Assay

    Mef2C regulates R2R3-LacZ and Hcn4 expression in vivo
    Figure Legend Snippet: Mef2C regulates R2R3-LacZ and Hcn4 expression in vivo

    Techniques Used: Expressing, In Vivo

    Identification and in vivo testing of Hcn4 regulatory elements
    Figure Legend Snippet: Identification and in vivo testing of Hcn4 regulatory elements

    Techniques Used: In Vivo

    In vivo temporal reporter activity directed by the combined R2-R3 Hcn4 Enhancers
    Figure Legend Snippet: In vivo temporal reporter activity directed by the combined R2-R3 Hcn4 Enhancers

    Techniques Used: In Vivo, Activity Assay

    In vivo expression directed by the R2R3 Hcn4 enhancer in the post-natal AV bundle and its interface with the AV Node
    Figure Legend Snippet: In vivo expression directed by the R2R3 Hcn4 enhancer in the post-natal AV bundle and its interface with the AV Node

    Techniques Used: In Vivo, Expressing

    Mef2C regulates R2R3 Hcn4 enhancer activity via a conserved Mef2 Site
    Figure Legend Snippet: Mef2C regulates R2R3 Hcn4 enhancer activity via a conserved Mef2 Site

    Techniques Used: Activity Assay

    Expression pattern of Hcn4 mRNA by whole-mount in-situ hybridization
    Figure Legend Snippet: Expression pattern of Hcn4 mRNA by whole-mount in-situ hybridization

    Techniques Used: Expressing, In Situ Hybridization

    9) Product Images from "Transcriptome of interstitial cells of Cajal reveals unique and selective gene signatures"

    Article Title: Transcriptome of interstitial cells of Cajal reveals unique and selective gene signatures

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0176031

    Identification of ICC-specific Hcn4 . (A) A genomic map view of Hcn4 variants expressed in JICC and CICC. Three alternative initial exons (V1-3) are circled in red and the differential last exon (E8) is boxed in blue. (B) Expression levels of total Hcn isoform genes in JICC and CICC. (C) Expression levels of Hcn4 transcriptional vaiants in JICC and CICC. (D) A topological map of HCN4 variants. Each circle denotes a single amino acid. Colors on amino acid sequence show distinct regions and domains. Green represents start codons found in alternatively initiated variants (V1-3). Six transmembrane domains (S1-6) and a pore region are shown. Red represe nts voltage sensor residues in S4. Two cAMP binding sites are in purple. (E) Western blot analysis showing HCN4 protein expressed in multiple tissues including stomach, jejunum, and colon muscularis. (F) Cryosection images of HCN4 protein in jejunum and colon. HCN4 was detected in the KIT + serosal layer, myenteric plexus, and deep muscular plexus in jejunum. It was also detected in the serosal layer and submuscular plexus in colon with much lower levels. (G) Whole mount images of HCN4 protein in jejunum (top panels) and colon (bottom panels) showing the protein found in KIT + ICC-SS, ICC-MY, and ICC-DMP in jejunum, and KIT + ICC-SS AND ICC-SMP in colon. All scale bars are 50 μm.
    Figure Legend Snippet: Identification of ICC-specific Hcn4 . (A) A genomic map view of Hcn4 variants expressed in JICC and CICC. Three alternative initial exons (V1-3) are circled in red and the differential last exon (E8) is boxed in blue. (B) Expression levels of total Hcn isoform genes in JICC and CICC. (C) Expression levels of Hcn4 transcriptional vaiants in JICC and CICC. (D) A topological map of HCN4 variants. Each circle denotes a single amino acid. Colors on amino acid sequence show distinct regions and domains. Green represents start codons found in alternatively initiated variants (V1-3). Six transmembrane domains (S1-6) and a pore region are shown. Red represe nts voltage sensor residues in S4. Two cAMP binding sites are in purple. (E) Western blot analysis showing HCN4 protein expressed in multiple tissues including stomach, jejunum, and colon muscularis. (F) Cryosection images of HCN4 protein in jejunum and colon. HCN4 was detected in the KIT + serosal layer, myenteric plexus, and deep muscular plexus in jejunum. It was also detected in the serosal layer and submuscular plexus in colon with much lower levels. (G) Whole mount images of HCN4 protein in jejunum (top panels) and colon (bottom panels) showing the protein found in KIT + ICC-SS, ICC-MY, and ICC-DMP in jejunum, and KIT + ICC-SS AND ICC-SMP in colon. All scale bars are 50 μm.

    Techniques Used: Immunocytochemistry, Expressing, Sequencing, Binding Assay, Western Blot

    10) Product Images from "The sinus venosus myocardium contributes to the atrioventricular canal: potential role during atrioventricular node development?"

    Article Title: The sinus venosus myocardium contributes to the atrioventricular canal: potential role during atrioventricular node development?

    Journal: Journal of Cellular and Molecular Medicine

    doi: 10.1111/jcmm.12525

    Characterization of the myocardial continuity between the sinus venosus and AV canal. (A) Whole mount ISL1 staining, posterior view of the heart. Arrows indicate ISL1+ cells in continuity sinus venosus and AV canal. Rectangle 1 shows region from which SV-AVC continuity cells were collected. Rectangle 2 shows origin of right ventricular cells for patch clamp. (B–E) mRNA expression after laser capture microscopy (LCM) of TNNT2, NKX2-5, HCN4 and ISL1 . (F–M) Results of single cell patch-clamp experiments. (F and J) Representative tracing of SV-AVC continuity cells (F, pacemaker-like tracing) and RV cells (J, ventricular-like phenotype). (G–I, K–M) Graphs showing the difference in maximum upstroke velocity (G), frequency (H), amplitude (I), APD50 (K), APD90 (L) and maximum diastolic potential (M). * Indicates P
    Figure Legend Snippet: Characterization of the myocardial continuity between the sinus venosus and AV canal. (A) Whole mount ISL1 staining, posterior view of the heart. Arrows indicate ISL1+ cells in continuity sinus venosus and AV canal. Rectangle 1 shows region from which SV-AVC continuity cells were collected. Rectangle 2 shows origin of right ventricular cells for patch clamp. (B–E) mRNA expression after laser capture microscopy (LCM) of TNNT2, NKX2-5, HCN4 and ISL1 . (F–M) Results of single cell patch-clamp experiments. (F and J) Representative tracing of SV-AVC continuity cells (F, pacemaker-like tracing) and RV cells (J, ventricular-like phenotype). (G–I, K–M) Graphs showing the difference in maximum upstroke velocity (G), frequency (H), amplitude (I), APD50 (K), APD90 (L) and maximum diastolic potential (M). * Indicates P

    Techniques Used: Staining, Patch Clamp, Expressing, Microscopy, Laser Capture Microdissection

    HCN4 expression in myocardial continuity between sinus venosus and posterior AV canal. (A) TNNI2 stain of HH19 chick embryo. Arrow indicates continuity between sinus venosus myocardium and posterior AV canal. (B) ISH shows HCN4 mRNA expression in continuity (arrow). (C) Merge of ISL1, TNNI2, HCN4 and DAPI staining of E11.5 mouse embryo, showing the myocardial continuity between the sinus venosus and posterior AV canal. The ISL1+/HCN4+/TNNI2+ SAN (arrowhead) and RVV (arrow) are shown. Note the thick endocardial cushions (asterisks) in the AVC. (D–F) Higher magnifications of boxed area in C and D. ISL1+/TNNI2+/HCN4+ cells are seen in the continuity between sinus venosus myocardium and posterior AV canal myocardium. (E and F) Grey values of ISL1 (E) and HCN4 (F). AVC: atrioventricular canal; LA: left atrium; LB: long bud; LCV: left cardinal vein; LV: left ventricle; OFT: outflow tract; RA: right atrium; RCV: right cardinal vein; SV: sinus venosus; V: ventricle. (C–F) White: TNNI2, green: ISL1, red: HCN4, blue: DAPI; scale bars: 50 μm.
    Figure Legend Snippet: HCN4 expression in myocardial continuity between sinus venosus and posterior AV canal. (A) TNNI2 stain of HH19 chick embryo. Arrow indicates continuity between sinus venosus myocardium and posterior AV canal. (B) ISH shows HCN4 mRNA expression in continuity (arrow). (C) Merge of ISL1, TNNI2, HCN4 and DAPI staining of E11.5 mouse embryo, showing the myocardial continuity between the sinus venosus and posterior AV canal. The ISL1+/HCN4+/TNNI2+ SAN (arrowhead) and RVV (arrow) are shown. Note the thick endocardial cushions (asterisks) in the AVC. (D–F) Higher magnifications of boxed area in C and D. ISL1+/TNNI2+/HCN4+ cells are seen in the continuity between sinus venosus myocardium and posterior AV canal myocardium. (E and F) Grey values of ISL1 (E) and HCN4 (F). AVC: atrioventricular canal; LA: left atrium; LB: long bud; LCV: left cardinal vein; LV: left ventricle; OFT: outflow tract; RA: right atrium; RCV: right cardinal vein; SV: sinus venosus; V: ventricle. (C–F) White: TNNI2, green: ISL1, red: HCN4, blue: DAPI; scale bars: 50 μm.

    Techniques Used: Expressing, Staining, In Situ Hybridization

    11) Product Images from "The sinus venosus myocardium contributes to the atrioventricular canal: potential role during atrioventricular node development?"

    Article Title: The sinus venosus myocardium contributes to the atrioventricular canal: potential role during atrioventricular node development?

    Journal: Journal of Cellular and Molecular Medicine

    doi: 10.1111/jcmm.12525

    Characterization of the myocardial continuity between the sinus venosus and AV canal. (A) Whole mount ISL1 staining, posterior view of the heart. Arrows indicate ISL1+ cells in continuity sinus venosus and AV canal. Rectangle 1 shows region from which SV-AVC continuity cells were collected. Rectangle 2 shows origin of right ventricular cells for patch clamp. (B–E) mRNA expression after laser capture microscopy (LCM) of TNNT2, NKX2-5, HCN4 and ISL1 . (F–M) Results of single cell patch-clamp experiments. (F and J) Representative tracing of SV-AVC continuity cells (F, pacemaker-like tracing) and RV cells (J, ventricular-like phenotype). (G–I, K–M) Graphs showing the difference in maximum upstroke velocity (G), frequency (H), amplitude (I), APD50 (K), APD90 (L) and maximum diastolic potential (M). * Indicates P
    Figure Legend Snippet: Characterization of the myocardial continuity between the sinus venosus and AV canal. (A) Whole mount ISL1 staining, posterior view of the heart. Arrows indicate ISL1+ cells in continuity sinus venosus and AV canal. Rectangle 1 shows region from which SV-AVC continuity cells were collected. Rectangle 2 shows origin of right ventricular cells for patch clamp. (B–E) mRNA expression after laser capture microscopy (LCM) of TNNT2, NKX2-5, HCN4 and ISL1 . (F–M) Results of single cell patch-clamp experiments. (F and J) Representative tracing of SV-AVC continuity cells (F, pacemaker-like tracing) and RV cells (J, ventricular-like phenotype). (G–I, K–M) Graphs showing the difference in maximum upstroke velocity (G), frequency (H), amplitude (I), APD50 (K), APD90 (L) and maximum diastolic potential (M). * Indicates P

    Techniques Used: Staining, Patch Clamp, Expressing, Microscopy, Laser Capture Microdissection

    HCN4 expression in myocardial continuity between sinus venosus and posterior AV canal. (A) TNNI2 stain of HH19 chick embryo. Arrow indicates continuity between sinus venosus myocardium and posterior AV canal. (B) ISH shows HCN4 mRNA expression in continuity (arrow). (C) Merge of ISL1, TNNI2, HCN4 and DAPI staining of E11.5 mouse embryo, showing the myocardial continuity between the sinus venosus and posterior AV canal. The ISL1+/HCN4+/TNNI2+ SAN (arrowhead) and RVV (arrow) are shown. Note the thick endocardial cushions (asterisks) in the AVC. (D–F) Higher magnifications of boxed area in C and D. ISL1+/TNNI2+/HCN4+ cells are seen in the continuity between sinus venosus myocardium and posterior AV canal myocardium. (E and F) Grey values of ISL1 (E) and HCN4 (F). AVC: atrioventricular canal; LA: left atrium; LB: long bud; LCV: left cardinal vein; LV: left ventricle; OFT: outflow tract; RA: right atrium; RCV: right cardinal vein; SV: sinus venosus; V: ventricle. (C–F) White: TNNI2, green: ISL1, red: HCN4, blue: DAPI; scale bars: 50 μm.
    Figure Legend Snippet: HCN4 expression in myocardial continuity between sinus venosus and posterior AV canal. (A) TNNI2 stain of HH19 chick embryo. Arrow indicates continuity between sinus venosus myocardium and posterior AV canal. (B) ISH shows HCN4 mRNA expression in continuity (arrow). (C) Merge of ISL1, TNNI2, HCN4 and DAPI staining of E11.5 mouse embryo, showing the myocardial continuity between the sinus venosus and posterior AV canal. The ISL1+/HCN4+/TNNI2+ SAN (arrowhead) and RVV (arrow) are shown. Note the thick endocardial cushions (asterisks) in the AVC. (D–F) Higher magnifications of boxed area in C and D. ISL1+/TNNI2+/HCN4+ cells are seen in the continuity between sinus venosus myocardium and posterior AV canal myocardium. (E and F) Grey values of ISL1 (E) and HCN4 (F). AVC: atrioventricular canal; LA: left atrium; LB: long bud; LCV: left cardinal vein; LV: left ventricle; OFT: outflow tract; RA: right atrium; RCV: right cardinal vein; SV: sinus venosus; V: ventricle. (C–F) White: TNNI2, green: ISL1, red: HCN4, blue: DAPI; scale bars: 50 μm.

    Techniques Used: Expressing, Staining, In Situ Hybridization

    12) Product Images from "Spatiotemporal Regulation of an Hcn4 Enhancer Defines a Role for Mef2c and HDACs in Cardiac Electrical Patterning"

    Article Title: Spatiotemporal Regulation of an Hcn4 Enhancer Defines a Role for Mef2c and HDACs in Cardiac Electrical Patterning

    Journal: Developmental biology

    doi: 10.1016/j.ydbio.2012.10.017

    Identification of minimal enhancer regions within R2R3 that direct reporter activity in the Hcn4 expression domain
    Figure Legend Snippet: Identification of minimal enhancer regions within R2R3 that direct reporter activity in the Hcn4 expression domain

    Techniques Used: Activity Assay, Expressing

    HDAC Activity Regulates Hcn4 R2R3
    Figure Legend Snippet: HDAC Activity Regulates Hcn4 R2R3

    Techniques Used: Activity Assay

    Mef2C regulates R2R3-LacZ and Hcn4 expression in vivo
    Figure Legend Snippet: Mef2C regulates R2R3-LacZ and Hcn4 expression in vivo

    Techniques Used: Expressing, In Vivo

    Identification and in vivo testing of Hcn4 regulatory elements
    Figure Legend Snippet: Identification and in vivo testing of Hcn4 regulatory elements

    Techniques Used: In Vivo

    In vivo temporal reporter activity directed by the combined R2-R3 Hcn4 Enhancers
    Figure Legend Snippet: In vivo temporal reporter activity directed by the combined R2-R3 Hcn4 Enhancers

    Techniques Used: In Vivo, Activity Assay

    In vivo expression directed by the R2R3 Hcn4 enhancer in the post-natal AV bundle and its interface with the AV Node
    Figure Legend Snippet: In vivo expression directed by the R2R3 Hcn4 enhancer in the post-natal AV bundle and its interface with the AV Node

    Techniques Used: In Vivo, Expressing

    Mef2C regulates R2R3 Hcn4 enhancer activity via a conserved Mef2 Site
    Figure Legend Snippet: Mef2C regulates R2R3 Hcn4 enhancer activity via a conserved Mef2 Site

    Techniques Used: Activity Assay

    Expression pattern of Hcn4 mRNA by whole-mount in-situ hybridization
    Figure Legend Snippet: Expression pattern of Hcn4 mRNA by whole-mount in-situ hybridization

    Techniques Used: Expressing, In Situ Hybridization

    13) Product Images from "Spatiotemporal Regulation of an Hcn4 Enhancer Defines a Role for Mef2c and HDACs in Cardiac Electrical Patterning"

    Article Title: Spatiotemporal Regulation of an Hcn4 Enhancer Defines a Role for Mef2c and HDACs in Cardiac Electrical Patterning

    Journal: Developmental biology

    doi: 10.1016/j.ydbio.2012.10.017

    Identification of minimal enhancer regions within R2R3 that direct reporter activity in the Hcn4 expression domain
    Figure Legend Snippet: Identification of minimal enhancer regions within R2R3 that direct reporter activity in the Hcn4 expression domain

    Techniques Used: Activity Assay, Expressing

    HDAC Activity Regulates Hcn4 R2R3
    Figure Legend Snippet: HDAC Activity Regulates Hcn4 R2R3

    Techniques Used: Activity Assay

    Mef2C regulates R2R3-LacZ and Hcn4 expression in vivo
    Figure Legend Snippet: Mef2C regulates R2R3-LacZ and Hcn4 expression in vivo

    Techniques Used: Expressing, In Vivo

    Identification and in vivo testing of Hcn4 regulatory elements
    Figure Legend Snippet: Identification and in vivo testing of Hcn4 regulatory elements

    Techniques Used: In Vivo

    In vivo temporal reporter activity directed by the combined R2-R3 Hcn4 Enhancers
    Figure Legend Snippet: In vivo temporal reporter activity directed by the combined R2-R3 Hcn4 Enhancers

    Techniques Used: In Vivo, Activity Assay

    In vivo expression directed by the R2R3 Hcn4 enhancer in the post-natal AV bundle and its interface with the AV Node
    Figure Legend Snippet: In vivo expression directed by the R2R3 Hcn4 enhancer in the post-natal AV bundle and its interface with the AV Node

    Techniques Used: In Vivo, Expressing

    Mef2C regulates R2R3 Hcn4 enhancer activity via a conserved Mef2 Site
    Figure Legend Snippet: Mef2C regulates R2R3 Hcn4 enhancer activity via a conserved Mef2 Site

    Techniques Used: Activity Assay

    Expression pattern of Hcn4 mRNA by whole-mount in-situ hybridization
    Figure Legend Snippet: Expression pattern of Hcn4 mRNA by whole-mount in-situ hybridization

    Techniques Used: Expressing, In Situ Hybridization

    14) Product Images from "The sinus venosus myocardium contributes to the atrioventricular canal: potential role during atrioventricular node development?"

    Article Title: The sinus venosus myocardium contributes to the atrioventricular canal: potential role during atrioventricular node development?

    Journal: Journal of Cellular and Molecular Medicine

    doi: 10.1111/jcmm.12525

    Characterization of the myocardial continuity between the sinus venosus and AV canal. (A) Whole mount ISL1 staining, posterior view of the heart. Arrows indicate ISL1+ cells in continuity sinus venosus and AV canal. Rectangle 1 shows region from which SV-AVC continuity cells were collected. Rectangle 2 shows origin of right ventricular cells for patch clamp. (B–E) mRNA expression after laser capture microscopy (LCM) of TNNT2, NKX2-5, HCN4 and ISL1 . (F–M) Results of single cell patch-clamp experiments. (F and J) Representative tracing of SV-AVC continuity cells (F, pacemaker-like tracing) and RV cells (J, ventricular-like phenotype). (G–I, K–M) Graphs showing the difference in maximum upstroke velocity (G), frequency (H), amplitude (I), APD50 (K), APD90 (L) and maximum diastolic potential (M). * Indicates P
    Figure Legend Snippet: Characterization of the myocardial continuity between the sinus venosus and AV canal. (A) Whole mount ISL1 staining, posterior view of the heart. Arrows indicate ISL1+ cells in continuity sinus venosus and AV canal. Rectangle 1 shows region from which SV-AVC continuity cells were collected. Rectangle 2 shows origin of right ventricular cells for patch clamp. (B–E) mRNA expression after laser capture microscopy (LCM) of TNNT2, NKX2-5, HCN4 and ISL1 . (F–M) Results of single cell patch-clamp experiments. (F and J) Representative tracing of SV-AVC continuity cells (F, pacemaker-like tracing) and RV cells (J, ventricular-like phenotype). (G–I, K–M) Graphs showing the difference in maximum upstroke velocity (G), frequency (H), amplitude (I), APD50 (K), APD90 (L) and maximum diastolic potential (M). * Indicates P

    Techniques Used: Staining, Patch Clamp, Expressing, Microscopy, Laser Capture Microdissection

    HCN4 expression in myocardial continuity between sinus venosus and posterior AV canal. (A) TNNI2 stain of HH19 chick embryo. Arrow indicates continuity between sinus venosus myocardium and posterior AV canal. (B) ISH shows HCN4 mRNA expression in continuity (arrow). (C) Merge of ISL1, TNNI2, HCN4 and DAPI staining of E11.5 mouse embryo, showing the myocardial continuity between the sinus venosus and posterior AV canal. The ISL1+/HCN4+/TNNI2+ SAN (arrowhead) and RVV (arrow) are shown. Note the thick endocardial cushions (asterisks) in the AVC. (D–F) Higher magnifications of boxed area in C and D. ISL1+/TNNI2+/HCN4+ cells are seen in the continuity between sinus venosus myocardium and posterior AV canal myocardium. (E and F) Grey values of ISL1 (E) and HCN4 (F). AVC: atrioventricular canal; LA: left atrium; LB: long bud; LCV: left cardinal vein; LV: left ventricle; OFT: outflow tract; RA: right atrium; RCV: right cardinal vein; SV: sinus venosus; V: ventricle. (C–F) White: TNNI2, green: ISL1, red: HCN4, blue: DAPI; scale bars: 50 μm.
    Figure Legend Snippet: HCN4 expression in myocardial continuity between sinus venosus and posterior AV canal. (A) TNNI2 stain of HH19 chick embryo. Arrow indicates continuity between sinus venosus myocardium and posterior AV canal. (B) ISH shows HCN4 mRNA expression in continuity (arrow). (C) Merge of ISL1, TNNI2, HCN4 and DAPI staining of E11.5 mouse embryo, showing the myocardial continuity between the sinus venosus and posterior AV canal. The ISL1+/HCN4+/TNNI2+ SAN (arrowhead) and RVV (arrow) are shown. Note the thick endocardial cushions (asterisks) in the AVC. (D–F) Higher magnifications of boxed area in C and D. ISL1+/TNNI2+/HCN4+ cells are seen in the continuity between sinus venosus myocardium and posterior AV canal myocardium. (E and F) Grey values of ISL1 (E) and HCN4 (F). AVC: atrioventricular canal; LA: left atrium; LB: long bud; LCV: left cardinal vein; LV: left ventricle; OFT: outflow tract; RA: right atrium; RCV: right cardinal vein; SV: sinus venosus; V: ventricle. (C–F) White: TNNI2, green: ISL1, red: HCN4, blue: DAPI; scale bars: 50 μm.

    Techniques Used: Expressing, Staining, In Situ Hybridization

    15) Product Images from "Transcriptome of interstitial cells of Cajal reveals unique and selective gene signatures"

    Article Title: Transcriptome of interstitial cells of Cajal reveals unique and selective gene signatures

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0176031

    Identification of ICC-specific Hcn4 . (A) A genomic map view of Hcn4 variants expressed in JICC and CICC. Three alternative initial exons (V1-3) are circled in red and the differential last exon (E8) is boxed in blue. (B) Expression levels of total Hcn isoform genes in JICC and CICC. (C) Expression levels of Hcn4 transcriptional vaiants in JICC and CICC. (D) A topological map of HCN4 variants. Each circle denotes a single amino acid. Colors on amino acid sequence show distinct regions and domains. Green represents start codons found in alternatively initiated variants (V1-3). Six transmembrane domains (S1-6) and a pore region are shown. Red represe nts voltage sensor residues in S4. Two cAMP binding sites are in purple. (E) Western blot analysis showing HCN4 protein expressed in multiple tissues including stomach, jejunum, and colon muscularis. (F) Cryosection images of HCN4 protein in jejunum and colon. HCN4 was detected in the KIT + serosal layer, myenteric plexus, and deep muscular plexus in jejunum. It was also detected in the serosal layer and submuscular plexus in colon with much lower levels. (G) Whole mount images of HCN4 protein in jejunum (top panels) and colon (bottom panels) showing the protein found in KIT + ICC-SS, ICC-MY, and ICC-DMP in jejunum, and KIT + ICC-SS AND ICC-SMP in colon. All scale bars are 50 μm.
    Figure Legend Snippet: Identification of ICC-specific Hcn4 . (A) A genomic map view of Hcn4 variants expressed in JICC and CICC. Three alternative initial exons (V1-3) are circled in red and the differential last exon (E8) is boxed in blue. (B) Expression levels of total Hcn isoform genes in JICC and CICC. (C) Expression levels of Hcn4 transcriptional vaiants in JICC and CICC. (D) A topological map of HCN4 variants. Each circle denotes a single amino acid. Colors on amino acid sequence show distinct regions and domains. Green represents start codons found in alternatively initiated variants (V1-3). Six transmembrane domains (S1-6) and a pore region are shown. Red represe nts voltage sensor residues in S4. Two cAMP binding sites are in purple. (E) Western blot analysis showing HCN4 protein expressed in multiple tissues including stomach, jejunum, and colon muscularis. (F) Cryosection images of HCN4 protein in jejunum and colon. HCN4 was detected in the KIT + serosal layer, myenteric plexus, and deep muscular plexus in jejunum. It was also detected in the serosal layer and submuscular plexus in colon with much lower levels. (G) Whole mount images of HCN4 protein in jejunum (top panels) and colon (bottom panels) showing the protein found in KIT + ICC-SS, ICC-MY, and ICC-DMP in jejunum, and KIT + ICC-SS AND ICC-SMP in colon. All scale bars are 50 μm.

    Techniques Used: Immunocytochemistry, Expressing, Sequencing, Binding Assay, Western Blot

    16) Product Images from "The sinus venosus myocardium contributes to the atrioventricular canal: potential role during atrioventricular node development?"

    Article Title: The sinus venosus myocardium contributes to the atrioventricular canal: potential role during atrioventricular node development?

    Journal: Journal of Cellular and Molecular Medicine

    doi: 10.1111/jcmm.12525

    Characterization of the myocardial continuity between the sinus venosus and AV canal. (A) Whole mount ISL1 staining, posterior view of the heart. Arrows indicate ISL1+ cells in continuity sinus venosus and AV canal. Rectangle 1 shows region from which SV-AVC continuity cells were collected. Rectangle 2 shows origin of right ventricular cells for patch clamp. (B–E) mRNA expression after laser capture microscopy (LCM) of TNNT2, NKX2-5, HCN4 and ISL1 . (F–M) Results of single cell patch-clamp experiments. (F and J) Representative tracing of SV-AVC continuity cells (F, pacemaker-like tracing) and RV cells (J, ventricular-like phenotype). (G–I, K–M) Graphs showing the difference in maximum upstroke velocity (G), frequency (H), amplitude (I), APD50 (K), APD90 (L) and maximum diastolic potential (M). * Indicates P
    Figure Legend Snippet: Characterization of the myocardial continuity between the sinus venosus and AV canal. (A) Whole mount ISL1 staining, posterior view of the heart. Arrows indicate ISL1+ cells in continuity sinus venosus and AV canal. Rectangle 1 shows region from which SV-AVC continuity cells were collected. Rectangle 2 shows origin of right ventricular cells for patch clamp. (B–E) mRNA expression after laser capture microscopy (LCM) of TNNT2, NKX2-5, HCN4 and ISL1 . (F–M) Results of single cell patch-clamp experiments. (F and J) Representative tracing of SV-AVC continuity cells (F, pacemaker-like tracing) and RV cells (J, ventricular-like phenotype). (G–I, K–M) Graphs showing the difference in maximum upstroke velocity (G), frequency (H), amplitude (I), APD50 (K), APD90 (L) and maximum diastolic potential (M). * Indicates P

    Techniques Used: Staining, Patch Clamp, Expressing, Microscopy, Laser Capture Microdissection

    HCN4 expression in myocardial continuity between sinus venosus and posterior AV canal. (A) TNNI2 stain of HH19 chick embryo. Arrow indicates continuity between sinus venosus myocardium and posterior AV canal. (B) ISH shows HCN4 mRNA expression in continuity (arrow). (C) Merge of ISL1, TNNI2, HCN4 and DAPI staining of E11.5 mouse embryo, showing the myocardial continuity between the sinus venosus and posterior AV canal. The ISL1+/HCN4+/TNNI2+ SAN (arrowhead) and RVV (arrow) are shown. Note the thick endocardial cushions (asterisks) in the AVC. (D–F) Higher magnifications of boxed area in C and D. ISL1+/TNNI2+/HCN4+ cells are seen in the continuity between sinus venosus myocardium and posterior AV canal myocardium. (E and F) Grey values of ISL1 (E) and HCN4 (F). AVC: atrioventricular canal; LA: left atrium; LB: long bud; LCV: left cardinal vein; LV: left ventricle; OFT: outflow tract; RA: right atrium; RCV: right cardinal vein; SV: sinus venosus; V: ventricle. (C–F) White: TNNI2, green: ISL1, red: HCN4, blue: DAPI; scale bars: 50 μm.
    Figure Legend Snippet: HCN4 expression in myocardial continuity between sinus venosus and posterior AV canal. (A) TNNI2 stain of HH19 chick embryo. Arrow indicates continuity between sinus venosus myocardium and posterior AV canal. (B) ISH shows HCN4 mRNA expression in continuity (arrow). (C) Merge of ISL1, TNNI2, HCN4 and DAPI staining of E11.5 mouse embryo, showing the myocardial continuity between the sinus venosus and posterior AV canal. The ISL1+/HCN4+/TNNI2+ SAN (arrowhead) and RVV (arrow) are shown. Note the thick endocardial cushions (asterisks) in the AVC. (D–F) Higher magnifications of boxed area in C and D. ISL1+/TNNI2+/HCN4+ cells are seen in the continuity between sinus venosus myocardium and posterior AV canal myocardium. (E and F) Grey values of ISL1 (E) and HCN4 (F). AVC: atrioventricular canal; LA: left atrium; LB: long bud; LCV: left cardinal vein; LV: left ventricle; OFT: outflow tract; RA: right atrium; RCV: right cardinal vein; SV: sinus venosus; V: ventricle. (C–F) White: TNNI2, green: ISL1, red: HCN4, blue: DAPI; scale bars: 50 μm.

    Techniques Used: Expressing, Staining, In Situ Hybridization

    17) Product Images from "Spatiotemporal Regulation of an Hcn4 Enhancer Defines a Role for Mef2c and HDACs in Cardiac Electrical Patterning"

    Article Title: Spatiotemporal Regulation of an Hcn4 Enhancer Defines a Role for Mef2c and HDACs in Cardiac Electrical Patterning

    Journal: Developmental biology

    doi: 10.1016/j.ydbio.2012.10.017

    Identification of minimal enhancer regions within R2R3 that direct reporter activity in the Hcn4 expression domain
    Figure Legend Snippet: Identification of minimal enhancer regions within R2R3 that direct reporter activity in the Hcn4 expression domain

    Techniques Used: Activity Assay, Expressing

    HDAC Activity Regulates Hcn4 R2R3
    Figure Legend Snippet: HDAC Activity Regulates Hcn4 R2R3

    Techniques Used: Activity Assay

    Mef2C regulates R2R3-LacZ and Hcn4 expression in vivo
    Figure Legend Snippet: Mef2C regulates R2R3-LacZ and Hcn4 expression in vivo

    Techniques Used: Expressing, In Vivo

    Identification and in vivo testing of Hcn4 regulatory elements
    Figure Legend Snippet: Identification and in vivo testing of Hcn4 regulatory elements

    Techniques Used: In Vivo

    In vivo temporal reporter activity directed by the combined R2-R3 Hcn4 Enhancers
    Figure Legend Snippet: In vivo temporal reporter activity directed by the combined R2-R3 Hcn4 Enhancers

    Techniques Used: In Vivo, Activity Assay

    In vivo expression directed by the R2R3 Hcn4 enhancer in the post-natal AV bundle and its interface with the AV Node
    Figure Legend Snippet: In vivo expression directed by the R2R3 Hcn4 enhancer in the post-natal AV bundle and its interface with the AV Node

    Techniques Used: In Vivo, Expressing

    Mef2C regulates R2R3 Hcn4 enhancer activity via a conserved Mef2 Site
    Figure Legend Snippet: Mef2C regulates R2R3 Hcn4 enhancer activity via a conserved Mef2 Site

    Techniques Used: Activity Assay

    Expression pattern of Hcn4 mRNA by whole-mount in-situ hybridization
    Figure Legend Snippet: Expression pattern of Hcn4 mRNA by whole-mount in-situ hybridization

    Techniques Used: Expressing, In Situ Hybridization

    18) Product Images from "Transcriptome of interstitial cells of Cajal reveals unique and selective gene signatures"

    Article Title: Transcriptome of interstitial cells of Cajal reveals unique and selective gene signatures

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0176031

    Identification of ICC-specific Hcn4 . (A) A genomic map view of Hcn4 variants expressed in JICC and CICC. Three alternative initial exons (V1-3) are circled in red and the differential last exon (E8) is boxed in blue. (B) Expression levels of total Hcn isoform genes in JICC and CICC. (C) Expression levels of Hcn4 transcriptional vaiants in JICC and CICC. (D) A topological map of HCN4 variants. Each circle denotes a single amino acid. Colors on amino acid sequence show distinct regions and domains. Green represents start codons found in alternatively initiated variants (V1-3). Six transmembrane domains (S1-6) and a pore region are shown. Red represe nts voltage sensor residues in S4. Two cAMP binding sites are in purple. (E) Western blot analysis showing HCN4 protein expressed in multiple tissues including stomach, jejunum, and colon muscularis. (F) Cryosection images of HCN4 protein in jejunum and colon. HCN4 was detected in the KIT + serosal layer, myenteric plexus, and deep muscular plexus in jejunum. It was also detected in the serosal layer and submuscular plexus in colon with much lower levels. (G) Whole mount images of HCN4 protein in jejunum (top panels) and colon (bottom panels) showing the protein found in KIT + ICC-SS, ICC-MY, and ICC-DMP in jejunum, and KIT + ICC-SS AND ICC-SMP in colon. All scale bars are 50 μm.
    Figure Legend Snippet: Identification of ICC-specific Hcn4 . (A) A genomic map view of Hcn4 variants expressed in JICC and CICC. Three alternative initial exons (V1-3) are circled in red and the differential last exon (E8) is boxed in blue. (B) Expression levels of total Hcn isoform genes in JICC and CICC. (C) Expression levels of Hcn4 transcriptional vaiants in JICC and CICC. (D) A topological map of HCN4 variants. Each circle denotes a single amino acid. Colors on amino acid sequence show distinct regions and domains. Green represents start codons found in alternatively initiated variants (V1-3). Six transmembrane domains (S1-6) and a pore region are shown. Red represe nts voltage sensor residues in S4. Two cAMP binding sites are in purple. (E) Western blot analysis showing HCN4 protein expressed in multiple tissues including stomach, jejunum, and colon muscularis. (F) Cryosection images of HCN4 protein in jejunum and colon. HCN4 was detected in the KIT + serosal layer, myenteric plexus, and deep muscular plexus in jejunum. It was also detected in the serosal layer and submuscular plexus in colon with much lower levels. (G) Whole mount images of HCN4 protein in jejunum (top panels) and colon (bottom panels) showing the protein found in KIT + ICC-SS, ICC-MY, and ICC-DMP in jejunum, and KIT + ICC-SS AND ICC-SMP in colon. All scale bars are 50 μm.

    Techniques Used: Immunocytochemistry, Expressing, Sequencing, Binding Assay, Western Blot

    19) Product Images from "The sinus venosus myocardium contributes to the atrioventricular canal: potential role during atrioventricular node development?"

    Article Title: The sinus venosus myocardium contributes to the atrioventricular canal: potential role during atrioventricular node development?

    Journal: Journal of Cellular and Molecular Medicine

    doi: 10.1111/jcmm.12525

    Characterization of the myocardial continuity between the sinus venosus and AV canal. (A) Whole mount ISL1 staining, posterior view of the heart. Arrows indicate ISL1+ cells in continuity sinus venosus and AV canal. Rectangle 1 shows region from which SV-AVC continuity cells were collected. Rectangle 2 shows origin of right ventricular cells for patch clamp. (B–E) mRNA expression after laser capture microscopy (LCM) of TNNT2, NKX2-5, HCN4 and ISL1 . (F–M) Results of single cell patch-clamp experiments. (F and J) Representative tracing of SV-AVC continuity cells (F, pacemaker-like tracing) and RV cells (J, ventricular-like phenotype). (G–I, K–M) Graphs showing the difference in maximum upstroke velocity (G), frequency (H), amplitude (I), APD50 (K), APD90 (L) and maximum diastolic potential (M). * Indicates P
    Figure Legend Snippet: Characterization of the myocardial continuity between the sinus venosus and AV canal. (A) Whole mount ISL1 staining, posterior view of the heart. Arrows indicate ISL1+ cells in continuity sinus venosus and AV canal. Rectangle 1 shows region from which SV-AVC continuity cells were collected. Rectangle 2 shows origin of right ventricular cells for patch clamp. (B–E) mRNA expression after laser capture microscopy (LCM) of TNNT2, NKX2-5, HCN4 and ISL1 . (F–M) Results of single cell patch-clamp experiments. (F and J) Representative tracing of SV-AVC continuity cells (F, pacemaker-like tracing) and RV cells (J, ventricular-like phenotype). (G–I, K–M) Graphs showing the difference in maximum upstroke velocity (G), frequency (H), amplitude (I), APD50 (K), APD90 (L) and maximum diastolic potential (M). * Indicates P

    Techniques Used: Staining, Patch Clamp, Expressing, Microscopy, Laser Capture Microdissection

    HCN4 expression in myocardial continuity between sinus venosus and posterior AV canal. (A) TNNI2 stain of HH19 chick embryo. Arrow indicates continuity between sinus venosus myocardium and posterior AV canal. (B) ISH shows HCN4 mRNA expression in continuity (arrow). (C) Merge of ISL1, TNNI2, HCN4 and DAPI staining of E11.5 mouse embryo, showing the myocardial continuity between the sinus venosus and posterior AV canal. The ISL1+/HCN4+/TNNI2+ SAN (arrowhead) and RVV (arrow) are shown. Note the thick endocardial cushions (asterisks) in the AVC. (D–F) Higher magnifications of boxed area in C and D. ISL1+/TNNI2+/HCN4+ cells are seen in the continuity between sinus venosus myocardium and posterior AV canal myocardium. (E and F) Grey values of ISL1 (E) and HCN4 (F). AVC: atrioventricular canal; LA: left atrium; LB: long bud; LCV: left cardinal vein; LV: left ventricle; OFT: outflow tract; RA: right atrium; RCV: right cardinal vein; SV: sinus venosus; V: ventricle. (C–F) White: TNNI2, green: ISL1, red: HCN4, blue: DAPI; scale bars: 50 μm.
    Figure Legend Snippet: HCN4 expression in myocardial continuity between sinus venosus and posterior AV canal. (A) TNNI2 stain of HH19 chick embryo. Arrow indicates continuity between sinus venosus myocardium and posterior AV canal. (B) ISH shows HCN4 mRNA expression in continuity (arrow). (C) Merge of ISL1, TNNI2, HCN4 and DAPI staining of E11.5 mouse embryo, showing the myocardial continuity between the sinus venosus and posterior AV canal. The ISL1+/HCN4+/TNNI2+ SAN (arrowhead) and RVV (arrow) are shown. Note the thick endocardial cushions (asterisks) in the AVC. (D–F) Higher magnifications of boxed area in C and D. ISL1+/TNNI2+/HCN4+ cells are seen in the continuity between sinus venosus myocardium and posterior AV canal myocardium. (E and F) Grey values of ISL1 (E) and HCN4 (F). AVC: atrioventricular canal; LA: left atrium; LB: long bud; LCV: left cardinal vein; LV: left ventricle; OFT: outflow tract; RA: right atrium; RCV: right cardinal vein; SV: sinus venosus; V: ventricle. (C–F) White: TNNI2, green: ISL1, red: HCN4, blue: DAPI; scale bars: 50 μm.

    Techniques Used: Expressing, Staining, In Situ Hybridization

    20) Product Images from "Spatiotemporal Regulation of an Hcn4 Enhancer Defines a Role for Mef2c and HDACs in Cardiac Electrical Patterning"

    Article Title: Spatiotemporal Regulation of an Hcn4 Enhancer Defines a Role for Mef2c and HDACs in Cardiac Electrical Patterning

    Journal: Developmental biology

    doi: 10.1016/j.ydbio.2012.10.017

    Identification of minimal enhancer regions within R2R3 that direct reporter activity in the Hcn4 expression domain
    Figure Legend Snippet: Identification of minimal enhancer regions within R2R3 that direct reporter activity in the Hcn4 expression domain

    Techniques Used: Activity Assay, Expressing

    HDAC Activity Regulates Hcn4 R2R3
    Figure Legend Snippet: HDAC Activity Regulates Hcn4 R2R3

    Techniques Used: Activity Assay

    Mef2C regulates R2R3-LacZ and Hcn4 expression in vivo
    Figure Legend Snippet: Mef2C regulates R2R3-LacZ and Hcn4 expression in vivo

    Techniques Used: Expressing, In Vivo

    Identification and in vivo testing of Hcn4 regulatory elements
    Figure Legend Snippet: Identification and in vivo testing of Hcn4 regulatory elements

    Techniques Used: In Vivo

    In vivo temporal reporter activity directed by the combined R2-R3 Hcn4 Enhancers
    Figure Legend Snippet: In vivo temporal reporter activity directed by the combined R2-R3 Hcn4 Enhancers

    Techniques Used: In Vivo, Activity Assay

    In vivo expression directed by the R2R3 Hcn4 enhancer in the post-natal AV bundle and its interface with the AV Node
    Figure Legend Snippet: In vivo expression directed by the R2R3 Hcn4 enhancer in the post-natal AV bundle and its interface with the AV Node

    Techniques Used: In Vivo, Expressing

    Mef2C regulates R2R3 Hcn4 enhancer activity via a conserved Mef2 Site
    Figure Legend Snippet: Mef2C regulates R2R3 Hcn4 enhancer activity via a conserved Mef2 Site

    Techniques Used: Activity Assay

    Expression pattern of Hcn4 mRNA by whole-mount in-situ hybridization
    Figure Legend Snippet: Expression pattern of Hcn4 mRNA by whole-mount in-situ hybridization

    Techniques Used: Expressing, In Situ Hybridization

    21) Product Images from "Spatiotemporal Regulation of an Hcn4 Enhancer Defines a Role for Mef2c and HDACs in Cardiac Electrical Patterning"

    Article Title: Spatiotemporal Regulation of an Hcn4 Enhancer Defines a Role for Mef2c and HDACs in Cardiac Electrical Patterning

    Journal: Developmental biology

    doi: 10.1016/j.ydbio.2012.10.017

    Identification of minimal enhancer regions within R2R3 that direct reporter activity in the Hcn4 expression domain
    Figure Legend Snippet: Identification of minimal enhancer regions within R2R3 that direct reporter activity in the Hcn4 expression domain

    Techniques Used: Activity Assay, Expressing

    HDAC Activity Regulates Hcn4 R2R3
    Figure Legend Snippet: HDAC Activity Regulates Hcn4 R2R3

    Techniques Used: Activity Assay

    Mef2C regulates R2R3-LacZ and Hcn4 expression in vivo
    Figure Legend Snippet: Mef2C regulates R2R3-LacZ and Hcn4 expression in vivo

    Techniques Used: Expressing, In Vivo

    Identification and in vivo testing of Hcn4 regulatory elements
    Figure Legend Snippet: Identification and in vivo testing of Hcn4 regulatory elements

    Techniques Used: In Vivo

    In vivo temporal reporter activity directed by the combined R2-R3 Hcn4 Enhancers
    Figure Legend Snippet: In vivo temporal reporter activity directed by the combined R2-R3 Hcn4 Enhancers

    Techniques Used: In Vivo, Activity Assay

    In vivo expression directed by the R2R3 Hcn4 enhancer in the post-natal AV bundle and its interface with the AV Node
    Figure Legend Snippet: In vivo expression directed by the R2R3 Hcn4 enhancer in the post-natal AV bundle and its interface with the AV Node

    Techniques Used: In Vivo, Expressing

    Mef2C regulates R2R3 Hcn4 enhancer activity via a conserved Mef2 Site
    Figure Legend Snippet: Mef2C regulates R2R3 Hcn4 enhancer activity via a conserved Mef2 Site

    Techniques Used: Activity Assay

    Expression pattern of Hcn4 mRNA by whole-mount in-situ hybridization
    Figure Legend Snippet: Expression pattern of Hcn4 mRNA by whole-mount in-situ hybridization

    Techniques Used: Expressing, In Situ Hybridization

    22) Product Images from "Spatiotemporal Regulation of an Hcn4 Enhancer Defines a Role for Mef2c and HDACs in Cardiac Electrical Patterning"

    Article Title: Spatiotemporal Regulation of an Hcn4 Enhancer Defines a Role for Mef2c and HDACs in Cardiac Electrical Patterning

    Journal: Developmental biology

    doi: 10.1016/j.ydbio.2012.10.017

    Identification of minimal enhancer regions within R2R3 that direct reporter activity in the Hcn4 expression domain
    Figure Legend Snippet: Identification of minimal enhancer regions within R2R3 that direct reporter activity in the Hcn4 expression domain

    Techniques Used: Activity Assay, Expressing

    HDAC Activity Regulates Hcn4 R2R3
    Figure Legend Snippet: HDAC Activity Regulates Hcn4 R2R3

    Techniques Used: Activity Assay

    Mef2C regulates R2R3-LacZ and Hcn4 expression in vivo
    Figure Legend Snippet: Mef2C regulates R2R3-LacZ and Hcn4 expression in vivo

    Techniques Used: Expressing, In Vivo

    Identification and in vivo testing of Hcn4 regulatory elements
    Figure Legend Snippet: Identification and in vivo testing of Hcn4 regulatory elements

    Techniques Used: In Vivo

    In vivo temporal reporter activity directed by the combined R2-R3 Hcn4 Enhancers
    Figure Legend Snippet: In vivo temporal reporter activity directed by the combined R2-R3 Hcn4 Enhancers

    Techniques Used: In Vivo, Activity Assay

    In vivo expression directed by the R2R3 Hcn4 enhancer in the post-natal AV bundle and its interface with the AV Node
    Figure Legend Snippet: In vivo expression directed by the R2R3 Hcn4 enhancer in the post-natal AV bundle and its interface with the AV Node

    Techniques Used: In Vivo, Expressing

    Mef2C regulates R2R3 Hcn4 enhancer activity via a conserved Mef2 Site
    Figure Legend Snippet: Mef2C regulates R2R3 Hcn4 enhancer activity via a conserved Mef2 Site

    Techniques Used: Activity Assay

    Expression pattern of Hcn4 mRNA by whole-mount in-situ hybridization
    Figure Legend Snippet: Expression pattern of Hcn4 mRNA by whole-mount in-situ hybridization

    Techniques Used: Expressing, In Situ Hybridization

    23) Product Images from "Transcriptome of interstitial cells of Cajal reveals unique and selective gene signatures"

    Article Title: Transcriptome of interstitial cells of Cajal reveals unique and selective gene signatures

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0176031

    Identification of ICC-specific Hcn4 . (A) A genomic map view of Hcn4 variants expressed in JICC and CICC. Three alternative initial exons (V1-3) are circled in red and the differential last exon (E8) is boxed in blue. (B) Expression levels of total Hcn isoform genes in JICC and CICC. (C) Expression levels of Hcn4 transcriptional vaiants in JICC and CICC. (D) A topological map of HCN4 variants. Each circle denotes a single amino acid. Colors on amino acid sequence show distinct regions and domains. Green represents start codons found in alternatively initiated variants (V1-3). Six transmembrane domains (S1-6) and a pore region are shown. Red represe nts voltage sensor residues in S4. Two cAMP binding sites are in purple. (E) Western blot analysis showing HCN4 protein expressed in multiple tissues including stomach, jejunum, and colon muscularis. (F) Cryosection images of HCN4 protein in jejunum and colon. HCN4 was detected in the KIT + serosal layer, myenteric plexus, and deep muscular plexus in jejunum. It was also detected in the serosal layer and submuscular plexus in colon with much lower levels. (G) Whole mount images of HCN4 protein in jejunum (top panels) and colon (bottom panels) showing the protein found in KIT + ICC-SS, ICC-MY, and ICC-DMP in jejunum, and KIT + ICC-SS AND ICC-SMP in colon. All scale bars are 50 μm.
    Figure Legend Snippet: Identification of ICC-specific Hcn4 . (A) A genomic map view of Hcn4 variants expressed in JICC and CICC. Three alternative initial exons (V1-3) are circled in red and the differential last exon (E8) is boxed in blue. (B) Expression levels of total Hcn isoform genes in JICC and CICC. (C) Expression levels of Hcn4 transcriptional vaiants in JICC and CICC. (D) A topological map of HCN4 variants. Each circle denotes a single amino acid. Colors on amino acid sequence show distinct regions and domains. Green represents start codons found in alternatively initiated variants (V1-3). Six transmembrane domains (S1-6) and a pore region are shown. Red represe nts voltage sensor residues in S4. Two cAMP binding sites are in purple. (E) Western blot analysis showing HCN4 protein expressed in multiple tissues including stomach, jejunum, and colon muscularis. (F) Cryosection images of HCN4 protein in jejunum and colon. HCN4 was detected in the KIT + serosal layer, myenteric plexus, and deep muscular plexus in jejunum. It was also detected in the serosal layer and submuscular plexus in colon with much lower levels. (G) Whole mount images of HCN4 protein in jejunum (top panels) and colon (bottom panels) showing the protein found in KIT + ICC-SS, ICC-MY, and ICC-DMP in jejunum, and KIT + ICC-SS AND ICC-SMP in colon. All scale bars are 50 μm.

    Techniques Used: Immunocytochemistry, Expressing, Sequencing, Binding Assay, Western Blot

    24) Product Images from "Transcriptome of interstitial cells of Cajal reveals unique and selective gene signatures"

    Article Title: Transcriptome of interstitial cells of Cajal reveals unique and selective gene signatures

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0176031

    Identification of ICC-specific Hcn4 . (A) A genomic map view of Hcn4 variants expressed in JICC and CICC. Three alternative initial exons (V1-3) are circled in red and the differential last exon (E8) is boxed in blue. (B) Expression levels of total Hcn isoform genes in JICC and CICC. (C) Expression levels of Hcn4 transcriptional vaiants in JICC and CICC. (D) A topological map of HCN4 variants. Each circle denotes a single amino acid. Colors on amino acid sequence show distinct regions and domains. Green represents start codons found in alternatively initiated variants (V1-3). Six transmembrane domains (S1-6) and a pore region are shown. Red represe nts voltage sensor residues in S4. Two cAMP binding sites are in purple. (E) Western blot analysis showing HCN4 protein expressed in multiple tissues including stomach, jejunum, and colon muscularis. (F) Cryosection images of HCN4 protein in jejunum and colon. HCN4 was detected in the KIT + serosal layer, myenteric plexus, and deep muscular plexus in jejunum. It was also detected in the serosal layer and submuscular plexus in colon with much lower levels. (G) Whole mount images of HCN4 protein in jejunum (top panels) and colon (bottom panels) showing the protein found in KIT + ICC-SS, ICC-MY, and ICC-DMP in jejunum, and KIT + ICC-SS AND ICC-SMP in colon. All scale bars are 50 μm.
    Figure Legend Snippet: Identification of ICC-specific Hcn4 . (A) A genomic map view of Hcn4 variants expressed in JICC and CICC. Three alternative initial exons (V1-3) are circled in red and the differential last exon (E8) is boxed in blue. (B) Expression levels of total Hcn isoform genes in JICC and CICC. (C) Expression levels of Hcn4 transcriptional vaiants in JICC and CICC. (D) A topological map of HCN4 variants. Each circle denotes a single amino acid. Colors on amino acid sequence show distinct regions and domains. Green represents start codons found in alternatively initiated variants (V1-3). Six transmembrane domains (S1-6) and a pore region are shown. Red represe nts voltage sensor residues in S4. Two cAMP binding sites are in purple. (E) Western blot analysis showing HCN4 protein expressed in multiple tissues including stomach, jejunum, and colon muscularis. (F) Cryosection images of HCN4 protein in jejunum and colon. HCN4 was detected in the KIT + serosal layer, myenteric plexus, and deep muscular plexus in jejunum. It was also detected in the serosal layer and submuscular plexus in colon with much lower levels. (G) Whole mount images of HCN4 protein in jejunum (top panels) and colon (bottom panels) showing the protein found in KIT + ICC-SS, ICC-MY, and ICC-DMP in jejunum, and KIT + ICC-SS AND ICC-SMP in colon. All scale bars are 50 μm.

    Techniques Used: Immunocytochemistry, Expressing, Sequencing, Binding Assay, Western Blot

    25) Product Images from "Spatiotemporal Regulation of an Hcn4 Enhancer Defines a Role for Mef2c and HDACs in Cardiac Electrical Patterning"

    Article Title: Spatiotemporal Regulation of an Hcn4 Enhancer Defines a Role for Mef2c and HDACs in Cardiac Electrical Patterning

    Journal: Developmental biology

    doi: 10.1016/j.ydbio.2012.10.017

    Identification of minimal enhancer regions within R2R3 that direct reporter activity in the Hcn4 expression domain
    Figure Legend Snippet: Identification of minimal enhancer regions within R2R3 that direct reporter activity in the Hcn4 expression domain

    Techniques Used: Activity Assay, Expressing

    HDAC Activity Regulates Hcn4 R2R3
    Figure Legend Snippet: HDAC Activity Regulates Hcn4 R2R3

    Techniques Used: Activity Assay

    Mef2C regulates R2R3-LacZ and Hcn4 expression in vivo
    Figure Legend Snippet: Mef2C regulates R2R3-LacZ and Hcn4 expression in vivo

    Techniques Used: Expressing, In Vivo

    Identification and in vivo testing of Hcn4 regulatory elements
    Figure Legend Snippet: Identification and in vivo testing of Hcn4 regulatory elements

    Techniques Used: In Vivo

    In vivo temporal reporter activity directed by the combined R2-R3 Hcn4 Enhancers
    Figure Legend Snippet: In vivo temporal reporter activity directed by the combined R2-R3 Hcn4 Enhancers

    Techniques Used: In Vivo, Activity Assay

    In vivo expression directed by the R2R3 Hcn4 enhancer in the post-natal AV bundle and its interface with the AV Node
    Figure Legend Snippet: In vivo expression directed by the R2R3 Hcn4 enhancer in the post-natal AV bundle and its interface with the AV Node

    Techniques Used: In Vivo, Expressing

    Mef2C regulates R2R3 Hcn4 enhancer activity via a conserved Mef2 Site
    Figure Legend Snippet: Mef2C regulates R2R3 Hcn4 enhancer activity via a conserved Mef2 Site

    Techniques Used: Activity Assay

    Expression pattern of Hcn4 mRNA by whole-mount in-situ hybridization
    Figure Legend Snippet: Expression pattern of Hcn4 mRNA by whole-mount in-situ hybridization

    Techniques Used: Expressing, In Situ Hybridization

    26) Product Images from "Spatiotemporal Regulation of an Hcn4 Enhancer Defines a Role for Mef2c and HDACs in Cardiac Electrical Patterning"

    Article Title: Spatiotemporal Regulation of an Hcn4 Enhancer Defines a Role for Mef2c and HDACs in Cardiac Electrical Patterning

    Journal: Developmental biology

    doi: 10.1016/j.ydbio.2012.10.017

    Identification of minimal enhancer regions within R2R3 that direct reporter activity in the Hcn4 expression domain
    Figure Legend Snippet: Identification of minimal enhancer regions within R2R3 that direct reporter activity in the Hcn4 expression domain

    Techniques Used: Activity Assay, Expressing

    HDAC Activity Regulates Hcn4 R2R3
    Figure Legend Snippet: HDAC Activity Regulates Hcn4 R2R3

    Techniques Used: Activity Assay

    Mef2C regulates R2R3-LacZ and Hcn4 expression in vivo
    Figure Legend Snippet: Mef2C regulates R2R3-LacZ and Hcn4 expression in vivo

    Techniques Used: Expressing, In Vivo

    Identification and in vivo testing of Hcn4 regulatory elements
    Figure Legend Snippet: Identification and in vivo testing of Hcn4 regulatory elements

    Techniques Used: In Vivo

    In vivo temporal reporter activity directed by the combined R2-R3 Hcn4 Enhancers
    Figure Legend Snippet: In vivo temporal reporter activity directed by the combined R2-R3 Hcn4 Enhancers

    Techniques Used: In Vivo, Activity Assay

    In vivo expression directed by the R2R3 Hcn4 enhancer in the post-natal AV bundle and its interface with the AV Node
    Figure Legend Snippet: In vivo expression directed by the R2R3 Hcn4 enhancer in the post-natal AV bundle and its interface with the AV Node

    Techniques Used: In Vivo, Expressing

    Mef2C regulates R2R3 Hcn4 enhancer activity via a conserved Mef2 Site
    Figure Legend Snippet: Mef2C regulates R2R3 Hcn4 enhancer activity via a conserved Mef2 Site

    Techniques Used: Activity Assay

    Expression pattern of Hcn4 mRNA by whole-mount in-situ hybridization
    Figure Legend Snippet: Expression pattern of Hcn4 mRNA by whole-mount in-situ hybridization

    Techniques Used: Expressing, In Situ Hybridization

    27) Product Images from "The sinus venosus myocardium contributes to the atrioventricular canal: potential role during atrioventricular node development?"

    Article Title: The sinus venosus myocardium contributes to the atrioventricular canal: potential role during atrioventricular node development?

    Journal: Journal of Cellular and Molecular Medicine

    doi: 10.1111/jcmm.12525

    Characterization of the myocardial continuity between the sinus venosus and AV canal. (A) Whole mount ISL1 staining, posterior view of the heart. Arrows indicate ISL1+ cells in continuity sinus venosus and AV canal. Rectangle 1 shows region from which SV-AVC continuity cells were collected. Rectangle 2 shows origin of right ventricular cells for patch clamp. (B–E) mRNA expression after laser capture microscopy (LCM) of TNNT2, NKX2-5, HCN4 and ISL1 . (F–M) Results of single cell patch-clamp experiments. (F and J) Representative tracing of SV-AVC continuity cells (F, pacemaker-like tracing) and RV cells (J, ventricular-like phenotype). (G–I, K–M) Graphs showing the difference in maximum upstroke velocity (G), frequency (H), amplitude (I), APD50 (K), APD90 (L) and maximum diastolic potential (M). * Indicates P
    Figure Legend Snippet: Characterization of the myocardial continuity between the sinus venosus and AV canal. (A) Whole mount ISL1 staining, posterior view of the heart. Arrows indicate ISL1+ cells in continuity sinus venosus and AV canal. Rectangle 1 shows region from which SV-AVC continuity cells were collected. Rectangle 2 shows origin of right ventricular cells for patch clamp. (B–E) mRNA expression after laser capture microscopy (LCM) of TNNT2, NKX2-5, HCN4 and ISL1 . (F–M) Results of single cell patch-clamp experiments. (F and J) Representative tracing of SV-AVC continuity cells (F, pacemaker-like tracing) and RV cells (J, ventricular-like phenotype). (G–I, K–M) Graphs showing the difference in maximum upstroke velocity (G), frequency (H), amplitude (I), APD50 (K), APD90 (L) and maximum diastolic potential (M). * Indicates P

    Techniques Used: Staining, Patch Clamp, Expressing, Microscopy, Laser Capture Microdissection

    HCN4 expression in myocardial continuity between sinus venosus and posterior AV canal. (A) TNNI2 stain of HH19 chick embryo. Arrow indicates continuity between sinus venosus myocardium and posterior AV canal. (B) ISH shows HCN4 mRNA expression in continuity (arrow). (C) Merge of ISL1, TNNI2, HCN4 and DAPI staining of E11.5 mouse embryo, showing the myocardial continuity between the sinus venosus and posterior AV canal. The ISL1+/HCN4+/TNNI2+ SAN (arrowhead) and RVV (arrow) are shown. Note the thick endocardial cushions (asterisks) in the AVC. (D–F) Higher magnifications of boxed area in C and D. ISL1+/TNNI2+/HCN4+ cells are seen in the continuity between sinus venosus myocardium and posterior AV canal myocardium. (E and F) Grey values of ISL1 (E) and HCN4 (F). AVC: atrioventricular canal; LA: left atrium; LB: long bud; LCV: left cardinal vein; LV: left ventricle; OFT: outflow tract; RA: right atrium; RCV: right cardinal vein; SV: sinus venosus; V: ventricle. (C–F) White: TNNI2, green: ISL1, red: HCN4, blue: DAPI; scale bars: 50 μm.
    Figure Legend Snippet: HCN4 expression in myocardial continuity between sinus venosus and posterior AV canal. (A) TNNI2 stain of HH19 chick embryo. Arrow indicates continuity between sinus venosus myocardium and posterior AV canal. (B) ISH shows HCN4 mRNA expression in continuity (arrow). (C) Merge of ISL1, TNNI2, HCN4 and DAPI staining of E11.5 mouse embryo, showing the myocardial continuity between the sinus venosus and posterior AV canal. The ISL1+/HCN4+/TNNI2+ SAN (arrowhead) and RVV (arrow) are shown. Note the thick endocardial cushions (asterisks) in the AVC. (D–F) Higher magnifications of boxed area in C and D. ISL1+/TNNI2+/HCN4+ cells are seen in the continuity between sinus venosus myocardium and posterior AV canal myocardium. (E and F) Grey values of ISL1 (E) and HCN4 (F). AVC: atrioventricular canal; LA: left atrium; LB: long bud; LCV: left cardinal vein; LV: left ventricle; OFT: outflow tract; RA: right atrium; RCV: right cardinal vein; SV: sinus venosus; V: ventricle. (C–F) White: TNNI2, green: ISL1, red: HCN4, blue: DAPI; scale bars: 50 μm.

    Techniques Used: Expressing, Staining, In Situ Hybridization

    28) Product Images from "Spatiotemporal Regulation of an Hcn4 Enhancer Defines a Role for Mef2c and HDACs in Cardiac Electrical Patterning"

    Article Title: Spatiotemporal Regulation of an Hcn4 Enhancer Defines a Role for Mef2c and HDACs in Cardiac Electrical Patterning

    Journal: Developmental biology

    doi: 10.1016/j.ydbio.2012.10.017

    Identification of minimal enhancer regions within R2R3 that direct reporter activity in the Hcn4 expression domain
    Figure Legend Snippet: Identification of minimal enhancer regions within R2R3 that direct reporter activity in the Hcn4 expression domain

    Techniques Used: Activity Assay, Expressing

    HDAC Activity Regulates Hcn4 R2R3
    Figure Legend Snippet: HDAC Activity Regulates Hcn4 R2R3

    Techniques Used: Activity Assay

    Mef2C regulates R2R3-LacZ and Hcn4 expression in vivo
    Figure Legend Snippet: Mef2C regulates R2R3-LacZ and Hcn4 expression in vivo

    Techniques Used: Expressing, In Vivo

    Identification and in vivo testing of Hcn4 regulatory elements
    Figure Legend Snippet: Identification and in vivo testing of Hcn4 regulatory elements

    Techniques Used: In Vivo

    In vivo temporal reporter activity directed by the combined R2-R3 Hcn4 Enhancers
    Figure Legend Snippet: In vivo temporal reporter activity directed by the combined R2-R3 Hcn4 Enhancers

    Techniques Used: In Vivo, Activity Assay

    In vivo expression directed by the R2R3 Hcn4 enhancer in the post-natal AV bundle and its interface with the AV Node
    Figure Legend Snippet: In vivo expression directed by the R2R3 Hcn4 enhancer in the post-natal AV bundle and its interface with the AV Node

    Techniques Used: In Vivo, Expressing

    Mef2C regulates R2R3 Hcn4 enhancer activity via a conserved Mef2 Site
    Figure Legend Snippet: Mef2C regulates R2R3 Hcn4 enhancer activity via a conserved Mef2 Site

    Techniques Used: Activity Assay

    Expression pattern of Hcn4 mRNA by whole-mount in-situ hybridization
    Figure Legend Snippet: Expression pattern of Hcn4 mRNA by whole-mount in-situ hybridization

    Techniques Used: Expressing, In Situ Hybridization

    29) Product Images from "Spatiotemporal Regulation of an Hcn4 Enhancer Defines a Role for Mef2c and HDACs in Cardiac Electrical Patterning"

    Article Title: Spatiotemporal Regulation of an Hcn4 Enhancer Defines a Role for Mef2c and HDACs in Cardiac Electrical Patterning

    Journal: Developmental biology

    doi: 10.1016/j.ydbio.2012.10.017

    Identification of minimal enhancer regions within R2R3 that direct reporter activity in the Hcn4 expression domain
    Figure Legend Snippet: Identification of minimal enhancer regions within R2R3 that direct reporter activity in the Hcn4 expression domain

    Techniques Used: Activity Assay, Expressing

    HDAC Activity Regulates Hcn4 R2R3
    Figure Legend Snippet: HDAC Activity Regulates Hcn4 R2R3

    Techniques Used: Activity Assay

    Mef2C regulates R2R3-LacZ and Hcn4 expression in vivo
    Figure Legend Snippet: Mef2C regulates R2R3-LacZ and Hcn4 expression in vivo

    Techniques Used: Expressing, In Vivo

    Identification and in vivo testing of Hcn4 regulatory elements
    Figure Legend Snippet: Identification and in vivo testing of Hcn4 regulatory elements

    Techniques Used: In Vivo

    In vivo temporal reporter activity directed by the combined R2-R3 Hcn4 Enhancers
    Figure Legend Snippet: In vivo temporal reporter activity directed by the combined R2-R3 Hcn4 Enhancers

    Techniques Used: In Vivo, Activity Assay

    In vivo expression directed by the R2R3 Hcn4 enhancer in the post-natal AV bundle and its interface with the AV Node
    Figure Legend Snippet: In vivo expression directed by the R2R3 Hcn4 enhancer in the post-natal AV bundle and its interface with the AV Node

    Techniques Used: In Vivo, Expressing

    Mef2C regulates R2R3 Hcn4 enhancer activity via a conserved Mef2 Site
    Figure Legend Snippet: Mef2C regulates R2R3 Hcn4 enhancer activity via a conserved Mef2 Site

    Techniques Used: Activity Assay

    Expression pattern of Hcn4 mRNA by whole-mount in-situ hybridization
    Figure Legend Snippet: Expression pattern of Hcn4 mRNA by whole-mount in-situ hybridization

    Techniques Used: Expressing, In Situ Hybridization

    30) Product Images from "Spatiotemporal Regulation of an Hcn4 Enhancer Defines a Role for Mef2c and HDACs in Cardiac Electrical Patterning"

    Article Title: Spatiotemporal Regulation of an Hcn4 Enhancer Defines a Role for Mef2c and HDACs in Cardiac Electrical Patterning

    Journal: Developmental biology

    doi: 10.1016/j.ydbio.2012.10.017

    Identification of minimal enhancer regions within R2R3 that direct reporter activity in the Hcn4 expression domain
    Figure Legend Snippet: Identification of minimal enhancer regions within R2R3 that direct reporter activity in the Hcn4 expression domain

    Techniques Used: Activity Assay, Expressing

    HDAC Activity Regulates Hcn4 R2R3
    Figure Legend Snippet: HDAC Activity Regulates Hcn4 R2R3

    Techniques Used: Activity Assay

    Mef2C regulates R2R3-LacZ and Hcn4 expression in vivo
    Figure Legend Snippet: Mef2C regulates R2R3-LacZ and Hcn4 expression in vivo

    Techniques Used: Expressing, In Vivo

    Identification and in vivo testing of Hcn4 regulatory elements
    Figure Legend Snippet: Identification and in vivo testing of Hcn4 regulatory elements

    Techniques Used: In Vivo

    In vivo temporal reporter activity directed by the combined R2-R3 Hcn4 Enhancers
    Figure Legend Snippet: In vivo temporal reporter activity directed by the combined R2-R3 Hcn4 Enhancers

    Techniques Used: In Vivo, Activity Assay

    In vivo expression directed by the R2R3 Hcn4 enhancer in the post-natal AV bundle and its interface with the AV Node
    Figure Legend Snippet: In vivo expression directed by the R2R3 Hcn4 enhancer in the post-natal AV bundle and its interface with the AV Node

    Techniques Used: In Vivo, Expressing

    Mef2C regulates R2R3 Hcn4 enhancer activity via a conserved Mef2 Site
    Figure Legend Snippet: Mef2C regulates R2R3 Hcn4 enhancer activity via a conserved Mef2 Site

    Techniques Used: Activity Assay

    Expression pattern of Hcn4 mRNA by whole-mount in-situ hybridization
    Figure Legend Snippet: Expression pattern of Hcn4 mRNA by whole-mount in-situ hybridization

    Techniques Used: Expressing, In Situ Hybridization

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    Alomone Labs rabbit anti hcn4
    The tdTomato-expressing retinal bipolar cells in the 5-HTR2a-cre mouse line are co-labeled with antibodies specific to type 4 and type 3b cone bipolar cells, and rod bipolar cells. A – C : In a retinal vertical section, the tdTomato-expressing retina ( A ) was immunostained for calsenilin ( B ). The overlay of A and B is shown in C . The double-positive bipolar cells are marked with stars. D – F : In a retinal whole mount with the focal plane at the distal portion of the inner nuclear layer (INL), the tdTomato-expressing retina ( D ) was immunostained for calsenilin ( E ). The overlay of D and E is shown in F . The majority of the tomato-expressing cells in the distal portion of the INL are calsenilin-positive. The tdTomato-expressing cells that do not show calsenilin staining are marked with arrowheads. The tdTomato-expressing retina was immunostained for PKCα in a retinal vertical section ( G–I ). The double-positive bipolar cells are marked with arrows in the somata and arrowheads pointing to the axon terminals. J – L : The tdTomato-expressing retina was immunostained for PKARIIβ. Two double-positive cells are marked with stars. The tdTomato-expressing retinal bipolar cells were not labeled by antibodies for <t>HCN4</t> ( M – O ) and Syt2 ( P – R ). Scale bars represent 50 µm. ONL, outer nuclear layer; IPL, inner plexiform layer; GCL, ganglion cell layer.
    Rabbit Anti Hcn4, supplied by Alomone Labs, used in various techniques. Bioz Stars score: 95/100, based on 6 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    The tdTomato-expressing retinal bipolar cells in the 5-HTR2a-cre mouse line are co-labeled with antibodies specific to type 4 and type 3b cone bipolar cells, and rod bipolar cells. A – C : In a retinal vertical section, the tdTomato-expressing retina ( A ) was immunostained for calsenilin ( B ). The overlay of A and B is shown in C . The double-positive bipolar cells are marked with stars. D – F : In a retinal whole mount with the focal plane at the distal portion of the inner nuclear layer (INL), the tdTomato-expressing retina ( D ) was immunostained for calsenilin ( E ). The overlay of D and E is shown in F . The majority of the tomato-expressing cells in the distal portion of the INL are calsenilin-positive. The tdTomato-expressing cells that do not show calsenilin staining are marked with arrowheads. The tdTomato-expressing retina was immunostained for PKCα in a retinal vertical section ( G–I ). The double-positive bipolar cells are marked with arrows in the somata and arrowheads pointing to the axon terminals. J – L : The tdTomato-expressing retina was immunostained for PKARIIβ. Two double-positive cells are marked with stars. The tdTomato-expressing retinal bipolar cells were not labeled by antibodies for HCN4 ( M – O ) and Syt2 ( P – R ). Scale bars represent 50 µm. ONL, outer nuclear layer; IPL, inner plexiform layer; GCL, ganglion cell layer.

    Journal: Molecular Vision

    Article Title: Cre-mediated recombination efficiency and transgene expression patterns of three retinal bipolar cell-expressing Cre transgenic mouse lines

    doi:

    Figure Lengend Snippet: The tdTomato-expressing retinal bipolar cells in the 5-HTR2a-cre mouse line are co-labeled with antibodies specific to type 4 and type 3b cone bipolar cells, and rod bipolar cells. A – C : In a retinal vertical section, the tdTomato-expressing retina ( A ) was immunostained for calsenilin ( B ). The overlay of A and B is shown in C . The double-positive bipolar cells are marked with stars. D – F : In a retinal whole mount with the focal plane at the distal portion of the inner nuclear layer (INL), the tdTomato-expressing retina ( D ) was immunostained for calsenilin ( E ). The overlay of D and E is shown in F . The majority of the tomato-expressing cells in the distal portion of the INL are calsenilin-positive. The tdTomato-expressing cells that do not show calsenilin staining are marked with arrowheads. The tdTomato-expressing retina was immunostained for PKCα in a retinal vertical section ( G–I ). The double-positive bipolar cells are marked with arrows in the somata and arrowheads pointing to the axon terminals. J – L : The tdTomato-expressing retina was immunostained for PKARIIβ. Two double-positive cells are marked with stars. The tdTomato-expressing retinal bipolar cells were not labeled by antibodies for HCN4 ( M – O ) and Syt2 ( P – R ). Scale bars represent 50 µm. ONL, outer nuclear layer; IPL, inner plexiform layer; GCL, ganglion cell layer.

    Article Snippet: The following antibodies were used in this study: rabbit anti-mCherry (1:500; 632,496, Clontech, Mountain View, CA); mouse anticalsenilin (1:2,000; kindly provided by W. Wasco, Harvard Medical School, Boston, MA); rabbit anti-PKC (1:20,000; catalog number 2056, Cell Signal, Danvers, MA); mouse anti-PKC (1:10000; catalog number sc8393, Santa Cruz, CA); mouse antisynaptotagmin II (Syt2; 1:600; Zebrafish International Resource Center, Eugene, OR); rabbit anti-HCN4 (1:500; Alomone Labs, Jerusalem, Israel); and mouse antiprotein kinase A (PKA) RIIβ (1:80,000; BD Biosciences, San Jose, CA).

    Techniques: Expressing, Labeling, Staining

    The tdTomato-expressing retinal bipolar cells in the Pcp2-cre mouse line are co-labeled with antibodies specific to rod bipolar cells, type 2 and 6 cone bipolar cells. A – C : In retinal vertical sections, the tdTomato-expressing retina ( A ) was immunostained for PKCα ( B ). The overlay of A and B is shown in C . The double-positive bipolar cells were marked with stars in the somata and with arrowheads pointing at the axon terminals. D–F : In the retinal whole mount with the focal plane in the INL, colabeling with tdTomato and PKCα. PKCα-negative tdTomato-expressing cells are marked with arrows. G–I : The tdTomato-expressing retina was immunostained for Syt2. The double-positive bipolar cells with axon terminals stratified at the distal portion of the IPL (type 2 bipolar cells) are marked with white stars in the somata and with white arrowheads pointing at the axon terminals. The double-positive bipolar cells with axon terminals stratified in the proximal portion of the IPL (type 6 bipolar cells) are marked with yellow arrows at the somata and yellow arrowheads at the axon terminals. A tdTomato-expressing bipolar cell with their axon terminals stratified slightly distal to Syt2-positive cells is marked with a blue arrowhead ( I ). The tdTomato-expressing retinal bipolar cells were not found to be labeled by PKARIIβ ( J – L ), HCN4 ( M – O ), and calsenilin ( P – R ). Scale bars represent 50 µm. ONL, outer nuclear layer; INL, inner nuclear layer; IPL, inner plexiform layer; GCL, ganglion cell layer.

    Journal: Molecular Vision

    Article Title: Cre-mediated recombination efficiency and transgene expression patterns of three retinal bipolar cell-expressing Cre transgenic mouse lines

    doi:

    Figure Lengend Snippet: The tdTomato-expressing retinal bipolar cells in the Pcp2-cre mouse line are co-labeled with antibodies specific to rod bipolar cells, type 2 and 6 cone bipolar cells. A – C : In retinal vertical sections, the tdTomato-expressing retina ( A ) was immunostained for PKCα ( B ). The overlay of A and B is shown in C . The double-positive bipolar cells were marked with stars in the somata and with arrowheads pointing at the axon terminals. D–F : In the retinal whole mount with the focal plane in the INL, colabeling with tdTomato and PKCα. PKCα-negative tdTomato-expressing cells are marked with arrows. G–I : The tdTomato-expressing retina was immunostained for Syt2. The double-positive bipolar cells with axon terminals stratified at the distal portion of the IPL (type 2 bipolar cells) are marked with white stars in the somata and with white arrowheads pointing at the axon terminals. The double-positive bipolar cells with axon terminals stratified in the proximal portion of the IPL (type 6 bipolar cells) are marked with yellow arrows at the somata and yellow arrowheads at the axon terminals. A tdTomato-expressing bipolar cell with their axon terminals stratified slightly distal to Syt2-positive cells is marked with a blue arrowhead ( I ). The tdTomato-expressing retinal bipolar cells were not found to be labeled by PKARIIβ ( J – L ), HCN4 ( M – O ), and calsenilin ( P – R ). Scale bars represent 50 µm. ONL, outer nuclear layer; INL, inner nuclear layer; IPL, inner plexiform layer; GCL, ganglion cell layer.

    Article Snippet: The following antibodies were used in this study: rabbit anti-mCherry (1:500; 632,496, Clontech, Mountain View, CA); mouse anticalsenilin (1:2,000; kindly provided by W. Wasco, Harvard Medical School, Boston, MA); rabbit anti-PKC (1:20,000; catalog number 2056, Cell Signal, Danvers, MA); mouse anti-PKC (1:10000; catalog number sc8393, Santa Cruz, CA); mouse antisynaptotagmin II (Syt2; 1:600; Zebrafish International Resource Center, Eugene, OR); rabbit anti-HCN4 (1:500; Alomone Labs, Jerusalem, Israel); and mouse antiprotein kinase A (PKA) RIIβ (1:80,000; BD Biosciences, San Jose, CA).

    Techniques: Expressing, Labeling

    Panels A and B : Optical slices (1 μm thick) of HCN4 immunoreactive cells at a depth of 30 μm and 60 μm from the endocardial surface. Panel C : a 3D reconstruction of 70 stacked HCN4 images (optically sliced via confocal microscope) of an immunolabeled whole-mount SAN preparation demonstrating the distribution of HCN4 immunoreactive cells within a depth of 70 μM from the endocardium. Tissue depth from the endocardial site is color coded on the right side of the panel.

    Journal: bioRxiv

    Article Title: Synchronized cardiac impulses emerge from multi-scale, heterogeneous local calcium signals within and among cells of heart pacemaker tissue

    doi: 10.1101/2020.04.14.039461

    Figure Lengend Snippet: Panels A and B : Optical slices (1 μm thick) of HCN4 immunoreactive cells at a depth of 30 μm and 60 μm from the endocardial surface. Panel C : a 3D reconstruction of 70 stacked HCN4 images (optically sliced via confocal microscope) of an immunolabeled whole-mount SAN preparation demonstrating the distribution of HCN4 immunoreactive cells within a depth of 70 μM from the endocardium. Tissue depth from the endocardial site is color coded on the right side of the panel.

    Article Snippet: Antibodies: HCN4+ cells were identified by rabbit polyclonal antibodies for hyperpolarization-activated, cyclic nucleotide-gate cation channels HCN4 (1:250; Alomone Labs).

    Techniques: Microscopy, Immunolabeling

    A dual immunolabeled HCN4 (red) and CX43 (green) together with F-actin labelling (cyan) merged into a single image is shown in both panels. Panel A: Stacked confocal images and reconstructed front view of optically sliced z-stack images of CX43 at a depth of 35 μm from endocardium. Gap junctions are color coded by depth and plotted within the z-stacks reconstructed from optical slices. Confocal images were acquired with a 40x oil immersion. Panel B: Stacked confocal images and reconstructed side view of optically sliced z-stack images of HCN4 + /F-actin - /(CX43) - cells (red) and HCN4 - /F-actin + /(CX43) + cells (cyan) at a depth of 40 μm and 50 μm from endocardium. Confocal images were acquired with a 40x oil immersion objective Panel C: Optical slice shows HCN4 + /F-actin - /(CX43) - cells (red) adjacent to HCN4 - /F-actin + /(CX43) + cells (cyan). CX43 protein (green) is expressed only in cyan cells.

    Journal: bioRxiv

    Article Title: Synchronized cardiac impulses emerge from multi-scale, heterogeneous local calcium signals within and among cells of heart pacemaker tissue

    doi: 10.1101/2020.04.14.039461

    Figure Lengend Snippet: A dual immunolabeled HCN4 (red) and CX43 (green) together with F-actin labelling (cyan) merged into a single image is shown in both panels. Panel A: Stacked confocal images and reconstructed front view of optically sliced z-stack images of CX43 at a depth of 35 μm from endocardium. Gap junctions are color coded by depth and plotted within the z-stacks reconstructed from optical slices. Confocal images were acquired with a 40x oil immersion. Panel B: Stacked confocal images and reconstructed side view of optically sliced z-stack images of HCN4 + /F-actin - /(CX43) - cells (red) and HCN4 - /F-actin + /(CX43) + cells (cyan) at a depth of 40 μm and 50 μm from endocardium. Confocal images were acquired with a 40x oil immersion objective Panel C: Optical slice shows HCN4 + /F-actin - /(CX43) - cells (red) adjacent to HCN4 - /F-actin + /(CX43) + cells (cyan). CX43 protein (green) is expressed only in cyan cells.

    Article Snippet: Antibodies: HCN4+ cells were identified by rabbit polyclonal antibodies for hyperpolarization-activated, cyclic nucleotide-gate cation channels HCN4 (1:250; Alomone Labs).

    Techniques: Immunolabeling

    Panel A and B: A dual immunolabeled HCN4 (red) and CX43 (green) together with F-actin labelling (cyan) merged into a single image is shown in both panels. Panel A: Spatial cytoarchitecture of SAN within whole mount preparations reconstructed from 36 tiled confocal images of the area of the 1350μm by 1350μm demonstrate meshwork (red)/network(cyan) intertwining. Green dots are immunolabelled CX43 proteins. Gray color tissue was imaged in transmitted light. Panel B: Spatial cytoarchitecture of SAN within whole mount preparations reconstructed from 16 tiled confocal images of the area of the 900μm by 900μm of another area of meshwork (red)/network(cyan) intertwining. Green dots are immunolabelled CX43 proteins. CX43 is detected only in F-actin labeled cells.

    Journal: bioRxiv

    Article Title: Synchronized cardiac impulses emerge from multi-scale, heterogeneous local calcium signals within and among cells of heart pacemaker tissue

    doi: 10.1101/2020.04.14.039461

    Figure Lengend Snippet: Panel A and B: A dual immunolabeled HCN4 (red) and CX43 (green) together with F-actin labelling (cyan) merged into a single image is shown in both panels. Panel A: Spatial cytoarchitecture of SAN within whole mount preparations reconstructed from 36 tiled confocal images of the area of the 1350μm by 1350μm demonstrate meshwork (red)/network(cyan) intertwining. Green dots are immunolabelled CX43 proteins. Gray color tissue was imaged in transmitted light. Panel B: Spatial cytoarchitecture of SAN within whole mount preparations reconstructed from 16 tiled confocal images of the area of the 900μm by 900μm of another area of meshwork (red)/network(cyan) intertwining. Green dots are immunolabelled CX43 proteins. CX43 is detected only in F-actin labeled cells.

    Article Snippet: Antibodies: HCN4+ cells were identified by rabbit polyclonal antibodies for hyperpolarization-activated, cyclic nucleotide-gate cation channels HCN4 (1:250; Alomone Labs).

    Techniques: Immunolabeling, Labeling

    Panel A : An immunolabelled, whole mount image of a SAN preparation at low (2.5x) optical magnification. Panel B: Stacked confocal images and reconstructed front view of optically sliced z-stack images of HCN4 + /F-actin - cells (red) and HCN4 - /F-actin + cells (cyan) at a depth of 30 μm from endocardium. Confocal images were acquired with a 40x oil immersion objective within ROI (yellow) shown in panel A. Panel C: Side views of the z-stack images in panel B, illustrating the intertwining cells of HCN4-meshwork and F-actin networks across the 30 μm depth.

    Journal: bioRxiv

    Article Title: Synchronized cardiac impulses emerge from multi-scale, heterogeneous local calcium signals within and among cells of heart pacemaker tissue

    doi: 10.1101/2020.04.14.039461

    Figure Lengend Snippet: Panel A : An immunolabelled, whole mount image of a SAN preparation at low (2.5x) optical magnification. Panel B: Stacked confocal images and reconstructed front view of optically sliced z-stack images of HCN4 + /F-actin - cells (red) and HCN4 - /F-actin + cells (cyan) at a depth of 30 μm from endocardium. Confocal images were acquired with a 40x oil immersion objective within ROI (yellow) shown in panel A. Panel C: Side views of the z-stack images in panel B, illustrating the intertwining cells of HCN4-meshwork and F-actin networks across the 30 μm depth.

    Article Snippet: Antibodies: HCN4+ cells were identified by rabbit polyclonal antibodies for hyperpolarization-activated, cyclic nucleotide-gate cation channels HCN4 (1:250; Alomone Labs).

    Techniques:

    Panel A: An Image of a whole mount SAN preparation at low (2.5x) optical magnification, demonstrating the distribution of HCN4 (red color) immunoreactive and F-actin (cyan color) labelled cells. The merged images between HCN4 and F-actin is shown in both panels. Panel B: Tiled Image of the HCN4 + /F-actin - cell meshwork (red) intertwined with the HCN4 - /F-actin + cell network (cyan) reconstructed from 4 images recorded via 10x water immersion objective within the red box in panel A.

    Journal: bioRxiv

    Article Title: Synchronized cardiac impulses emerge from multi-scale, heterogeneous local calcium signals within and among cells of heart pacemaker tissue

    doi: 10.1101/2020.04.14.039461

    Figure Lengend Snippet: Panel A: An Image of a whole mount SAN preparation at low (2.5x) optical magnification, demonstrating the distribution of HCN4 (red color) immunoreactive and F-actin (cyan color) labelled cells. The merged images between HCN4 and F-actin is shown in both panels. Panel B: Tiled Image of the HCN4 + /F-actin - cell meshwork (red) intertwined with the HCN4 - /F-actin + cell network (cyan) reconstructed from 4 images recorded via 10x water immersion objective within the red box in panel A.

    Article Snippet: Antibodies: HCN4+ cells were identified by rabbit polyclonal antibodies for hyperpolarization-activated, cyclic nucleotide-gate cation channels HCN4 (1:250; Alomone Labs).

    Techniques:

    Upper panels - HCN4 immunoreactive SAN cells: elongated (magenta arrows in Panel A ), novel, pyramidal-like shape cells (yellow arrow in Panels B and D ), spider-like ( Panel C ), and spindle cells ( Panel E , blue arrow). Lower panels - SAN cells loaded with Fluo-4 AM have similar shapes to immunolabelled HCN4 + cells in the upper panel. Spider-like cells ( Panel F ) are indicated by the red arrow. Novel cells with a pyramidal-like soma ( Panel G ) are indicated by yellow arrows. Spindle cells are indicated by the blue arrow ( Panel G and H ). Elongated cells ( Panel H ) are indicated by magenta arrows.

    Journal: bioRxiv

    Article Title: Synchronized cardiac impulses emerge from multi-scale, heterogeneous local calcium signals within and among cells of heart pacemaker tissue

    doi: 10.1101/2020.04.14.039461

    Figure Lengend Snippet: Upper panels - HCN4 immunoreactive SAN cells: elongated (magenta arrows in Panel A ), novel, pyramidal-like shape cells (yellow arrow in Panels B and D ), spider-like ( Panel C ), and spindle cells ( Panel E , blue arrow). Lower panels - SAN cells loaded with Fluo-4 AM have similar shapes to immunolabelled HCN4 + cells in the upper panel. Spider-like cells ( Panel F ) are indicated by the red arrow. Novel cells with a pyramidal-like soma ( Panel G ) are indicated by yellow arrows. Spindle cells are indicated by the blue arrow ( Panel G and H ). Elongated cells ( Panel H ) are indicated by magenta arrows.

    Article Snippet: Antibodies: HCN4+ cells were identified by rabbit polyclonal antibodies for hyperpolarization-activated, cyclic nucleotide-gate cation channels HCN4 (1:250; Alomone Labs).

    Techniques:

    Panel A: A dual immunolabeled HCN4 (red) and CX43 (green) whole mount SAN image at low optical magnification (2.5x). Merged (CX43 and HCN4) immunoreactivity is shown in all three panels. Panel B: Image within the ROI in panel A reconstructed from 4 tile images of the HCN4 + /CX43 - meshwork (red) intertwined with HCN4 - /(CX43) + network (green) taken with 10x water immersion objective. Panels C : Confocal images from the area within the ROI in panel B showing: HCN4 + cells that do not express CX43 (upper image); intertwining areas between HCN4 + /(Cx43) - meshwork (red color), and penetrating HCN4-/ CX43 + cells outlined by green dots corresponding to CX43 protein on the cell membranes (middle and lower panel). Note that HCN4 expressing cells in all three images do not express CX43.

    Journal: bioRxiv

    Article Title: Synchronized cardiac impulses emerge from multi-scale, heterogeneous local calcium signals within and among cells of heart pacemaker tissue

    doi: 10.1101/2020.04.14.039461

    Figure Lengend Snippet: Panel A: A dual immunolabeled HCN4 (red) and CX43 (green) whole mount SAN image at low optical magnification (2.5x). Merged (CX43 and HCN4) immunoreactivity is shown in all three panels. Panel B: Image within the ROI in panel A reconstructed from 4 tile images of the HCN4 + /CX43 - meshwork (red) intertwined with HCN4 - /(CX43) + network (green) taken with 10x water immersion objective. Panels C : Confocal images from the area within the ROI in panel B showing: HCN4 + cells that do not express CX43 (upper image); intertwining areas between HCN4 + /(Cx43) - meshwork (red color), and penetrating HCN4-/ CX43 + cells outlined by green dots corresponding to CX43 protein on the cell membranes (middle and lower panel). Note that HCN4 expressing cells in all three images do not express CX43.

    Article Snippet: Antibodies: HCN4+ cells were identified by rabbit polyclonal antibodies for hyperpolarization-activated, cyclic nucleotide-gate cation channels HCN4 (1:250; Alomone Labs).

    Techniques: Immunolabeling, Expressing

    The HCN4 expressions in the hippocampus of the normal and epileptic animal groups following SE. HCN4 immunoreactivities are rarely detected in all hippocampal regions, whereas its expression is observed in some interneurons (arrows in panels A2-A4). However, the immunoreactivities of HCN4 in the hippocampus following SE are unchanged depending on the time course after pilocarpine treatment (B1-D4). Bar = 280 μm (panels A1, B1, C1, and D1), 50 μm (panels A2-A4, B2-B4, C2-C4, and D2-D4). Quantitative analyses of HCN4 immunoreactivity in the normal and epileptic hippocampi following SE (E, mean ± S.E.M).

    Journal: BMB Reports

    Article Title: Alterations in hyperpolarization-activated cyclic nucleotidegated cation channel (HCN) expression in the hippocampus following pilocarpine-induced status epilepticus

    doi: 10.5483/BMBRep.2012.45.11.091

    Figure Lengend Snippet: The HCN4 expressions in the hippocampus of the normal and epileptic animal groups following SE. HCN4 immunoreactivities are rarely detected in all hippocampal regions, whereas its expression is observed in some interneurons (arrows in panels A2-A4). However, the immunoreactivities of HCN4 in the hippocampus following SE are unchanged depending on the time course after pilocarpine treatment (B1-D4). Bar = 280 μm (panels A1, B1, C1, and D1), 50 μm (panels A2-A4, B2-B4, C2-C4, and D2-D4). Quantitative analyses of HCN4 immunoreactivity in the normal and epileptic hippocampi following SE (E, mean ± S.E.M).

    Article Snippet: Sections were then incubated in below mentioned primary antibodies in PBS containing 0.3% triton X-100 overnight at room temperature; the antibodies employed were rabbit anti-HCN1, HCN2, and HCN4 IgG (Alomone labs, Israel, diluted 1:200).

    Techniques: Expressing

    The p.Pro257Ser mutant channel does not produce a measurable current and does not express on the cell membrane A, Currents were elicited from a holding current of -35 mV to a test pulse of -150 mV (fully-activated voltage) for 4 seconds and returned back to the holding current. The wild type HCN4 channel produced a current and the p.Pro257Ser mutant did not. B, confocal micrographs of wild type HCN4 and p.Pro257Ser channels expressed in CHO cells. The cells were stained with rabbit anti-HCN4 antibody (green) and DAPI (blue). The wild type HCN4 channel is expressed on the cell membrane and in the cytoplasm, whereas the p.Pro257Ser mutant channel is restricted to the cytoplasm. The scale bar denotes 50μm.

    Journal: Heart rhythm : the official journal of the Heart Rhythm Society

    Article Title: A Novel Trafficking-defective HCN4 Mutation is Associated with Early-Onset Atrial Fibrillation

    doi: 10.1016/j.hrthm.2014.03.002

    Figure Lengend Snippet: The p.Pro257Ser mutant channel does not produce a measurable current and does not express on the cell membrane A, Currents were elicited from a holding current of -35 mV to a test pulse of -150 mV (fully-activated voltage) for 4 seconds and returned back to the holding current. The wild type HCN4 channel produced a current and the p.Pro257Ser mutant did not. B, confocal micrographs of wild type HCN4 and p.Pro257Ser channels expressed in CHO cells. The cells were stained with rabbit anti-HCN4 antibody (green) and DAPI (blue). The wild type HCN4 channel is expressed on the cell membrane and in the cytoplasm, whereas the p.Pro257Ser mutant channel is restricted to the cytoplasm. The scale bar denotes 50μm.

    Article Snippet: Thirty-six hours after transfection, the cells were rinsed with PBS, fixed in 4% cold paraformaldehyde, blocked in 10% horse serum and immunolabeled with primary rabbit anti-HCN4 (Alomone labs) and Alexa546-conjugated secondary antibody (Invitrogen).

    Techniques: Mutagenesis, Produced, Staining

    Co-expression of the wild type HCN4 channel the p.Pro257Ser mutant channel produce currents that are not functionally different from wild type A, current recordings of wild type HCN4 (2 μg) and wild type HCN4 (1 μg)+p.Pro257Ser (1 μg). B, plot of current density (pA/pF) measured at -150 mV for wild type HCN4 and wild type HCN4+p.Pro257Ser. C, plots of activation curves for wild type HCN4 and wild type HCN4+p.Pro257Ser. The number in parentheses represents the number of cells. D, confocal micrographs of co-expressed wild type HCN4 and p.Pro257Ser constructs tagged with unique C-terminal epitopes in CHO cells (see methods); i) wild type HCN4-myc (green)+ wild type HCN4-V5 (red) and ii) wild type HCN4-myc (green)+p.Pro257Ser-V5(red). Co-expressed wild type HCN4-myc and wild type HCN4-V5 channels both traffick and are distributed together on cell membrane. ii) wild type HCN4-myc +p.Pro257Ser-V5 images show the wild type HCN4-myc channel expressed on the cell membrane and the p.Pro257Ser-V5 channel distributed in the cytoplasm and not on the cell membrane. Cells were also stained with DAPI (blue) to visualize the nucleus which is shown in the merged images. The scale bar denotes 50μm.

    Journal: Heart rhythm : the official journal of the Heart Rhythm Society

    Article Title: A Novel Trafficking-defective HCN4 Mutation is Associated with Early-Onset Atrial Fibrillation

    doi: 10.1016/j.hrthm.2014.03.002

    Figure Lengend Snippet: Co-expression of the wild type HCN4 channel the p.Pro257Ser mutant channel produce currents that are not functionally different from wild type A, current recordings of wild type HCN4 (2 μg) and wild type HCN4 (1 μg)+p.Pro257Ser (1 μg). B, plot of current density (pA/pF) measured at -150 mV for wild type HCN4 and wild type HCN4+p.Pro257Ser. C, plots of activation curves for wild type HCN4 and wild type HCN4+p.Pro257Ser. The number in parentheses represents the number of cells. D, confocal micrographs of co-expressed wild type HCN4 and p.Pro257Ser constructs tagged with unique C-terminal epitopes in CHO cells (see methods); i) wild type HCN4-myc (green)+ wild type HCN4-V5 (red) and ii) wild type HCN4-myc (green)+p.Pro257Ser-V5(red). Co-expressed wild type HCN4-myc and wild type HCN4-V5 channels both traffick and are distributed together on cell membrane. ii) wild type HCN4-myc +p.Pro257Ser-V5 images show the wild type HCN4-myc channel expressed on the cell membrane and the p.Pro257Ser-V5 channel distributed in the cytoplasm and not on the cell membrane. Cells were also stained with DAPI (blue) to visualize the nucleus which is shown in the merged images. The scale bar denotes 50μm.

    Article Snippet: Thirty-six hours after transfection, the cells were rinsed with PBS, fixed in 4% cold paraformaldehyde, blocked in 10% horse serum and immunolabeled with primary rabbit anti-HCN4 (Alomone labs) and Alexa546-conjugated secondary antibody (Invitrogen).

    Techniques: Expressing, Mutagenesis, Activation Assay, Construct, Staining

    ECG recordings from AF-22 who carries the HCN4 trafficking-defective p.Pro257Ser mutation Surface ECG of leads II and V1 from AF-22. AF-22 presents with the absence of p-waves indicative of AF and conduction pauses at the time of enrollment and remains in AF 12 years later.

    Journal: Heart rhythm : the official journal of the Heart Rhythm Society

    Article Title: A Novel Trafficking-defective HCN4 Mutation is Associated with Early-Onset Atrial Fibrillation

    doi: 10.1016/j.hrthm.2014.03.002

    Figure Lengend Snippet: ECG recordings from AF-22 who carries the HCN4 trafficking-defective p.Pro257Ser mutation Surface ECG of leads II and V1 from AF-22. AF-22 presents with the absence of p-waves indicative of AF and conduction pauses at the time of enrollment and remains in AF 12 years later.

    Article Snippet: Thirty-six hours after transfection, the cells were rinsed with PBS, fixed in 4% cold paraformaldehyde, blocked in 10% horse serum and immunolabeled with primary rabbit anti-HCN4 (Alomone labs) and Alexa546-conjugated secondary antibody (Invitrogen).

    Techniques: Mutagenesis

    Location of novel HCN4 coding variants in early-onset AF cases and referents A B, illustration of a HCN4 -subunit with the cytoplasmic NH2- and COOH-terminus, six transmembrane segments (S1–S6), including the S4 voltage sensor (‘+’ sign denotes amino acid residues with positive charge), the pore loop between S5 and S6, and the C-linker (CL) with the cyclic-nucleotide binding domain (CNBD). The red (seven) and blue (three) circles denote the location of the novel variants identified in the early-onset AF cases and referents, respectively.

    Journal: Heart rhythm : the official journal of the Heart Rhythm Society

    Article Title: A Novel Trafficking-defective HCN4 Mutation is Associated with Early-Onset Atrial Fibrillation

    doi: 10.1016/j.hrthm.2014.03.002

    Figure Lengend Snippet: Location of novel HCN4 coding variants in early-onset AF cases and referents A B, illustration of a HCN4 -subunit with the cytoplasmic NH2- and COOH-terminus, six transmembrane segments (S1–S6), including the S4 voltage sensor (‘+’ sign denotes amino acid residues with positive charge), the pore loop between S5 and S6, and the C-linker (CL) with the cyclic-nucleotide binding domain (CNBD). The red (seven) and blue (three) circles denote the location of the novel variants identified in the early-onset AF cases and referents, respectively.

    Article Snippet: Thirty-six hours after transfection, the cells were rinsed with PBS, fixed in 4% cold paraformaldehyde, blocked in 10% horse serum and immunolabeled with primary rabbit anti-HCN4 (Alomone labs) and Alexa546-conjugated secondary antibody (Invitrogen).

    Techniques: Binding Assay