hcn1  (Alomone Labs)


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    Name:
    Anti HCN1 Antibody
    Description:
    Anti HCN1 Antibody APC 056 is a highly specific antibody directed against an epitope of the rat protein The antibody can be used in western blot immunohistochemistry and immunocytochemistry applications It has been designed to recognize HCN1 from human rat and mouse samples
    Catalog Number:
    APC-056
    Price:
    397.0
    Category:
    Primary Antibody
    Applications:
    Immunocytochemistry, Immunofluorescence, Immunohistochemistry, Western Blot
    Purity:
    Affinity purified on immobilized antigen.
    Immunogen:
    Synthetic peptide
    Size:
    25 mcl
    Antibody Type:
    Polyclonal Primary Antibodies
    Format:
    Lyophilized Powder
    Host:
    Rabbit
    Isotype:
    Rabbit IgG
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    Structured Review

    Alomone Labs hcn1
    Anti HCN1 Antibody
    Anti HCN1 Antibody APC 056 is a highly specific antibody directed against an epitope of the rat protein The antibody can be used in western blot immunohistochemistry and immunocytochemistry applications It has been designed to recognize HCN1 from human rat and mouse samples
    https://www.bioz.com/result/hcn1/product/Alomone Labs
    Average 95 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    hcn1 - by Bioz Stars, 2021-09
    95/100 stars

    Images

    1) Product Images from "Alternatively spliced isoforms of TRIP8b differentially control h channel trafficking and function"

    Article Title: Alternatively spliced isoforms of TRIP8b differentially control h channel trafficking and function

    Journal: The Journal of neuroscience : the official journal of the Society for Neuroscience

    doi: 10.1523/JNEUROSCI.0856-09.2009

    TRIP8b interaction with HCN1 is independent of TRIP8b alternative splicing and is bipartite
    Figure Legend Snippet: TRIP8b interaction with HCN1 is independent of TRIP8b alternative splicing and is bipartite

    Techniques Used:

    TRIP8b isoforms increase or decrease I h peak current in HCN1 cotransfected HEK293T cells depending on isoform identity
    Figure Legend Snippet: TRIP8b isoforms increase or decrease I h peak current in HCN1 cotransfected HEK293T cells depending on isoform identity

    Techniques Used:

    TRIP8b isoforms bidirectionally modify HCN1 protein surface expression as determined by flow cytometry
    Figure Legend Snippet: TRIP8b isoforms bidirectionally modify HCN1 protein surface expression as determined by flow cytometry

    Techniques Used: Expressing, Flow Cytometry

    TRIP8b isoforms A4, B2, and B3 alter amount of surface HCN1 protein
    Figure Legend Snippet: TRIP8b isoforms A4, B2, and B3 alter amount of surface HCN1 protein

    Techniques Used:

    Isoforms of TRIP8b alter the trafficking of HCN1 in cultured hippocampal neurons
    Figure Legend Snippet: Isoforms of TRIP8b alter the trafficking of HCN1 in cultured hippocampal neurons

    Techniques Used: Cell Culture

    2) Product Images from "Alternatively spliced isoforms of TRIP8b differentially control h channel trafficking and function"

    Article Title: Alternatively spliced isoforms of TRIP8b differentially control h channel trafficking and function

    Journal: The Journal of neuroscience : the official journal of the Society for Neuroscience

    doi: 10.1523/JNEUROSCI.0856-09.2009

    TRIP8b interaction with HCN1 is independent of TRIP8b alternative splicing and is bipartite
    Figure Legend Snippet: TRIP8b interaction with HCN1 is independent of TRIP8b alternative splicing and is bipartite

    Techniques Used:

    TRIP8b isoforms increase or decrease I h peak current in HCN1 cotransfected HEK293T cells depending on isoform identity
    Figure Legend Snippet: TRIP8b isoforms increase or decrease I h peak current in HCN1 cotransfected HEK293T cells depending on isoform identity

    Techniques Used:

    TRIP8b isoforms bidirectionally modify HCN1 protein surface expression as determined by flow cytometry
    Figure Legend Snippet: TRIP8b isoforms bidirectionally modify HCN1 protein surface expression as determined by flow cytometry

    Techniques Used: Expressing, Flow Cytometry, Cytometry

    TRIP8b isoforms A4, B2, and B3 alter amount of surface HCN1 protein
    Figure Legend Snippet: TRIP8b isoforms A4, B2, and B3 alter amount of surface HCN1 protein

    Techniques Used:

    Isoforms of TRIP8b alter the trafficking of HCN1 in cultured hippocampal neurons
    Figure Legend Snippet: Isoforms of TRIP8b alter the trafficking of HCN1 in cultured hippocampal neurons

    Techniques Used: Cell Culture

    3) Product Images from "Differential Distribution and Function of Hyperpolarization-Activated Channels in Sensory Neurons and Mechanosensitive Fibers"

    Article Title: Differential Distribution and Function of Hyperpolarization-Activated Channels in Sensory Neurons and Mechanosensitive Fibers

    Journal: The Journal of Neuroscience

    doi: 10.1523/JNEUROSCI.5156-03.2004

    HCN immunoreactivity in club endings of myelinated fibers accompanied by fine unmyelinated fibers. Left, HCN1 (FITC) colocalized (yellow) with peripherin (Rhodamine Red-X) in a club ending but not with fine unmyelinated fiber (arrowhead). Center, HCN2 (Rhodamine Red-X) appeared in both the fine unmyelinated fibers and in a club ending of a myelinated fiber (yellow). Right, Both club endings and fine fibers (arrowhead) labeled with FITC-NF mixture also expressed HCN4 (Rhodamine Red-X). HCN2 and HCN4 (Rhodamine Red-X) are also found in cells surrounding the fibers.
    Figure Legend Snippet: HCN immunoreactivity in club endings of myelinated fibers accompanied by fine unmyelinated fibers. Left, HCN1 (FITC) colocalized (yellow) with peripherin (Rhodamine Red-X) in a club ending but not with fine unmyelinated fiber (arrowhead). Center, HCN2 (Rhodamine Red-X) appeared in both the fine unmyelinated fibers and in a club ending of a myelinated fiber (yellow). Right, Both club endings and fine fibers (arrowhead) labeled with FITC-NF mixture also expressed HCN4 (Rhodamine Red-X). HCN2 and HCN4 (Rhodamine Red-X) are also found in cells surrounding the fibers.

    Techniques Used: Labeling

    HCN1, HCN2, and HCN4 immunoreactivity in nodose neurons. A–C , HCN immunoreactivity identified in 6–10 μm sections of nodose ganglion. HCN1 immunoreactivity was localized to a small subpopulation of neurons and, in most of these cells, was heavily localized at the plasma membrane ( A ). HCN2 ( B ) and HCN4 ( C ) immunoreactivity was present in all neurons in the ganglion. D–F , Single confocal sections through cultured nodose neurons selected for expression of HCN1 ( D ), HCN2 ( E ), and HCN4 ( F ). Heavy labeling at the membrane is again shown for HCN1 ( D ). Patches of HCN2 and HCN4 immunoreactivity were located at the cell perimeter; examples are indicated by the arrows ( E, F ). The light microscopic differential interference contrast image is also shown for each neuron. The calibration bar in C also applies to A and B , whereas calibration in F applies to E . The antibodies were preabsorbed with the immunizing peptide as shown in the figure. A control for nonspecific staining omitted the primary Ab (data not shown).
    Figure Legend Snippet: HCN1, HCN2, and HCN4 immunoreactivity in nodose neurons. A–C , HCN immunoreactivity identified in 6–10 μm sections of nodose ganglion. HCN1 immunoreactivity was localized to a small subpopulation of neurons and, in most of these cells, was heavily localized at the plasma membrane ( A ). HCN2 ( B ) and HCN4 ( C ) immunoreactivity was present in all neurons in the ganglion. D–F , Single confocal sections through cultured nodose neurons selected for expression of HCN1 ( D ), HCN2 ( E ), and HCN4 ( F ). Heavy labeling at the membrane is again shown for HCN1 ( D ). Patches of HCN2 and HCN4 immunoreactivity were located at the cell perimeter; examples are indicated by the arrows ( E, F ). The light microscopic differential interference contrast image is also shown for each neuron. The calibration bar in C also applies to A and B , whereas calibration in F applies to E . The antibodies were preabsorbed with the immunizing peptide as shown in the figure. A control for nonspecific staining omitted the primary Ab (data not shown).

    Techniques Used: Cell Culture, Expressing, Labeling, Staining

    Colocalization of HCN1 and IB4 or VR1. A , No dose ganglion section immunolabeled with rabbit HCN1 Ab (left) and IB4 lectin (right). HCN1 neurons do not contain IB4. Three of the HCN1-labeled neurons are identified by an asterisk. B , Nodose section labeled with anti-HCN1 Ab (left) and anti-VR1 Ab (center) is shown overlaid on the right. The neurons with strong labeling for HCN1 did not coexpress VR1, but weaker HCN1 staining is seen on two VR1(*)-immunoreactive neurons. The arrow indicates an example of HCN1 axonal labeling.
    Figure Legend Snippet: Colocalization of HCN1 and IB4 or VR1. A , No dose ganglion section immunolabeled with rabbit HCN1 Ab (left) and IB4 lectin (right). HCN1 neurons do not contain IB4. Three of the HCN1-labeled neurons are identified by an asterisk. B , Nodose section labeled with anti-HCN1 Ab (left) and anti-VR1 Ab (center) is shown overlaid on the right. The neurons with strong labeling for HCN1 did not coexpress VR1, but weaker HCN1 staining is seen on two VR1(*)-immunoreactive neurons. The arrow indicates an example of HCN1 axonal labeling.

    Techniques Used: Immunolabeling, Labeling, Staining

    HCN1, HCN2, HCN3, and HCN4 mRNA is expressed in nodose ganglia. PCR products resulting from the amplification of first-strand cDNA prepared with (+) or without (–) RT from nodose ganglia or brain poly A+ RNA with HCN1-, HCN2-, HCN3-, and HCN4-specific oligonucleotides were separated by electrophoresis and transferred to nylon membranes (Ambion). After Southern hybridization with 32 P-labeled specific internal oligomers, the autoradiogram showed a positive signal for all four channels from nodose and rat brain in the (+) RT lanes and no signals in the control (–) RT. The oligonucleotide probes amplify cDNA of 641 bp for HCN1, 638 bp for HCN2, 509 bp for HCN3, and 635 bp for HCN4.
    Figure Legend Snippet: HCN1, HCN2, HCN3, and HCN4 mRNA is expressed in nodose ganglia. PCR products resulting from the amplification of first-strand cDNA prepared with (+) or without (–) RT from nodose ganglia or brain poly A+ RNA with HCN1-, HCN2-, HCN3-, and HCN4-specific oligonucleotides were separated by electrophoresis and transferred to nylon membranes (Ambion). After Southern hybridization with 32 P-labeled specific internal oligomers, the autoradiogram showed a positive signal for all four channels from nodose and rat brain in the (+) RT lanes and no signals in the control (–) RT. The oligonucleotide probes amplify cDNA of 641 bp for HCN1, 638 bp for HCN2, 509 bp for HCN3, and 635 bp for HCN4.

    Techniques Used: Polymerase Chain Reaction, Amplification, Electrophoresis, Hybridization, Labeling

    HCN immunoreactivity in aortic baroreceptor terminals of myelinated fibers. Top, A collapsed Z-series stack of 0.4 μm confocal sections through a bush baroreceptor terminal shows localization of HCN1 (left) and PGP9.5 (right). PGP9.5 is a ubiquitin hydrolase expressed in neuronal–neuroendocrine cells. Middle, Bush ending is colabeled with HCN2 on the left and the neurofilament mixture on the right. Bottom, HCN4 immunoreactivity on the left is localized to the bush ending identified using the neurofilament mixture (right).
    Figure Legend Snippet: HCN immunoreactivity in aortic baroreceptor terminals of myelinated fibers. Top, A collapsed Z-series stack of 0.4 μm confocal sections through a bush baroreceptor terminal shows localization of HCN1 (left) and PGP9.5 (right). PGP9.5 is a ubiquitin hydrolase expressed in neuronal–neuroendocrine cells. Middle, Bush ending is colabeled with HCN2 on the left and the neurofilament mixture on the right. Bottom, HCN4 immunoreactivity on the left is localized to the bush ending identified using the neurofilament mixture (right).

    Techniques Used:

    Related Articles

    Immunolabeling:

    Article Title: Presynaptic HCN channels constrain GABAergic synaptic transmission in pyramidal cells of the medial prefrontal cortex
    Article Snippet: .. Sequential primary immunolabeling for HCN1, HCN2 or HCN4 was performed using anti-HCN1, HCN2 or HCN4 rabbit antibodies (1:40; Alomone Laboratories, Israel,. ..

    Protease Inhibitor:

    Article Title: Novel Potassium Channels in Kidney Mitochondria: The Hyperpolarization-Activated and Cyclic Nucleotide-Gated HCN Channels
    Article Snippet: .. Anti-HCN1, anti-HCN2, anti-HCN3 and anti-HCN4 were purchased from Alomone (Jerusalem, Israel). cOmplete™ Protease Inhibitor Cocktail Roche, Mexico. ..

    Incubation:

    Article Title: Functional and Molecular Analysis of Proprioceptive Sensory Neuron Excitability in Mice
    Article Snippet: .. The sections were then incubated in a cocktail of primary antibodies containing guinea pig anti-PV (1:500; Frontier Institute Cat# PV-GP-Af1000, RRID:AB_2336938 ) with either rabbit anti HCN1 (1:250; Alomone Labs Cat# APC-056, RRID:AB_2039900 ), rabbit anti-HCN2 (1:500; Alomone Labs Cat# APC-030, RRID:AB_2313726 ) or mouse anti-HCN4 (diluted 1:500; UC Davis/NIH NeuroMab Facility Cat# 73–150, RRID:AB_10673158). ..

    Article Title: cAMP-dependent regulation of HCN4 controls the tonic entrainment process in sinoatrial node pacemaker cells
    Article Snippet: .. Another three washing steps were followed by overnight incubation at 4 °C with the guinea pig anti-HCN4 and rabbit anti-HCN1 antibodies (1:200 in PBS, Alomone labs, Israel). ..

    Article Title: Hyperpolarization-Activated Cyclic Nucleotide-Gated Ion (HCN) Channels Regulate PC12 Cell Differentiation Toward Sympathetic Neuron
    Article Snippet: .. After washing six times with PBS (5 min for each wash), cells were permeated by incubation in 0.5% Triton X-100 in PBS for 5 min, rinsed six times with PBS, blocked with 10% sheep serum in PBS for 45 min, washed once with 1% sheep serum in PBS, and incubated in a refrigerator (4°C) overnight in the presence or absence of the primary antibodies diluted with 1% sheep serum in PBS: 1:50 dilution for rabbit anti-HCN1 (APC-056), anti-HCN2 (APC-030), anti-HCN3 (APC-057), anti-HCN4 (AGP-004) (Alomone Labs, Jerusalem, Israel), anti-GAP43 (sc-135915) and anti-TH (sc-136100) (Santa Cruz Biotechnology, Inc., United States). ..

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  • 94
    Alomone Labs anti hcn1
    <t>HCN1</t> and HCN2 immunostaining. A and B : ABC immunostaining. A: with X100 objective, HCN1 and HCN2 ring staining (white arrows) is over the neuron perimeter and not in satellite cells (black arrows) thus probably membrane associated, (sn = satellite cell nucleus). B: in adjacent sections HCN1 and HCN2 show co-localisation, with some differences in intensity or location. Symbols in B : ‡ ring staining for both HCN1 and HCN2; x neither, o clear HCN1 ring but weaker HCN2; • clear HCN2 ring but weaker HCN1. C–F: Double immunofluorescence staining in L5 DRG neurons. C and D : HCN1 (C) or HCN2 (D) ring staining (red) are present in large, myelinated, neurofilament-rich (RT97 positive, green) neurons. There is also cytoplasmic HCN2 staining in a few RT97 negative, unmyelinated, small neurons (fine arrow in D). Symbols in C and D indicate examples of staining with: ‡ both antibodies, x neither, o clear HCN2 cytoplasmic but not neurofilament staining. E and F: Representative X100 images of HCN1 (E) and HCN2 (F) (in red) with Ankyrin G (AnkG, green) to indicate the nodes of Ranvier (arrows) in longitudinal sections of normal L5 dorsal root nerve. Yellow is indicative of co-localisation.
    Anti Hcn1, supplied by Alomone Labs, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/anti hcn1/product/Alomone Labs
    Average 94 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    anti hcn1 - by Bioz Stars, 2021-09
    94/100 stars
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    HCN1 and HCN2 immunostaining. A and B : ABC immunostaining. A: with X100 objective, HCN1 and HCN2 ring staining (white arrows) is over the neuron perimeter and not in satellite cells (black arrows) thus probably membrane associated, (sn = satellite cell nucleus). B: in adjacent sections HCN1 and HCN2 show co-localisation, with some differences in intensity or location. Symbols in B : ‡ ring staining for both HCN1 and HCN2; x neither, o clear HCN1 ring but weaker HCN2; • clear HCN2 ring but weaker HCN1. C–F: Double immunofluorescence staining in L5 DRG neurons. C and D : HCN1 (C) or HCN2 (D) ring staining (red) are present in large, myelinated, neurofilament-rich (RT97 positive, green) neurons. There is also cytoplasmic HCN2 staining in a few RT97 negative, unmyelinated, small neurons (fine arrow in D). Symbols in C and D indicate examples of staining with: ‡ both antibodies, x neither, o clear HCN2 cytoplasmic but not neurofilament staining. E and F: Representative X100 images of HCN1 (E) and HCN2 (F) (in red) with Ankyrin G (AnkG, green) to indicate the nodes of Ranvier (arrows) in longitudinal sections of normal L5 dorsal root nerve. Yellow is indicative of co-localisation.

    Journal: PLoS ONE

    Article Title: HCN1 and HCN2 in Rat DRG Neurons: Levels in Nociceptors and Non-Nociceptors, NT3-Dependence and Influence of CFA-Induced Skin Inflammation on HCN2 and NT3 Expression

    doi: 10.1371/journal.pone.0050442

    Figure Lengend Snippet: HCN1 and HCN2 immunostaining. A and B : ABC immunostaining. A: with X100 objective, HCN1 and HCN2 ring staining (white arrows) is over the neuron perimeter and not in satellite cells (black arrows) thus probably membrane associated, (sn = satellite cell nucleus). B: in adjacent sections HCN1 and HCN2 show co-localisation, with some differences in intensity or location. Symbols in B : ‡ ring staining for both HCN1 and HCN2; x neither, o clear HCN1 ring but weaker HCN2; • clear HCN2 ring but weaker HCN1. C–F: Double immunofluorescence staining in L5 DRG neurons. C and D : HCN1 (C) or HCN2 (D) ring staining (red) are present in large, myelinated, neurofilament-rich (RT97 positive, green) neurons. There is also cytoplasmic HCN2 staining in a few RT97 negative, unmyelinated, small neurons (fine arrow in D). Symbols in C and D indicate examples of staining with: ‡ both antibodies, x neither, o clear HCN2 cytoplasmic but not neurofilament staining. E and F: Representative X100 images of HCN1 (E) and HCN2 (F) (in red) with Ankyrin G (AnkG, green) to indicate the nodes of Ranvier (arrows) in longitudinal sections of normal L5 dorsal root nerve. Yellow is indicative of co-localisation.

    Article Snippet: Primary Antibodies The anti-HCN1 and anti-HCN2 antibodies were rabbit polyclonal antibodies from Alomone Labs Ltd, Jerusalem, Israel.

    Techniques: Immunostaining, Staining, Double Immunofluorescence Staining

    HCN1 and HCN2 dependence on NT3. A to D : DRG neurons cultured for 2 days in absence ( A and B ) or presence ( C and D ) of NT3 (40 ng/ml). HCN1 (A and C) and HCN2 (B and D) immunofluorescence examples are shown. Immunostaining for both was much reduced in the absence of NT3. Arrows in A and B show neurons visible under interference contrast; many were so weakly stained they were not/weakly visible under fluorescence. Note that in B , the left hand image includes smaller neurons; the right image includes a medium-sized and a large neuron. In C and D (with NT3), staining was stronger at the cell perimeter of medium to large neurons for both HCN1 and HCN2. Some smaller neurons showed elevated HCN2 staining ( D) . E and F : Quantitation of the effect of NT3 (40 ng/ml) on HCN1 and HCN2 edge staining after 2 days in culture. Medians for HCN1 and HCN2-edge immunostaining were both significantly greater (Mann Whitney test) for medium and large neurons in the presence of NT3. For HCN1 only, there was also a significant elevation in edge staining in small neurons with NT3. * P

    Journal: PLoS ONE

    Article Title: HCN1 and HCN2 in Rat DRG Neurons: Levels in Nociceptors and Non-Nociceptors, NT3-Dependence and Influence of CFA-Induced Skin Inflammation on HCN2 and NT3 Expression

    doi: 10.1371/journal.pone.0050442

    Figure Lengend Snippet: HCN1 and HCN2 dependence on NT3. A to D : DRG neurons cultured for 2 days in absence ( A and B ) or presence ( C and D ) of NT3 (40 ng/ml). HCN1 (A and C) and HCN2 (B and D) immunofluorescence examples are shown. Immunostaining for both was much reduced in the absence of NT3. Arrows in A and B show neurons visible under interference contrast; many were so weakly stained they were not/weakly visible under fluorescence. Note that in B , the left hand image includes smaller neurons; the right image includes a medium-sized and a large neuron. In C and D (with NT3), staining was stronger at the cell perimeter of medium to large neurons for both HCN1 and HCN2. Some smaller neurons showed elevated HCN2 staining ( D) . E and F : Quantitation of the effect of NT3 (40 ng/ml) on HCN1 and HCN2 edge staining after 2 days in culture. Medians for HCN1 and HCN2-edge immunostaining were both significantly greater (Mann Whitney test) for medium and large neurons in the presence of NT3. For HCN1 only, there was also a significant elevation in edge staining in small neurons with NT3. * P

    Article Snippet: Primary Antibodies The anti-HCN1 and anti-HCN2 antibodies were rabbit polyclonal antibodies from Alomone Labs Ltd, Jerusalem, Israel.

    Techniques: Cell Culture, Immunofluorescence, Immunostaining, Staining, Fluorescence, Quantitation Assay, MANN-WHITNEY

    HCN2 immunointensity versus sensory properties and CV. A :Top row dye-injected neurons; bottom row; the same neurons after ABC HCN2 immunocytochemistry; all with X40 objective. Large arrows indicate the dye-injected neurons. From left to right: C-unresponsive neuron shows cytoplasmic but no ring staining; the Aβ-nociceptor illustrated shows no, or a very weak, ring; Aα/β cutaneous LTM (field unit) with clear, not intense, ring; in the same field the asterisk shows a (non-dye-labelled) neuron identified as an MSA by strong staining on an adjacent section (not shown) for the α3 Na + pump, which selectively labels MSAs. Also in that image, fine arrows show satellite cells unstained by HCN2. The right hand dye-injected neuron is a muscle spindle afferent (MSA), with a clear black ring. B : Cytoplasmic HCN2 relative intensities are elevated in MSAs versus other Aα/β neurons and all other neurons. C : HCN2 ring/edge staining is higher in MSAs than other Aα/β-neurons and than all other neurons; it is higher in Aβ-nociceptors than C-nociceptors, which showed no rings. Two out of three C-LTMs showed weak detectable ring staining. D: For non-MSA neurons ring intensities for HCN2 are highly significantly correlated with those for HCN1 (measured in different sections of the same neurons). For MSAs the linear regression was not significant (ns). The best linear regression line for MSAs was highly significantly elevated relative to non MSAs. E : HCN2 ring relative intensity is positively correlated with CV. For more detail and abbreviations see Fig. 2; for statistics see Methods.

    Journal: PLoS ONE

    Article Title: HCN1 and HCN2 in Rat DRG Neurons: Levels in Nociceptors and Non-Nociceptors, NT3-Dependence and Influence of CFA-Induced Skin Inflammation on HCN2 and NT3 Expression

    doi: 10.1371/journal.pone.0050442

    Figure Lengend Snippet: HCN2 immunointensity versus sensory properties and CV. A :Top row dye-injected neurons; bottom row; the same neurons after ABC HCN2 immunocytochemistry; all with X40 objective. Large arrows indicate the dye-injected neurons. From left to right: C-unresponsive neuron shows cytoplasmic but no ring staining; the Aβ-nociceptor illustrated shows no, or a very weak, ring; Aα/β cutaneous LTM (field unit) with clear, not intense, ring; in the same field the asterisk shows a (non-dye-labelled) neuron identified as an MSA by strong staining on an adjacent section (not shown) for the α3 Na + pump, which selectively labels MSAs. Also in that image, fine arrows show satellite cells unstained by HCN2. The right hand dye-injected neuron is a muscle spindle afferent (MSA), with a clear black ring. B : Cytoplasmic HCN2 relative intensities are elevated in MSAs versus other Aα/β neurons and all other neurons. C : HCN2 ring/edge staining is higher in MSAs than other Aα/β-neurons and than all other neurons; it is higher in Aβ-nociceptors than C-nociceptors, which showed no rings. Two out of three C-LTMs showed weak detectable ring staining. D: For non-MSA neurons ring intensities for HCN2 are highly significantly correlated with those for HCN1 (measured in different sections of the same neurons). For MSAs the linear regression was not significant (ns). The best linear regression line for MSAs was highly significantly elevated relative to non MSAs. E : HCN2 ring relative intensity is positively correlated with CV. For more detail and abbreviations see Fig. 2; for statistics see Methods.

    Article Snippet: Primary Antibodies The anti-HCN1 and anti-HCN2 antibodies were rabbit polyclonal antibodies from Alomone Labs Ltd, Jerusalem, Israel.

    Techniques: Injection, Immunocytochemistry, Staining

    HCN1 immunointensity, sensory properties and CV. Images captured with 40X objective. A: From the left, a C-nociceptor, an Aδ-nociceptor, an Aα/β-nociceptor and an Aα/β-LTM SA unit all indicated by arrows. Top: fluorescent dye-injected neurons; bottom: bright field images of the same neurons after HCN1 immunocytochemistry (ABC). B–D: Relative intensities (see Methods) of HCN1 ring and cytoplasmic staining as a percentage of maximum ring intensity are plotted. B and C: scatter plots of HCN1 relative immunostaining intensity for different sensory properties. B: cytoplasmic staining, C: membrane-associated (ring) staining; above 20% (dotted line) ring staining is usually visible. Each symbol represents a single dye-injected neuron. Abbreviations: Noci: nociceptors; LTM: low threshold mechanoreceptor; G/F G hair or field unit; SA: slowly adapting; RA: rapidly adapting; MSA: muscle spindle afferent. Star symbols indicate C− and Aδ-nociceptor-type neurons. Tests were Kruskall-Wallis (solid lines) for comparison of 3 or more groups, or Mann-Whitney tests (for two groups, dotted lines). For more detail and significance levels see Methods. D: HCN1 ring relative intensity versus CV; vertical dotted lines indicate borderlines between C−, Aδ-, and Aα/β-fiber CVs. HCN1 ring intensities were highly correlated with CV in nociceptors (Spearman’s correlation, r s = 0.7, p

    Journal: PLoS ONE

    Article Title: HCN1 and HCN2 in Rat DRG Neurons: Levels in Nociceptors and Non-Nociceptors, NT3-Dependence and Influence of CFA-Induced Skin Inflammation on HCN2 and NT3 Expression

    doi: 10.1371/journal.pone.0050442

    Figure Lengend Snippet: HCN1 immunointensity, sensory properties and CV. Images captured with 40X objective. A: From the left, a C-nociceptor, an Aδ-nociceptor, an Aα/β-nociceptor and an Aα/β-LTM SA unit all indicated by arrows. Top: fluorescent dye-injected neurons; bottom: bright field images of the same neurons after HCN1 immunocytochemistry (ABC). B–D: Relative intensities (see Methods) of HCN1 ring and cytoplasmic staining as a percentage of maximum ring intensity are plotted. B and C: scatter plots of HCN1 relative immunostaining intensity for different sensory properties. B: cytoplasmic staining, C: membrane-associated (ring) staining; above 20% (dotted line) ring staining is usually visible. Each symbol represents a single dye-injected neuron. Abbreviations: Noci: nociceptors; LTM: low threshold mechanoreceptor; G/F G hair or field unit; SA: slowly adapting; RA: rapidly adapting; MSA: muscle spindle afferent. Star symbols indicate C− and Aδ-nociceptor-type neurons. Tests were Kruskall-Wallis (solid lines) for comparison of 3 or more groups, or Mann-Whitney tests (for two groups, dotted lines). For more detail and significance levels see Methods. D: HCN1 ring relative intensity versus CV; vertical dotted lines indicate borderlines between C−, Aδ-, and Aα/β-fiber CVs. HCN1 ring intensities were highly correlated with CV in nociceptors (Spearman’s correlation, r s = 0.7, p

    Article Snippet: Primary Antibodies The anti-HCN1 and anti-HCN2 antibodies were rabbit polyclonal antibodies from Alomone Labs Ltd, Jerusalem, Israel.

    Techniques: Injection, Immunocytochemistry, Staining, Immunostaining, MANN-WHITNEY

    Ring staining for HCN1 and HCN2 versus I h in different neuronal groups. Median values for HCN1 and HCN2 ring intensity for the different neuronal subgroups plotted against published median I h values taken from Table 1 in Gao et al., 2012, for the same neuronal subgroups, thus each symbol represents medians from one subgroup. Note that unlike the immunocytochemistry, the I h was measured in non-dye injected neurons. For ranges of values in each group see Fig. 2 and 3 and Gao et al 2012. Because I h and HCN2 were both significantly higher in MSAs than for any other neurons, we show the linear regressions for all groups excluding MSAs (black lines) and for all groups including MSAs (grey dashed lines).

    Journal: PLoS ONE

    Article Title: HCN1 and HCN2 in Rat DRG Neurons: Levels in Nociceptors and Non-Nociceptors, NT3-Dependence and Influence of CFA-Induced Skin Inflammation on HCN2 and NT3 Expression

    doi: 10.1371/journal.pone.0050442

    Figure Lengend Snippet: Ring staining for HCN1 and HCN2 versus I h in different neuronal groups. Median values for HCN1 and HCN2 ring intensity for the different neuronal subgroups plotted against published median I h values taken from Table 1 in Gao et al., 2012, for the same neuronal subgroups, thus each symbol represents medians from one subgroup. Note that unlike the immunocytochemistry, the I h was measured in non-dye injected neurons. For ranges of values in each group see Fig. 2 and 3 and Gao et al 2012. Because I h and HCN2 were both significantly higher in MSAs than for any other neurons, we show the linear regressions for all groups excluding MSAs (black lines) and for all groups including MSAs (grey dashed lines).

    Article Snippet: Primary Antibodies The anti-HCN1 and anti-HCN2 antibodies were rabbit polyclonal antibodies from Alomone Labs Ltd, Jerusalem, Israel.

    Techniques: Staining, Immunocytochemistry, Injection

    Quantitative polymerase chain reaction (qPCR) ΔCt analysis for hyperpolarization-activated cyclic nucleotide-gated channel (HCN channel) subunits in proprioceptive DRG cells. (A) Box and whisker plot show group data comparing ΔCt for GFP expression in GFP+ (green) and GFP− (blue) samples (normalized to β actin expression). The low GFP ΔCt in GFP+, but not GFP−, confirm high expression in this sample and the reliability of this sampling approach. (B) Box whisker plots plot shows group data comparing ΔCt values for HCN1–4 in GFP+ (green) and GFP− (blue) samples. The expression of all HCN subunits was significantly different between GFP+ and GFP− groups (where * p

    Journal: Frontiers in Molecular Neuroscience

    Article Title: Functional and Molecular Analysis of Proprioceptive Sensory Neuron Excitability in Mice

    doi: 10.3389/fnmol.2020.00036

    Figure Lengend Snippet: Quantitative polymerase chain reaction (qPCR) ΔCt analysis for hyperpolarization-activated cyclic nucleotide-gated channel (HCN channel) subunits in proprioceptive DRG cells. (A) Box and whisker plot show group data comparing ΔCt for GFP expression in GFP+ (green) and GFP− (blue) samples (normalized to β actin expression). The low GFP ΔCt in GFP+, but not GFP−, confirm high expression in this sample and the reliability of this sampling approach. (B) Box whisker plots plot shows group data comparing ΔCt values for HCN1–4 in GFP+ (green) and GFP− (blue) samples. The expression of all HCN subunits was significantly different between GFP+ and GFP− groups (where * p

    Article Snippet: The sections were then incubated in a cocktail of primary antibodies containing guinea pig anti-PV (1:500; Frontier Institute Cat# PV-GP-Af1000, RRID:AB_2336938 ) with either rabbit anti HCN1 (1:250; Alomone Labs Cat# APC-056, RRID:AB_2039900 ), rabbit anti-HCN2 (1:500; Alomone Labs Cat# APC-030, RRID:AB_2313726 ) or mouse anti-HCN4 (diluted 1:500; UC Davis/NIH NeuroMab Facility Cat# 73–150, RRID:AB_10673158).

    Techniques: Real-time Polymerase Chain Reaction, Whisker Assay, Expressing, Sampling

    Immunohistochemical localization of HCN subunits in PV-expressing DRG neurons. (A,B) PV-IR DRG neurons (green) in wild-type mice were shown to express both HCN1 ( A ; red) and HCN2 ( B ; red), with examples denoted by double arrowheads. (C,D) In DRGs from PVeGFP mice, GFP-expressing cells (green) showed immunolabeling for both HCN2 ( C ; red) and HCN4 ( D ; red). Immunolabeling for each of the HCN subunits was restricted to the cell membrane. Scale Bars (μm): (A,B) = 50; (C,D) = 10.

    Journal: Frontiers in Molecular Neuroscience

    Article Title: Functional and Molecular Analysis of Proprioceptive Sensory Neuron Excitability in Mice

    doi: 10.3389/fnmol.2020.00036

    Figure Lengend Snippet: Immunohistochemical localization of HCN subunits in PV-expressing DRG neurons. (A,B) PV-IR DRG neurons (green) in wild-type mice were shown to express both HCN1 ( A ; red) and HCN2 ( B ; red), with examples denoted by double arrowheads. (C,D) In DRGs from PVeGFP mice, GFP-expressing cells (green) showed immunolabeling for both HCN2 ( C ; red) and HCN4 ( D ; red). Immunolabeling for each of the HCN subunits was restricted to the cell membrane. Scale Bars (μm): (A,B) = 50; (C,D) = 10.

    Article Snippet: The sections were then incubated in a cocktail of primary antibodies containing guinea pig anti-PV (1:500; Frontier Institute Cat# PV-GP-Af1000, RRID:AB_2336938 ) with either rabbit anti HCN1 (1:250; Alomone Labs Cat# APC-056, RRID:AB_2039900 ), rabbit anti-HCN2 (1:500; Alomone Labs Cat# APC-030, RRID:AB_2313726 ) or mouse anti-HCN4 (diluted 1:500; UC Davis/NIH NeuroMab Facility Cat# 73–150, RRID:AB_10673158).

    Techniques: Immunohistochemistry, Expressing, Mouse Assay, Immunolabeling

    HCN3 channel expression in mitochondria isolated from renal cortex and HEK293 cells. ( a ) Immunoblotting of HCN1, HCN2, HCN3 and HCN4 with polyclonal antibodies detected only HCN2 (~94 kDa) and HCN3 (~86 kDa) in renal mitochondria. α-tubulin (~50 kDa) and voltage dependent anion channel (VDAC, ~31 kDa) were used as negative and positive controls of mitochondria abundance, respectively. ( b ) Immunoblotting of HCN1, HCN2, HCN3 and HCN4 in supernatant (S) and mitochondria (M) of control HEK293 cells. ( c ) Immunoblotting of HEK293 cells transfected with HCN1, HCN2, HCN3 and HCN4 in supernatant (S) and mitochondria (M). Only HCN3 (86 kDa) and HCN4 (160 kDa) were observed in HEK293 mitochondria. Immunogold electron microscopy localization of HCN3 (arrows) in the inner mitochondrial membrane of mitochondria from ( d ) rat (renal cortex) and ( e ) human kidney. ( f ) Negative control was performed in the absence of the primary antibody.

    Journal: International Journal of Molecular Sciences

    Article Title: Novel Potassium Channels in Kidney Mitochondria: The Hyperpolarization-Activated and Cyclic Nucleotide-Gated HCN Channels

    doi: 10.3390/ijms20204995

    Figure Lengend Snippet: HCN3 channel expression in mitochondria isolated from renal cortex and HEK293 cells. ( a ) Immunoblotting of HCN1, HCN2, HCN3 and HCN4 with polyclonal antibodies detected only HCN2 (~94 kDa) and HCN3 (~86 kDa) in renal mitochondria. α-tubulin (~50 kDa) and voltage dependent anion channel (VDAC, ~31 kDa) were used as negative and positive controls of mitochondria abundance, respectively. ( b ) Immunoblotting of HCN1, HCN2, HCN3 and HCN4 in supernatant (S) and mitochondria (M) of control HEK293 cells. ( c ) Immunoblotting of HEK293 cells transfected with HCN1, HCN2, HCN3 and HCN4 in supernatant (S) and mitochondria (M). Only HCN3 (86 kDa) and HCN4 (160 kDa) were observed in HEK293 mitochondria. Immunogold electron microscopy localization of HCN3 (arrows) in the inner mitochondrial membrane of mitochondria from ( d ) rat (renal cortex) and ( e ) human kidney. ( f ) Negative control was performed in the absence of the primary antibody.

    Article Snippet: Anti-HCN1, anti-HCN2, anti-HCN3 and anti-HCN4 were purchased from Alomone (Jerusalem, Israel). cOmplete™ Protease Inhibitor Cocktail Roche, Mexico.

    Techniques: Expressing, Isolation, Transfection, Electron Microscopy, Negative Control

    Inwardly potassium (K + ) currents recorded in isolated mitochondria from the renal cortex and HEK293 cells correspond to HCN3. ( a ) Fluorescence microscopy imaging from isolated mitochondria obtained from the renal cortex. Mitochondria were incubated for 10 min with 5 µM of the fluorescent indicator MitoTracker Green FM. Excitation was 490 nm and emission collected at 525 nm. By fluorescence microscopy it was possible to separate mitochondria from debris during patch clamp experiments. ( b ) Whole-mitochondria voltage clamp currents were recorded with 20 mV steps from −120 mV to +60 mV and a holding potential of 0 mV. Basal shows a representative family of HCN currents obtained from a single mitochondrion and the same family of currents after 2 min of incubation with 50 µM of the inhibitor ZD7288. ( c ) Currents obtained at −120 mV from different independent mitochondria under basal conditions and after addition of ZD7288. Each filled circle represents a single measurement and the horizontal line indicates the mean value. ( d ) HEK293T cells labeled with the fluorescent indicator MitoTracker Green FM. ( e ) Isolated mitochondria from HEK293 cells labeled with MitoTracker Green FM. ( f ) Family of currents obtained in whole-cell configuration and ( g ) in whole-mitochondria, under basal conditions and from cells overexpressing HCN1, HCN2 and HCN3. ( h ) Currents recorded at −120 mV from mitochondria isolated from HEK293 control cells (Basal) and cells overexpressing HCN1, HCN2, HCN3 and HCN3 + ZD7288. Each filled circle represents a single measurement and the horizontal line indicates the mean value.

    Journal: International Journal of Molecular Sciences

    Article Title: Novel Potassium Channels in Kidney Mitochondria: The Hyperpolarization-Activated and Cyclic Nucleotide-Gated HCN Channels

    doi: 10.3390/ijms20204995

    Figure Lengend Snippet: Inwardly potassium (K + ) currents recorded in isolated mitochondria from the renal cortex and HEK293 cells correspond to HCN3. ( a ) Fluorescence microscopy imaging from isolated mitochondria obtained from the renal cortex. Mitochondria were incubated for 10 min with 5 µM of the fluorescent indicator MitoTracker Green FM. Excitation was 490 nm and emission collected at 525 nm. By fluorescence microscopy it was possible to separate mitochondria from debris during patch clamp experiments. ( b ) Whole-mitochondria voltage clamp currents were recorded with 20 mV steps from −120 mV to +60 mV and a holding potential of 0 mV. Basal shows a representative family of HCN currents obtained from a single mitochondrion and the same family of currents after 2 min of incubation with 50 µM of the inhibitor ZD7288. ( c ) Currents obtained at −120 mV from different independent mitochondria under basal conditions and after addition of ZD7288. Each filled circle represents a single measurement and the horizontal line indicates the mean value. ( d ) HEK293T cells labeled with the fluorescent indicator MitoTracker Green FM. ( e ) Isolated mitochondria from HEK293 cells labeled with MitoTracker Green FM. ( f ) Family of currents obtained in whole-cell configuration and ( g ) in whole-mitochondria, under basal conditions and from cells overexpressing HCN1, HCN2 and HCN3. ( h ) Currents recorded at −120 mV from mitochondria isolated from HEK293 control cells (Basal) and cells overexpressing HCN1, HCN2, HCN3 and HCN3 + ZD7288. Each filled circle represents a single measurement and the horizontal line indicates the mean value.

    Article Snippet: Anti-HCN1, anti-HCN2, anti-HCN3 and anti-HCN4 were purchased from Alomone (Jerusalem, Israel). cOmplete™ Protease Inhibitor Cocktail Roche, Mexico.

    Techniques: Isolation, Fluorescence, Microscopy, Imaging, Incubation, Patch Clamp, Labeling

    TRIP8b interaction with HCN1 is independent of TRIP8b alternative splicing and is bipartite

    Journal: The Journal of neuroscience : the official journal of the Society for Neuroscience

    Article Title: Alternatively spliced isoforms of TRIP8b differentially control h channel trafficking and function

    doi: 10.1523/JNEUROSCI.0856-09.2009

    Figure Lengend Snippet: TRIP8b interaction with HCN1 is independent of TRIP8b alternative splicing and is bipartite

    Article Snippet: The following primary antibodies were used: mouse (ms) monoclonal antibodies to α-tubulin (DM1A, Upstate Biotechnology, Lake Placid, NY); rabbit (rab) polyclonal antibodies to HCN1 (Alomone Labs, Jerusalem); ms monoclonal to HA epitope (F-7, Santa Cruz Biotechnology, Santa Cruz, CA); Guinea pig (gp) polyclonal antibodies to HCN1 ( ); gp polyclonal antibodies to HCN2 ( ); rab and gp polyclonal antibodies to green fluorescent protein (see antibody generation and , available at as ); rab polyclonal antibodies to TRIP8b ( ); gp polyclonal antibodies to TRIP8b (see antibody generation and , available at as ); gp polyclonal antibodies to HCN4 ( ); gp polyclonal antibodies to TRIP8b exon 1a-5, exon 2, and exon 4 (see antibody generation and ); ms monoclonal to MAP-2 (Sigma, St. Louis, MO).

    Techniques:

    TRIP8b isoforms increase or decrease I h peak current in HCN1 cotransfected HEK293T cells depending on isoform identity

    Journal: The Journal of neuroscience : the official journal of the Society for Neuroscience

    Article Title: Alternatively spliced isoforms of TRIP8b differentially control h channel trafficking and function

    doi: 10.1523/JNEUROSCI.0856-09.2009

    Figure Lengend Snippet: TRIP8b isoforms increase or decrease I h peak current in HCN1 cotransfected HEK293T cells depending on isoform identity

    Article Snippet: The following primary antibodies were used: mouse (ms) monoclonal antibodies to α-tubulin (DM1A, Upstate Biotechnology, Lake Placid, NY); rabbit (rab) polyclonal antibodies to HCN1 (Alomone Labs, Jerusalem); ms monoclonal to HA epitope (F-7, Santa Cruz Biotechnology, Santa Cruz, CA); Guinea pig (gp) polyclonal antibodies to HCN1 ( ); gp polyclonal antibodies to HCN2 ( ); rab and gp polyclonal antibodies to green fluorescent protein (see antibody generation and , available at as ); rab polyclonal antibodies to TRIP8b ( ); gp polyclonal antibodies to TRIP8b (see antibody generation and , available at as ); gp polyclonal antibodies to HCN4 ( ); gp polyclonal antibodies to TRIP8b exon 1a-5, exon 2, and exon 4 (see antibody generation and ); ms monoclonal to MAP-2 (Sigma, St. Louis, MO).

    Techniques:

    TRIP8b isoforms bidirectionally modify HCN1 protein surface expression as determined by flow cytometry

    Journal: The Journal of neuroscience : the official journal of the Society for Neuroscience

    Article Title: Alternatively spliced isoforms of TRIP8b differentially control h channel trafficking and function

    doi: 10.1523/JNEUROSCI.0856-09.2009

    Figure Lengend Snippet: TRIP8b isoforms bidirectionally modify HCN1 protein surface expression as determined by flow cytometry

    Article Snippet: The following primary antibodies were used: mouse (ms) monoclonal antibodies to α-tubulin (DM1A, Upstate Biotechnology, Lake Placid, NY); rabbit (rab) polyclonal antibodies to HCN1 (Alomone Labs, Jerusalem); ms monoclonal to HA epitope (F-7, Santa Cruz Biotechnology, Santa Cruz, CA); Guinea pig (gp) polyclonal antibodies to HCN1 ( ); gp polyclonal antibodies to HCN2 ( ); rab and gp polyclonal antibodies to green fluorescent protein (see antibody generation and , available at as ); rab polyclonal antibodies to TRIP8b ( ); gp polyclonal antibodies to TRIP8b (see antibody generation and , available at as ); gp polyclonal antibodies to HCN4 ( ); gp polyclonal antibodies to TRIP8b exon 1a-5, exon 2, and exon 4 (see antibody generation and ); ms monoclonal to MAP-2 (Sigma, St. Louis, MO).

    Techniques: Expressing, Flow Cytometry

    TRIP8b isoforms A4, B2, and B3 alter amount of surface HCN1 protein

    Journal: The Journal of neuroscience : the official journal of the Society for Neuroscience

    Article Title: Alternatively spliced isoforms of TRIP8b differentially control h channel trafficking and function

    doi: 10.1523/JNEUROSCI.0856-09.2009

    Figure Lengend Snippet: TRIP8b isoforms A4, B2, and B3 alter amount of surface HCN1 protein

    Article Snippet: The following primary antibodies were used: mouse (ms) monoclonal antibodies to α-tubulin (DM1A, Upstate Biotechnology, Lake Placid, NY); rabbit (rab) polyclonal antibodies to HCN1 (Alomone Labs, Jerusalem); ms monoclonal to HA epitope (F-7, Santa Cruz Biotechnology, Santa Cruz, CA); Guinea pig (gp) polyclonal antibodies to HCN1 ( ); gp polyclonal antibodies to HCN2 ( ); rab and gp polyclonal antibodies to green fluorescent protein (see antibody generation and , available at as ); rab polyclonal antibodies to TRIP8b ( ); gp polyclonal antibodies to TRIP8b (see antibody generation and , available at as ); gp polyclonal antibodies to HCN4 ( ); gp polyclonal antibodies to TRIP8b exon 1a-5, exon 2, and exon 4 (see antibody generation and ); ms monoclonal to MAP-2 (Sigma, St. Louis, MO).

    Techniques:

    Isoforms of TRIP8b alter the trafficking of HCN1 in cultured hippocampal neurons

    Journal: The Journal of neuroscience : the official journal of the Society for Neuroscience

    Article Title: Alternatively spliced isoforms of TRIP8b differentially control h channel trafficking and function

    doi: 10.1523/JNEUROSCI.0856-09.2009

    Figure Lengend Snippet: Isoforms of TRIP8b alter the trafficking of HCN1 in cultured hippocampal neurons

    Article Snippet: The following primary antibodies were used: mouse (ms) monoclonal antibodies to α-tubulin (DM1A, Upstate Biotechnology, Lake Placid, NY); rabbit (rab) polyclonal antibodies to HCN1 (Alomone Labs, Jerusalem); ms monoclonal to HA epitope (F-7, Santa Cruz Biotechnology, Santa Cruz, CA); Guinea pig (gp) polyclonal antibodies to HCN1 ( ); gp polyclonal antibodies to HCN2 ( ); rab and gp polyclonal antibodies to green fluorescent protein (see antibody generation and , available at as ); rab polyclonal antibodies to TRIP8b ( ); gp polyclonal antibodies to TRIP8b (see antibody generation and , available at as ); gp polyclonal antibodies to HCN4 ( ); gp polyclonal antibodies to TRIP8b exon 1a-5, exon 2, and exon 4 (see antibody generation and ); ms monoclonal to MAP-2 (Sigma, St. Louis, MO).

    Techniques: Cell Culture