rabbit anti kv7 1  (Alomone Labs)


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    Structured Review

    Alomone Labs rabbit anti kv7 1
    Mutation of phosphorylated serines does not affect subcellular localization of <t>Kv7.1.</t> Alanine ( A ) and aspartate ( B ) mutants of the identified serine phosphorylation sites were coexpressed with the ER marker pDsRed2-ER in MDCK cells. The cells were stained
    Rabbit Anti Kv7 1, supplied by Alomone Labs, used in various techniques. Bioz Stars score: 94/100, based on 3 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/rabbit anti kv7 1/product/Alomone Labs
    Average 94 stars, based on 3 article reviews
    Price from $9.99 to $1999.99
    rabbit anti kv7 1 - by Bioz Stars, 2022-10
    94/100 stars

    Images

    1) Product Images from "Protein kinase A stimulates Kv7.1 surface expression by regulating Nedd4-2-dependent endocytic trafficking"

    Article Title: Protein kinase A stimulates Kv7.1 surface expression by regulating Nedd4-2-dependent endocytic trafficking

    Journal: American Journal of Physiology - Cell Physiology

    doi: 10.1152/ajpcell.00383.2014

    Mutation of phosphorylated serines does not affect subcellular localization of Kv7.1. Alanine ( A ) and aspartate ( B ) mutants of the identified serine phosphorylation sites were coexpressed with the ER marker pDsRed2-ER in MDCK cells. The cells were stained
    Figure Legend Snippet: Mutation of phosphorylated serines does not affect subcellular localization of Kv7.1. Alanine ( A ) and aspartate ( B ) mutants of the identified serine phosphorylation sites were coexpressed with the ER marker pDsRed2-ER in MDCK cells. The cells were stained

    Techniques Used: Mutagenesis, Marker, Staining

    Summary of proteomics results for Kv7.1. A : topological representation of the Kv7.1 channel, including the amino acid sequence. The amino acid sequences highlighted in blue were covered in the mass spectrometry experiments, and the identified phosphorylation
    Figure Legend Snippet: Summary of proteomics results for Kv7.1. A : topological representation of the Kv7.1 channel, including the amino acid sequence. The amino acid sequences highlighted in blue were covered in the mass spectrometry experiments, and the identified phosphorylation

    Techniques Used: Sequencing, Mass Spectrometry

    Inhibition of PKA leads to internalization and possibly lysosomal degradation of Kv7.1.
    Figure Legend Snippet: Inhibition of PKA leads to internalization and possibly lysosomal degradation of Kv7.1.

    Techniques Used: Inhibition

    Kv7.1 accumulates intracellularly upon exposure to PKA inhibitors KT-5720 and myristoylated protein kinase A inhibitor 14–22 amide. A : MDCK cells stably expressing Kv7.1 were subjected to a calcium switch for 24 h (control) followed by treatment
    Figure Legend Snippet: Kv7.1 accumulates intracellularly upon exposure to PKA inhibitors KT-5720 and myristoylated protein kinase A inhibitor 14–22 amide. A : MDCK cells stably expressing Kv7.1 were subjected to a calcium switch for 24 h (control) followed by treatment

    Techniques Used: Stable Transfection, Expressing

    Kv7.1 codistributes with late endosomal/lysosomal markers in response to H-89 treatment. A : confocal scans of MDCK-Kv7.1 cells treated with 20 μM H-89 for 3 h and colabeled with the late endosomal/lysosomal marker LAMP-2. Scale bar = 10 ( top )
    Figure Legend Snippet: Kv7.1 codistributes with late endosomal/lysosomal markers in response to H-89 treatment. A : confocal scans of MDCK-Kv7.1 cells treated with 20 μM H-89 for 3 h and colabeled with the late endosomal/lysosomal marker LAMP-2. Scale bar = 10 ( top )

    Techniques Used: Marker

    Small-interfering RNA (siRNA) knockdown of Nedd4-2 protects Kv7.1 from internalization upon PKA inhibition. A : MDCK cells stably expressing Kv7.1 were transiently transfected with siRNA targeting Nedd4-2 or with noncoding control siRNA. Enhanced GFP (eGFP)
    Figure Legend Snippet: Small-interfering RNA (siRNA) knockdown of Nedd4-2 protects Kv7.1 from internalization upon PKA inhibition. A : MDCK cells stably expressing Kv7.1 were transiently transfected with siRNA targeting Nedd4-2 or with noncoding control siRNA. Enhanced GFP (eGFP)

    Techniques Used: Small Interfering RNA, Inhibition, Stable Transfection, Expressing, Transfection

    Kv7.1-YA localization is unaffected by PKA inhibition. MDCK cells stably expressing the Kv7.1-YA mutant were treated with 20 μM H-89 for 3–5 h. A : confocal scans of MDCK-Kv7.1-YA cells before (0 h) and 3 (3 h H-89) and 5 (5 h H-89) h after
    Figure Legend Snippet: Kv7.1-YA localization is unaffected by PKA inhibition. MDCK cells stably expressing the Kv7.1-YA mutant were treated with 20 μM H-89 for 3–5 h. A : confocal scans of MDCK-Kv7.1-YA cells before (0 h) and 3 (3 h H-89) and 5 (5 h H-89) h after

    Techniques Used: Inhibition, Stable Transfection, Expressing, Mutagenesis

    Kv7.1 redistribution is not caused by increased Nedd4-2 binding or ubiquitinylation. A : representative Western blot showing the effects of 20 μM H-89 (3 h exposure) and 50 μM forskolin (1 h exposure) on the abundance of S 468 -phosphorylated
    Figure Legend Snippet: Kv7.1 redistribution is not caused by increased Nedd4-2 binding or ubiquitinylation. A : representative Western blot showing the effects of 20 μM H-89 (3 h exposure) and 50 μM forskolin (1 h exposure) on the abundance of S 468 -phosphorylated

    Techniques Used: Binding Assay, Western Blot

    Kv7.1 accumulates intracellularly upon inhibition of protein kinase A (PKA). Madin-Darby canine kidney (MDCK) cells stably expressing Kv7.1 were subjected to a calcium switch for 24 h followed by treatment with 20 μM H-89 for 3–5 h. A
    Figure Legend Snippet: Kv7.1 accumulates intracellularly upon inhibition of protein kinase A (PKA). Madin-Darby canine kidney (MDCK) cells stably expressing Kv7.1 were subjected to a calcium switch for 24 h followed by treatment with 20 μM H-89 for 3–5 h. A

    Techniques Used: Inhibition, Stable Transfection, Expressing

    Intracellular accumulation in response to PKA inhibition is specific for Kv7.1. MDCK cells were transfected with Kv7.4 and allowed to polarize before being subjected to either 20 μM H-89 or 10 μM KT-5720 treatments for 3 h. As demonstrated,
    Figure Legend Snippet: Intracellular accumulation in response to PKA inhibition is specific for Kv7.1. MDCK cells were transfected with Kv7.4 and allowed to polarize before being subjected to either 20 μM H-89 or 10 μM KT-5720 treatments for 3 h. As demonstrated,

    Techniques Used: Inhibition, Transfection

    PKA activation promotes Kv7.1 surface expression in polarized MDCK cells. MDCK cells stably expressing Kv7.1 were grown to confluence and allowed to polarize. The cells were then treated with 50 μM forskolin for 1 h. Cells were fixed before and
    Figure Legend Snippet: PKA activation promotes Kv7.1 surface expression in polarized MDCK cells. MDCK cells stably expressing Kv7.1 were grown to confluence and allowed to polarize. The cells were then treated with 50 μM forskolin for 1 h. Cells were fixed before and

    Techniques Used: Activation Assay, Expressing, Stable Transfection

    Mimicked phosphorylation of T470 results in reduced membrane expression of Kv7.1, but Kv7.1-T470A is still endocytosed in response to PKA inhibition. Kv7.1-T470A and Kv7.1-T470D mutants were coexpressed with DsRed2-ER in MDCK cells, which were allowed
    Figure Legend Snippet: Mimicked phosphorylation of T470 results in reduced membrane expression of Kv7.1, but Kv7.1-T470A is still endocytosed in response to PKA inhibition. Kv7.1-T470A and Kv7.1-T470D mutants were coexpressed with DsRed2-ER in MDCK cells, which were allowed

    Techniques Used: Expressing, Inhibition

    2) Product Images from "Protein kinase A stimulates Kv7.1 surface expression by regulating Nedd4-2-dependent endocytic trafficking"

    Article Title: Protein kinase A stimulates Kv7.1 surface expression by regulating Nedd4-2-dependent endocytic trafficking

    Journal: American Journal of Physiology - Cell Physiology

    doi: 10.1152/ajpcell.00383.2014

    Mutation of phosphorylated serines does not affect subcellular localization of Kv7.1. Alanine ( A ) and aspartate ( B ) mutants of the identified serine phosphorylation sites were coexpressed with the ER marker pDsRed2-ER in MDCK cells. The cells were stained
    Figure Legend Snippet: Mutation of phosphorylated serines does not affect subcellular localization of Kv7.1. Alanine ( A ) and aspartate ( B ) mutants of the identified serine phosphorylation sites were coexpressed with the ER marker pDsRed2-ER in MDCK cells. The cells were stained

    Techniques Used: Mutagenesis, Marker, Staining

    Summary of proteomics results for Kv7.1. A : topological representation of the Kv7.1 channel, including the amino acid sequence. The amino acid sequences highlighted in blue were covered in the mass spectrometry experiments, and the identified phosphorylation
    Figure Legend Snippet: Summary of proteomics results for Kv7.1. A : topological representation of the Kv7.1 channel, including the amino acid sequence. The amino acid sequences highlighted in blue were covered in the mass spectrometry experiments, and the identified phosphorylation

    Techniques Used: Sequencing, Mass Spectrometry

    Inhibition of PKA leads to internalization and possibly lysosomal degradation of Kv7.1.
    Figure Legend Snippet: Inhibition of PKA leads to internalization and possibly lysosomal degradation of Kv7.1.

    Techniques Used: Inhibition

    Kv7.1 accumulates intracellularly upon exposure to PKA inhibitors KT-5720 and myristoylated protein kinase A inhibitor 14–22 amide. A : MDCK cells stably expressing Kv7.1 were subjected to a calcium switch for 24 h (control) followed by treatment
    Figure Legend Snippet: Kv7.1 accumulates intracellularly upon exposure to PKA inhibitors KT-5720 and myristoylated protein kinase A inhibitor 14–22 amide. A : MDCK cells stably expressing Kv7.1 were subjected to a calcium switch for 24 h (control) followed by treatment

    Techniques Used: Stable Transfection, Expressing

    Kv7.1 codistributes with late endosomal/lysosomal markers in response to H-89 treatment. A : confocal scans of MDCK-Kv7.1 cells treated with 20 μM H-89 for 3 h and colabeled with the late endosomal/lysosomal marker LAMP-2. Scale bar = 10 ( top )
    Figure Legend Snippet: Kv7.1 codistributes with late endosomal/lysosomal markers in response to H-89 treatment. A : confocal scans of MDCK-Kv7.1 cells treated with 20 μM H-89 for 3 h and colabeled with the late endosomal/lysosomal marker LAMP-2. Scale bar = 10 ( top )

    Techniques Used: Marker

    Small-interfering RNA (siRNA) knockdown of Nedd4-2 protects Kv7.1 from internalization upon PKA inhibition. A : MDCK cells stably expressing Kv7.1 were transiently transfected with siRNA targeting Nedd4-2 or with noncoding control siRNA. Enhanced GFP (eGFP)
    Figure Legend Snippet: Small-interfering RNA (siRNA) knockdown of Nedd4-2 protects Kv7.1 from internalization upon PKA inhibition. A : MDCK cells stably expressing Kv7.1 were transiently transfected with siRNA targeting Nedd4-2 or with noncoding control siRNA. Enhanced GFP (eGFP)

    Techniques Used: Small Interfering RNA, Inhibition, Stable Transfection, Expressing, Transfection

    Kv7.1-YA localization is unaffected by PKA inhibition. MDCK cells stably expressing the Kv7.1-YA mutant were treated with 20 μM H-89 for 3–5 h. A : confocal scans of MDCK-Kv7.1-YA cells before (0 h) and 3 (3 h H-89) and 5 (5 h H-89) h after
    Figure Legend Snippet: Kv7.1-YA localization is unaffected by PKA inhibition. MDCK cells stably expressing the Kv7.1-YA mutant were treated with 20 μM H-89 for 3–5 h. A : confocal scans of MDCK-Kv7.1-YA cells before (0 h) and 3 (3 h H-89) and 5 (5 h H-89) h after

    Techniques Used: Inhibition, Stable Transfection, Expressing, Mutagenesis

    Kv7.1 redistribution is not caused by increased Nedd4-2 binding or ubiquitinylation. A : representative Western blot showing the effects of 20 μM H-89 (3 h exposure) and 50 μM forskolin (1 h exposure) on the abundance of S 468 -phosphorylated
    Figure Legend Snippet: Kv7.1 redistribution is not caused by increased Nedd4-2 binding or ubiquitinylation. A : representative Western blot showing the effects of 20 μM H-89 (3 h exposure) and 50 μM forskolin (1 h exposure) on the abundance of S 468 -phosphorylated

    Techniques Used: Binding Assay, Western Blot

    Kv7.1 accumulates intracellularly upon inhibition of protein kinase A (PKA). Madin-Darby canine kidney (MDCK) cells stably expressing Kv7.1 were subjected to a calcium switch for 24 h followed by treatment with 20 μM H-89 for 3–5 h. A
    Figure Legend Snippet: Kv7.1 accumulates intracellularly upon inhibition of protein kinase A (PKA). Madin-Darby canine kidney (MDCK) cells stably expressing Kv7.1 were subjected to a calcium switch for 24 h followed by treatment with 20 μM H-89 for 3–5 h. A

    Techniques Used: Inhibition, Stable Transfection, Expressing

    Intracellular accumulation in response to PKA inhibition is specific for Kv7.1. MDCK cells were transfected with Kv7.4 and allowed to polarize before being subjected to either 20 μM H-89 or 10 μM KT-5720 treatments for 3 h. As demonstrated,
    Figure Legend Snippet: Intracellular accumulation in response to PKA inhibition is specific for Kv7.1. MDCK cells were transfected with Kv7.4 and allowed to polarize before being subjected to either 20 μM H-89 or 10 μM KT-5720 treatments for 3 h. As demonstrated,

    Techniques Used: Inhibition, Transfection

    PKA activation promotes Kv7.1 surface expression in polarized MDCK cells. MDCK cells stably expressing Kv7.1 were grown to confluence and allowed to polarize. The cells were then treated with 50 μM forskolin for 1 h. Cells were fixed before and
    Figure Legend Snippet: PKA activation promotes Kv7.1 surface expression in polarized MDCK cells. MDCK cells stably expressing Kv7.1 were grown to confluence and allowed to polarize. The cells were then treated with 50 μM forskolin for 1 h. Cells were fixed before and

    Techniques Used: Activation Assay, Expressing, Stable Transfection

    Mimicked phosphorylation of T470 results in reduced membrane expression of Kv7.1, but Kv7.1-T470A is still endocytosed in response to PKA inhibition. Kv7.1-T470A and Kv7.1-T470D mutants were coexpressed with DsRed2-ER in MDCK cells, which were allowed
    Figure Legend Snippet: Mimicked phosphorylation of T470 results in reduced membrane expression of Kv7.1, but Kv7.1-T470A is still endocytosed in response to PKA inhibition. Kv7.1-T470A and Kv7.1-T470D mutants were coexpressed with DsRed2-ER in MDCK cells, which were allowed

    Techniques Used: Expressing, Inhibition

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    Alomone Labs rabbit anti kv7 1 antibody
    Mutation of phosphorylated serines does not affect subcellular localization of <t>Kv7.1.</t> Alanine ( A ) and aspartate ( B ) mutants of the identified serine phosphorylation sites were coexpressed with the ER marker pDsRed2-ER in MDCK cells. The cells were stained
    Rabbit Anti Kv7 1 Antibody, supplied by Alomone Labs, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/rabbit anti kv7 1 antibody/product/Alomone Labs
    Average 94 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    rabbit anti kv7 1 antibody - by Bioz Stars, 2022-10
    94/100 stars
      Buy from Supplier

    94
    Alomone Labs rabbit anti kv7 4
    Dopamine activates Kv7/M current in NAc-projecting VTA DA neurons. (A) Kv7/M currents recorded from DA neurons of WT (i,ii) and <t>Kv7.4</t> −/− (iii,iv) mice. Shown are time-courses of M current amplitudes recorded using whole-cell patch-clamp from VTA slices (for details, see “Materials and Methods” section). Summarized data are presented in (B) . Error bars represent SEM. *** p
    Rabbit Anti Kv7 4, supplied by Alomone Labs, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/rabbit anti kv7 4/product/Alomone Labs
    Average 94 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    rabbit anti kv7 4 - by Bioz Stars, 2022-10
    94/100 stars
      Buy from Supplier

    93
    Alomone Labs rabbit anti kv7 2
    NP1 interacts and colocalizes with <t>Kv7.2</t> at presynaptic terminals of excitatory synapses and axonal growth cones. A – D , Representative Western blots of immunoprecipitation eluates separated in 6% Tris-Glycine (for high molecular weight proteins) and 4–12% Bis-Tris gels (for low molecular weight proteins). TL, Total protein lysate; Syx, syntaxin. A , Both native Kv7.2 and syntaxin 1A, but not Kv7.3, coprecipitate with NP1 in total brain extracts. B , Both native Kv7.2 and NP1 coprecipitate with syntaxin in total brain extracts. C , D , Recombinant NP1 coprecipitates Kv7.2, but not syntaxin or Kv7.3, in 293T cells transfected with NP1, 5Myc-Kv7.2, 2HA-Kv7.3, and syntaxin 1A cDNAs. Kv7.2 and Kv7.3 were immunoprecipitated with antibodies against their respective Myc and HA tags. E , F , Immunofluorescence studies and confocal microscopy were performed in high-density ( E ) or low-density ( F ) isolated cortical neurons. E , Top, Confocal sections of 0.772 μm in the z -plane showing immunofluorescence of NP1, VGLUT1, Kv7.2, and negative control (omitting primary antibodies). Bottom, Colocalization (in white) of the excitatory presynaptic marker VGLUT1 (blue) with NP1 (green) and Kv7.2 (red) is shown in a single section with the corresponding orthogonal views of the stack of confocal sections. White arrows indicate sites of colocalization. F , NP1 (green) and KV7.2 (magenta) immunofluorescence and DIC images of an isolated cortical cultured neuron (1×) with its corresponding axonal growth cone highlighted in a white square box, shown in a confocal section of 0.772 μm in the z -plane at higher (5×) magnification. The negative control for primary antibodies is shown in another growth cone on the right. The image in the bottom is the merge of NP1 and Kv7.2 immunofluorescence images in the single confocal section obtained at 5× showing colocalization (white) of NP1 and Kv7.2 in the growth cone, with the corresponding orthogonal views from its respective stack of confocal sections. Images were acquired using restricted spectral emission wavelength ranges chosen to avoid crosstalk or bleed-through between the three different channels. Scale bar, 5 μm.
    Rabbit Anti Kv7 2, supplied by Alomone Labs, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/rabbit anti kv7 2/product/Alomone Labs
    Average 93 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    rabbit anti kv7 2 - by Bioz Stars, 2022-10
    93/100 stars
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    Image Search Results


    Mutation of phosphorylated serines does not affect subcellular localization of Kv7.1. Alanine ( A ) and aspartate ( B ) mutants of the identified serine phosphorylation sites were coexpressed with the ER marker pDsRed2-ER in MDCK cells. The cells were stained

    Journal: American Journal of Physiology - Cell Physiology

    Article Title: Protein kinase A stimulates Kv7.1 surface expression by regulating Nedd4-2-dependent endocytic trafficking

    doi: 10.1152/ajpcell.00383.2014

    Figure Lengend Snippet: Mutation of phosphorylated serines does not affect subcellular localization of Kv7.1. Alanine ( A ) and aspartate ( B ) mutants of the identified serine phosphorylation sites were coexpressed with the ER marker pDsRed2-ER in MDCK cells. The cells were stained

    Article Snippet: The precleared lysate was subsequently incubated with 3 μg of rabbit anti-Kv7.1 antibody (Alomone Labs), or, as a control, 3 μg normal rabbit IgG (Santa Cruz Biotechnology) for 1 h at 4°C.

    Techniques: Mutagenesis, Marker, Staining

    Summary of proteomics results for Kv7.1. A : topological representation of the Kv7.1 channel, including the amino acid sequence. The amino acid sequences highlighted in blue were covered in the mass spectrometry experiments, and the identified phosphorylation

    Journal: American Journal of Physiology - Cell Physiology

    Article Title: Protein kinase A stimulates Kv7.1 surface expression by regulating Nedd4-2-dependent endocytic trafficking

    doi: 10.1152/ajpcell.00383.2014

    Figure Lengend Snippet: Summary of proteomics results for Kv7.1. A : topological representation of the Kv7.1 channel, including the amino acid sequence. The amino acid sequences highlighted in blue were covered in the mass spectrometry experiments, and the identified phosphorylation

    Article Snippet: The precleared lysate was subsequently incubated with 3 μg of rabbit anti-Kv7.1 antibody (Alomone Labs), or, as a control, 3 μg normal rabbit IgG (Santa Cruz Biotechnology) for 1 h at 4°C.

    Techniques: Sequencing, Mass Spectrometry

    Inhibition of PKA leads to internalization and possibly lysosomal degradation of Kv7.1.

    Journal: American Journal of Physiology - Cell Physiology

    Article Title: Protein kinase A stimulates Kv7.1 surface expression by regulating Nedd4-2-dependent endocytic trafficking

    doi: 10.1152/ajpcell.00383.2014

    Figure Lengend Snippet: Inhibition of PKA leads to internalization and possibly lysosomal degradation of Kv7.1.

    Article Snippet: The precleared lysate was subsequently incubated with 3 μg of rabbit anti-Kv7.1 antibody (Alomone Labs), or, as a control, 3 μg normal rabbit IgG (Santa Cruz Biotechnology) for 1 h at 4°C.

    Techniques: Inhibition

    Kv7.1 accumulates intracellularly upon exposure to PKA inhibitors KT-5720 and myristoylated protein kinase A inhibitor 14–22 amide. A : MDCK cells stably expressing Kv7.1 were subjected to a calcium switch for 24 h (control) followed by treatment

    Journal: American Journal of Physiology - Cell Physiology

    Article Title: Protein kinase A stimulates Kv7.1 surface expression by regulating Nedd4-2-dependent endocytic trafficking

    doi: 10.1152/ajpcell.00383.2014

    Figure Lengend Snippet: Kv7.1 accumulates intracellularly upon exposure to PKA inhibitors KT-5720 and myristoylated protein kinase A inhibitor 14–22 amide. A : MDCK cells stably expressing Kv7.1 were subjected to a calcium switch for 24 h (control) followed by treatment

    Article Snippet: The precleared lysate was subsequently incubated with 3 μg of rabbit anti-Kv7.1 antibody (Alomone Labs), or, as a control, 3 μg normal rabbit IgG (Santa Cruz Biotechnology) for 1 h at 4°C.

    Techniques: Stable Transfection, Expressing

    Kv7.1 codistributes with late endosomal/lysosomal markers in response to H-89 treatment. A : confocal scans of MDCK-Kv7.1 cells treated with 20 μM H-89 for 3 h and colabeled with the late endosomal/lysosomal marker LAMP-2. Scale bar = 10 ( top )

    Journal: American Journal of Physiology - Cell Physiology

    Article Title: Protein kinase A stimulates Kv7.1 surface expression by regulating Nedd4-2-dependent endocytic trafficking

    doi: 10.1152/ajpcell.00383.2014

    Figure Lengend Snippet: Kv7.1 codistributes with late endosomal/lysosomal markers in response to H-89 treatment. A : confocal scans of MDCK-Kv7.1 cells treated with 20 μM H-89 for 3 h and colabeled with the late endosomal/lysosomal marker LAMP-2. Scale bar = 10 ( top )

    Article Snippet: The precleared lysate was subsequently incubated with 3 μg of rabbit anti-Kv7.1 antibody (Alomone Labs), or, as a control, 3 μg normal rabbit IgG (Santa Cruz Biotechnology) for 1 h at 4°C.

    Techniques: Marker

    Small-interfering RNA (siRNA) knockdown of Nedd4-2 protects Kv7.1 from internalization upon PKA inhibition. A : MDCK cells stably expressing Kv7.1 were transiently transfected with siRNA targeting Nedd4-2 or with noncoding control siRNA. Enhanced GFP (eGFP)

    Journal: American Journal of Physiology - Cell Physiology

    Article Title: Protein kinase A stimulates Kv7.1 surface expression by regulating Nedd4-2-dependent endocytic trafficking

    doi: 10.1152/ajpcell.00383.2014

    Figure Lengend Snippet: Small-interfering RNA (siRNA) knockdown of Nedd4-2 protects Kv7.1 from internalization upon PKA inhibition. A : MDCK cells stably expressing Kv7.1 were transiently transfected with siRNA targeting Nedd4-2 or with noncoding control siRNA. Enhanced GFP (eGFP)

    Article Snippet: The precleared lysate was subsequently incubated with 3 μg of rabbit anti-Kv7.1 antibody (Alomone Labs), or, as a control, 3 μg normal rabbit IgG (Santa Cruz Biotechnology) for 1 h at 4°C.

    Techniques: Small Interfering RNA, Inhibition, Stable Transfection, Expressing, Transfection

    Kv7.1-YA localization is unaffected by PKA inhibition. MDCK cells stably expressing the Kv7.1-YA mutant were treated with 20 μM H-89 for 3–5 h. A : confocal scans of MDCK-Kv7.1-YA cells before (0 h) and 3 (3 h H-89) and 5 (5 h H-89) h after

    Journal: American Journal of Physiology - Cell Physiology

    Article Title: Protein kinase A stimulates Kv7.1 surface expression by regulating Nedd4-2-dependent endocytic trafficking

    doi: 10.1152/ajpcell.00383.2014

    Figure Lengend Snippet: Kv7.1-YA localization is unaffected by PKA inhibition. MDCK cells stably expressing the Kv7.1-YA mutant were treated with 20 μM H-89 for 3–5 h. A : confocal scans of MDCK-Kv7.1-YA cells before (0 h) and 3 (3 h H-89) and 5 (5 h H-89) h after

    Article Snippet: The precleared lysate was subsequently incubated with 3 μg of rabbit anti-Kv7.1 antibody (Alomone Labs), or, as a control, 3 μg normal rabbit IgG (Santa Cruz Biotechnology) for 1 h at 4°C.

    Techniques: Inhibition, Stable Transfection, Expressing, Mutagenesis

    Kv7.1 redistribution is not caused by increased Nedd4-2 binding or ubiquitinylation. A : representative Western blot showing the effects of 20 μM H-89 (3 h exposure) and 50 μM forskolin (1 h exposure) on the abundance of S 468 -phosphorylated

    Journal: American Journal of Physiology - Cell Physiology

    Article Title: Protein kinase A stimulates Kv7.1 surface expression by regulating Nedd4-2-dependent endocytic trafficking

    doi: 10.1152/ajpcell.00383.2014

    Figure Lengend Snippet: Kv7.1 redistribution is not caused by increased Nedd4-2 binding or ubiquitinylation. A : representative Western blot showing the effects of 20 μM H-89 (3 h exposure) and 50 μM forskolin (1 h exposure) on the abundance of S 468 -phosphorylated

    Article Snippet: The precleared lysate was subsequently incubated with 3 μg of rabbit anti-Kv7.1 antibody (Alomone Labs), or, as a control, 3 μg normal rabbit IgG (Santa Cruz Biotechnology) for 1 h at 4°C.

    Techniques: Binding Assay, Western Blot

    Kv7.1 accumulates intracellularly upon inhibition of protein kinase A (PKA). Madin-Darby canine kidney (MDCK) cells stably expressing Kv7.1 were subjected to a calcium switch for 24 h followed by treatment with 20 μM H-89 for 3–5 h. A

    Journal: American Journal of Physiology - Cell Physiology

    Article Title: Protein kinase A stimulates Kv7.1 surface expression by regulating Nedd4-2-dependent endocytic trafficking

    doi: 10.1152/ajpcell.00383.2014

    Figure Lengend Snippet: Kv7.1 accumulates intracellularly upon inhibition of protein kinase A (PKA). Madin-Darby canine kidney (MDCK) cells stably expressing Kv7.1 were subjected to a calcium switch for 24 h followed by treatment with 20 μM H-89 for 3–5 h. A

    Article Snippet: The precleared lysate was subsequently incubated with 3 μg of rabbit anti-Kv7.1 antibody (Alomone Labs), or, as a control, 3 μg normal rabbit IgG (Santa Cruz Biotechnology) for 1 h at 4°C.

    Techniques: Inhibition, Stable Transfection, Expressing

    Intracellular accumulation in response to PKA inhibition is specific for Kv7.1. MDCK cells were transfected with Kv7.4 and allowed to polarize before being subjected to either 20 μM H-89 or 10 μM KT-5720 treatments for 3 h. As demonstrated,

    Journal: American Journal of Physiology - Cell Physiology

    Article Title: Protein kinase A stimulates Kv7.1 surface expression by regulating Nedd4-2-dependent endocytic trafficking

    doi: 10.1152/ajpcell.00383.2014

    Figure Lengend Snippet: Intracellular accumulation in response to PKA inhibition is specific for Kv7.1. MDCK cells were transfected with Kv7.4 and allowed to polarize before being subjected to either 20 μM H-89 or 10 μM KT-5720 treatments for 3 h. As demonstrated,

    Article Snippet: The precleared lysate was subsequently incubated with 3 μg of rabbit anti-Kv7.1 antibody (Alomone Labs), or, as a control, 3 μg normal rabbit IgG (Santa Cruz Biotechnology) for 1 h at 4°C.

    Techniques: Inhibition, Transfection

    PKA activation promotes Kv7.1 surface expression in polarized MDCK cells. MDCK cells stably expressing Kv7.1 were grown to confluence and allowed to polarize. The cells were then treated with 50 μM forskolin for 1 h. Cells were fixed before and

    Journal: American Journal of Physiology - Cell Physiology

    Article Title: Protein kinase A stimulates Kv7.1 surface expression by regulating Nedd4-2-dependent endocytic trafficking

    doi: 10.1152/ajpcell.00383.2014

    Figure Lengend Snippet: PKA activation promotes Kv7.1 surface expression in polarized MDCK cells. MDCK cells stably expressing Kv7.1 were grown to confluence and allowed to polarize. The cells were then treated with 50 μM forskolin for 1 h. Cells were fixed before and

    Article Snippet: The precleared lysate was subsequently incubated with 3 μg of rabbit anti-Kv7.1 antibody (Alomone Labs), or, as a control, 3 μg normal rabbit IgG (Santa Cruz Biotechnology) for 1 h at 4°C.

    Techniques: Activation Assay, Expressing, Stable Transfection

    Mimicked phosphorylation of T470 results in reduced membrane expression of Kv7.1, but Kv7.1-T470A is still endocytosed in response to PKA inhibition. Kv7.1-T470A and Kv7.1-T470D mutants were coexpressed with DsRed2-ER in MDCK cells, which were allowed

    Journal: American Journal of Physiology - Cell Physiology

    Article Title: Protein kinase A stimulates Kv7.1 surface expression by regulating Nedd4-2-dependent endocytic trafficking

    doi: 10.1152/ajpcell.00383.2014

    Figure Lengend Snippet: Mimicked phosphorylation of T470 results in reduced membrane expression of Kv7.1, but Kv7.1-T470A is still endocytosed in response to PKA inhibition. Kv7.1-T470A and Kv7.1-T470D mutants were coexpressed with DsRed2-ER in MDCK cells, which were allowed

    Article Snippet: The precleared lysate was subsequently incubated with 3 μg of rabbit anti-Kv7.1 antibody (Alomone Labs), or, as a control, 3 μg normal rabbit IgG (Santa Cruz Biotechnology) for 1 h at 4°C.

    Techniques: Expressing, Inhibition

    Kv7.1 and Kv7.5 expression in optimal cutting temperature compound (OCT)-embedded malignant vascular tumors. Consecutive sections of vascular tumor samples were stained by hematoxylin-eosin (HE) and immunofluorescent labeling for Kv7.1 and Kv7.5. ( Aa – Ac ) Cutaneous angiosarcoma. ( Ba – Bc ) Epithelioid haemangioendothelioma. ( Ca – Cc ) Testicular teratoma-derived angiosarcoma. ( Aa , Ba , Ca ) HE was used to identify structures. BV: healthy blood vessels; *: tumoral blood vessels. Scale bars 150 μm. Representative immunolabeling images showing merged Kv7.1 ( Ab , Bb , Cb ) and Kv7.5 ( Ac , Bc , Cc ) and tumor biopsies. Red, channels; green, α-actin (muscle); blue, DAPI nuclear staining. Scale bars 150 μm. ( D ) Tumor-to-normal (TN) ratios, in arbitrary units, were calculated from the protein expression levels obtained in immunofluorescence assays and represented in a forest plot. The analysis was performed for Kv7.1 ( top ) and Kv7.5 ( bottom ) channels, plotted in horizontal bars. The reference value (1) is indicated by a dotted line. Squares represent the mean ± SEM of the TN ratio for each patient and tumor. Gray, cutaneous angiosarcoma. White, epithelioid haemangioendothelioma. Black, testicular teratoma-derived angiosarcoma. Mean ± SEM, only represented when multiple samples and regions of interest (ROIs) were available.

    Journal: International Journal of Molecular Sciences

    Article Title: Remodeling of Kv7.1 and Kv7.5 Expression in Vascular Tumors

    doi: 10.3390/ijms21176019

    Figure Lengend Snippet: Kv7.1 and Kv7.5 expression in optimal cutting temperature compound (OCT)-embedded malignant vascular tumors. Consecutive sections of vascular tumor samples were stained by hematoxylin-eosin (HE) and immunofluorescent labeling for Kv7.1 and Kv7.5. ( Aa – Ac ) Cutaneous angiosarcoma. ( Ba – Bc ) Epithelioid haemangioendothelioma. ( Ca – Cc ) Testicular teratoma-derived angiosarcoma. ( Aa , Ba , Ca ) HE was used to identify structures. BV: healthy blood vessels; *: tumoral blood vessels. Scale bars 150 μm. Representative immunolabeling images showing merged Kv7.1 ( Ab , Bb , Cb ) and Kv7.5 ( Ac , Bc , Cc ) and tumor biopsies. Red, channels; green, α-actin (muscle); blue, DAPI nuclear staining. Scale bars 150 μm. ( D ) Tumor-to-normal (TN) ratios, in arbitrary units, were calculated from the protein expression levels obtained in immunofluorescence assays and represented in a forest plot. The analysis was performed for Kv7.1 ( top ) and Kv7.5 ( bottom ) channels, plotted in horizontal bars. The reference value (1) is indicated by a dotted line. Squares represent the mean ± SEM of the TN ratio for each patient and tumor. Gray, cutaneous angiosarcoma. White, epithelioid haemangioendothelioma. Black, testicular teratoma-derived angiosarcoma. Mean ± SEM, only represented when multiple samples and regions of interest (ROIs) were available.

    Article Snippet: After blocking for 1 h (5% nonfat powder milk, 10% goat serum, and 0.3% Triton X-100 in PBS-G), samples were incubated with anti-Kv7.1 (1:200, Alomone) or anti-Kv7.5 (1:200, Alomone) and anti-α-actin (1:500, Alomone) antibodies.

    Techniques: Expressing, Staining, Labeling, Derivative Assay, Immunolabeling, Immunofluorescence

    Kv7.1 and Kv7.5 expression according to tumor malignancy. Kv7.1 ( A ) and Kv7.5 ( B ) TN ratios (tumor-to-normal ratios) plotted against their mitotic index as a marker of the clinical aggressiveness of vascular tumors. A Pearson’s correlation coefficient of r = 0.683 with p = 0.061 indicates no apparent correlation between Kv7.1 expression and tumor aggressiveness. A Pearson’s correlation coefficient of r = 0.862 with p = 0.006 indicates a direct correlation between Kv7.5 expression and tumor aggressiveness. Values are the mean ± SEM. Cv: cavernous angioma; Cp: capillary angioma; GT: glomus tumor; KS: Kaposi’s sarcoma; EHE: epithelioid haemangioendothelioma; As: angiosarcoma; TAs: testicular teratoma-derived angiosarcoma; and CAs: cutaneous angiosarcoma.

    Journal: International Journal of Molecular Sciences

    Article Title: Remodeling of Kv7.1 and Kv7.5 Expression in Vascular Tumors

    doi: 10.3390/ijms21176019

    Figure Lengend Snippet: Kv7.1 and Kv7.5 expression according to tumor malignancy. Kv7.1 ( A ) and Kv7.5 ( B ) TN ratios (tumor-to-normal ratios) plotted against their mitotic index as a marker of the clinical aggressiveness of vascular tumors. A Pearson’s correlation coefficient of r = 0.683 with p = 0.061 indicates no apparent correlation between Kv7.1 expression and tumor aggressiveness. A Pearson’s correlation coefficient of r = 0.862 with p = 0.006 indicates a direct correlation between Kv7.5 expression and tumor aggressiveness. Values are the mean ± SEM. Cv: cavernous angioma; Cp: capillary angioma; GT: glomus tumor; KS: Kaposi’s sarcoma; EHE: epithelioid haemangioendothelioma; As: angiosarcoma; TAs: testicular teratoma-derived angiosarcoma; and CAs: cutaneous angiosarcoma.

    Article Snippet: After blocking for 1 h (5% nonfat powder milk, 10% goat serum, and 0.3% Triton X-100 in PBS-G), samples were incubated with anti-Kv7.1 (1:200, Alomone) or anti-Kv7.5 (1:200, Alomone) and anti-α-actin (1:500, Alomone) antibodies.

    Techniques: Expressing, Marker, Derivative Assay

    Voltage-dependent potassium (Kv)7.1 and Kv7.5 are differentially expressed in human blood vessels. ( A ) Membrane protein extracts were obtained from several human blood vessels (aorta and carotid arteries and inferior cava and subclavian veins) and rat tissue samples (brain, skeletal muscle and heart). ( B and C ) Fresh human samples were obtained and processed as described in the Materials and Methods. Sections were stained with anti-Kv7.1 ( B ) or anti-Kv7.5 ( C ) antibodies. Then, α-actin was used to identify muscle structures. Red, Kv7.1 ( B ) and Kv7.5 ( C ) channels; green, α-actin; merge, superimposed channel, α-actin and nuclear DAPI staining (in blue). TI: tunica intima; TM: tunica media; TE: tunica externa; e: endothelial layer; se: subendothelial layer. Scale bars: 50 μm in cava and subclavian veins; 100 μm in carotid artery; 250 μm in aorta artery.

    Journal: International Journal of Molecular Sciences

    Article Title: Remodeling of Kv7.1 and Kv7.5 Expression in Vascular Tumors

    doi: 10.3390/ijms21176019

    Figure Lengend Snippet: Voltage-dependent potassium (Kv)7.1 and Kv7.5 are differentially expressed in human blood vessels. ( A ) Membrane protein extracts were obtained from several human blood vessels (aorta and carotid arteries and inferior cava and subclavian veins) and rat tissue samples (brain, skeletal muscle and heart). ( B and C ) Fresh human samples were obtained and processed as described in the Materials and Methods. Sections were stained with anti-Kv7.1 ( B ) or anti-Kv7.5 ( C ) antibodies. Then, α-actin was used to identify muscle structures. Red, Kv7.1 ( B ) and Kv7.5 ( C ) channels; green, α-actin; merge, superimposed channel, α-actin and nuclear DAPI staining (in blue). TI: tunica intima; TM: tunica media; TE: tunica externa; e: endothelial layer; se: subendothelial layer. Scale bars: 50 μm in cava and subclavian veins; 100 μm in carotid artery; 250 μm in aorta artery.

    Article Snippet: After blocking for 1 h (5% nonfat powder milk, 10% goat serum, and 0.3% Triton X-100 in PBS-G), samples were incubated with anti-Kv7.1 (1:200, Alomone) or anti-Kv7.5 (1:200, Alomone) and anti-α-actin (1:500, Alomone) antibodies.

    Techniques: Staining

    Tumor-to-normal (TN) Kv7.1 and Kv7.5 ratios. TN ratios were calculated as described in the Materials and Methods from protein expression levels in immunofluorescence assays, and the data are represented in a box-and-whisker plot. The analysis was performed for Kv7.1 ( A ) and Kv7.5 ( B ). The reference value (1) is indicated by a dotted vertical line. Boxes represent the interquartile range (IQR: 25–75%) of the values. The inside-box line represents the mean of each group. Minimum and maximum values are indicated outside the box. Cavernous angioma, in magenta; Capillary angioma, in green; Kaposi’s sarcoma, in orange; Angiosarcoma, in blue; Glomus tumor, in red. * p

    Journal: International Journal of Molecular Sciences

    Article Title: Remodeling of Kv7.1 and Kv7.5 Expression in Vascular Tumors

    doi: 10.3390/ijms21176019

    Figure Lengend Snippet: Tumor-to-normal (TN) Kv7.1 and Kv7.5 ratios. TN ratios were calculated as described in the Materials and Methods from protein expression levels in immunofluorescence assays, and the data are represented in a box-and-whisker plot. The analysis was performed for Kv7.1 ( A ) and Kv7.5 ( B ). The reference value (1) is indicated by a dotted vertical line. Boxes represent the interquartile range (IQR: 25–75%) of the values. The inside-box line represents the mean of each group. Minimum and maximum values are indicated outside the box. Cavernous angioma, in magenta; Capillary angioma, in green; Kaposi’s sarcoma, in orange; Angiosarcoma, in blue; Glomus tumor, in red. * p

    Article Snippet: After blocking for 1 h (5% nonfat powder milk, 10% goat serum, and 0.3% Triton X-100 in PBS-G), samples were incubated with anti-Kv7.1 (1:200, Alomone) or anti-Kv7.5 (1:200, Alomone) and anti-α-actin (1:500, Alomone) antibodies.

    Techniques: Expressing, Immunofluorescence, Whisker Assay

    Kv7.1 and Kv7.5 expression in paraffin-embedded vascular tumors. Consecutive sections of vascular tumor samples were stained with hematoxylin-eosin (HE) and were immunofluorescently labeled for Kv7.1 and Kv7.5. ( Aa – Ae ) Cavernous angioma. ( Ba – Be ) Capillary hemangioma. ( Ca – Ce ) Kaposi’s sarcoma. ( Da – De ) Angiosarcoma. ( Ea – Ee ) Glomus tumor. HE was used to identify structures. BV: healthy blood vessels; *: tumoral blood vessels. Scale bars: 50 μm, capillary angioma; 100 μm, cavernous angioma, Kaposi’s sarcoma and angiosarcoma; 200 μm, glomus tumor. Representative immunolabeling images showing merged Kv7.1 in healthy ( Ab , Bb , Cb , Db , Eb ) and tumor ( Ac , Bc , Cc , Dc , Ec ) biopsies. Representative immunolabeling images showing merged Kv7.5 in healthy (Ad, Bd, Cd, Dd, Ed) and tumor ( Ae , Be , Ce , De , Ee ) biopsies. Red, channels; green, α-actin (muscle); blue, DAPI nuclear staining; gray, far red detection of red blood cells showing high autofluorescence Scale bars: 100 μm in cavernous and capillary angioma; 80 μm in Kaposi’s sarcoma, angiosarcoma and glomus tumor.

    Journal: International Journal of Molecular Sciences

    Article Title: Remodeling of Kv7.1 and Kv7.5 Expression in Vascular Tumors

    doi: 10.3390/ijms21176019

    Figure Lengend Snippet: Kv7.1 and Kv7.5 expression in paraffin-embedded vascular tumors. Consecutive sections of vascular tumor samples were stained with hematoxylin-eosin (HE) and were immunofluorescently labeled for Kv7.1 and Kv7.5. ( Aa – Ae ) Cavernous angioma. ( Ba – Be ) Capillary hemangioma. ( Ca – Ce ) Kaposi’s sarcoma. ( Da – De ) Angiosarcoma. ( Ea – Ee ) Glomus tumor. HE was used to identify structures. BV: healthy blood vessels; *: tumoral blood vessels. Scale bars: 50 μm, capillary angioma; 100 μm, cavernous angioma, Kaposi’s sarcoma and angiosarcoma; 200 μm, glomus tumor. Representative immunolabeling images showing merged Kv7.1 in healthy ( Ab , Bb , Cb , Db , Eb ) and tumor ( Ac , Bc , Cc , Dc , Ec ) biopsies. Representative immunolabeling images showing merged Kv7.5 in healthy (Ad, Bd, Cd, Dd, Ed) and tumor ( Ae , Be , Ce , De , Ee ) biopsies. Red, channels; green, α-actin (muscle); blue, DAPI nuclear staining; gray, far red detection of red blood cells showing high autofluorescence Scale bars: 100 μm in cavernous and capillary angioma; 80 μm in Kaposi’s sarcoma, angiosarcoma and glomus tumor.

    Article Snippet: After blocking for 1 h (5% nonfat powder milk, 10% goat serum, and 0.3% Triton X-100 in PBS-G), samples were incubated with anti-Kv7.1 (1:200, Alomone) or anti-Kv7.5 (1:200, Alomone) and anti-α-actin (1:500, Alomone) antibodies.

    Techniques: Expressing, Staining, Labeling, Immunolabeling

    Dopamine activates Kv7/M current in NAc-projecting VTA DA neurons. (A) Kv7/M currents recorded from DA neurons of WT (i,ii) and Kv7.4 −/− (iii,iv) mice. Shown are time-courses of M current amplitudes recorded using whole-cell patch-clamp from VTA slices (for details, see “Materials and Methods” section). Summarized data are presented in (B) . Error bars represent SEM. *** p

    Journal: Frontiers in Cellular Neuroscience

    Article Title: Kv7.4 Channel Contribute to Projection-Specific Auto-Inhibition of Dopamine Neurons in the Ventral Tegmental Area

    doi: 10.3389/fncel.2019.00557

    Figure Lengend Snippet: Dopamine activates Kv7/M current in NAc-projecting VTA DA neurons. (A) Kv7/M currents recorded from DA neurons of WT (i,ii) and Kv7.4 −/− (iii,iv) mice. Shown are time-courses of M current amplitudes recorded using whole-cell patch-clamp from VTA slices (for details, see “Materials and Methods” section). Summarized data are presented in (B) . Error bars represent SEM. *** p

    Article Snippet: Primary antibodies: mouse anti-TH (tyrosine hydrolyze; 1:400, Merck Millipore, Darmstadt, Germany), rabbit anti-Kv7.4 (1:100, AlomoneLabs, Jerusalem, Israel), goat anti-Kir3.2 (1:400, Santa Cruz Biotechnology, Santa Cruz, CA, USA).

    Techniques: Mouse Assay, Patch Clamp

    Kv7.4 contribute to the dopamine (DA)-induced inhibition of NAc-projecting DA neuron firing. (A) Single-cell polymerase chain reaction (PCR) analysis in retrogradely labeled ventral tegmental area (VTA) DA neurons from different projections. (B–D) VTA DA neuron firing recorded with loose cell-attached patch recordings. XE991 (3 μM), a Kv7 blocker, reversed the DA-induced inhibition of neuron firing in WT (B) and Kir3.2 −/− (D) mice, but not in Kv7.4 −/− mice (C) . (E) The effect of DA on spontaneous firing in DA neurons from Kv7.4 −/− /Kir3.2 −/− mice. (F) The percentage of VTA-NAc DA neurons with firing rate being inhibited to less than 20% (complete inhibition) in WT, Kv7.4 −/− , Kir3.2 −/− and Kv7.4 −/− /Kir3.2 −/− mice. (G) Average DA inhibition on firing rate in all recorded VTA-NAc DA neurons from WT, Kv7.4 −/− , Kir3.2 −/− and Kv7.4 −/− /Kir3.2 −/− mice. One-way repeated-measures ANOVA with Bonferroni post hoc test. *** p

    Journal: Frontiers in Cellular Neuroscience

    Article Title: Kv7.4 Channel Contribute to Projection-Specific Auto-Inhibition of Dopamine Neurons in the Ventral Tegmental Area

    doi: 10.3389/fncel.2019.00557

    Figure Lengend Snippet: Kv7.4 contribute to the dopamine (DA)-induced inhibition of NAc-projecting DA neuron firing. (A) Single-cell polymerase chain reaction (PCR) analysis in retrogradely labeled ventral tegmental area (VTA) DA neurons from different projections. (B–D) VTA DA neuron firing recorded with loose cell-attached patch recordings. XE991 (3 μM), a Kv7 blocker, reversed the DA-induced inhibition of neuron firing in WT (B) and Kir3.2 −/− (D) mice, but not in Kv7.4 −/− mice (C) . (E) The effect of DA on spontaneous firing in DA neurons from Kv7.4 −/− /Kir3.2 −/− mice. (F) The percentage of VTA-NAc DA neurons with firing rate being inhibited to less than 20% (complete inhibition) in WT, Kv7.4 −/− , Kir3.2 −/− and Kv7.4 −/− /Kir3.2 −/− mice. (G) Average DA inhibition on firing rate in all recorded VTA-NAc DA neurons from WT, Kv7.4 −/− , Kir3.2 −/− and Kv7.4 −/− /Kir3.2 −/− mice. One-way repeated-measures ANOVA with Bonferroni post hoc test. *** p

    Article Snippet: Primary antibodies: mouse anti-TH (tyrosine hydrolyze; 1:400, Merck Millipore, Darmstadt, Germany), rabbit anti-Kv7.4 (1:100, AlomoneLabs, Jerusalem, Israel), goat anti-Kir3.2 (1:400, Santa Cruz Biotechnology, Santa Cruz, CA, USA).

    Techniques: Inhibition, Polymerase Chain Reaction, Labeling, Mouse Assay

    DA-induced inhibition on the firing of NAc-projecting VTA DA neurons is reduced in the social defeat model of mice. (A,B) Example traces from cell-attached recordings of NAc-projecting VTA DA neurons are shown before and after administration of fasudil (a selective Kv7.4 opener; 10 μM) and DA (20 μM) in the control and the social defeat model mice. (C) Summarized the effect of fasudil and DA on the spontaneous firing frequency of NAc-projecting VTA DA neurons. ** p

    Journal: Frontiers in Cellular Neuroscience

    Article Title: Kv7.4 Channel Contribute to Projection-Specific Auto-Inhibition of Dopamine Neurons in the Ventral Tegmental Area

    doi: 10.3389/fncel.2019.00557

    Figure Lengend Snippet: DA-induced inhibition on the firing of NAc-projecting VTA DA neurons is reduced in the social defeat model of mice. (A,B) Example traces from cell-attached recordings of NAc-projecting VTA DA neurons are shown before and after administration of fasudil (a selective Kv7.4 opener; 10 μM) and DA (20 μM) in the control and the social defeat model mice. (C) Summarized the effect of fasudil and DA on the spontaneous firing frequency of NAc-projecting VTA DA neurons. ** p

    Article Snippet: Primary antibodies: mouse anti-TH (tyrosine hydrolyze; 1:400, Merck Millipore, Darmstadt, Germany), rabbit anti-Kv7.4 (1:100, AlomoneLabs, Jerusalem, Israel), goat anti-Kir3.2 (1:400, Santa Cruz Biotechnology, Santa Cruz, CA, USA).

    Techniques: Inhibition, Mouse Assay

    NP1 interacts and colocalizes with Kv7.2 at presynaptic terminals of excitatory synapses and axonal growth cones. A – D , Representative Western blots of immunoprecipitation eluates separated in 6% Tris-Glycine (for high molecular weight proteins) and 4–12% Bis-Tris gels (for low molecular weight proteins). TL, Total protein lysate; Syx, syntaxin. A , Both native Kv7.2 and syntaxin 1A, but not Kv7.3, coprecipitate with NP1 in total brain extracts. B , Both native Kv7.2 and NP1 coprecipitate with syntaxin in total brain extracts. C , D , Recombinant NP1 coprecipitates Kv7.2, but not syntaxin or Kv7.3, in 293T cells transfected with NP1, 5Myc-Kv7.2, 2HA-Kv7.3, and syntaxin 1A cDNAs. Kv7.2 and Kv7.3 were immunoprecipitated with antibodies against their respective Myc and HA tags. E , F , Immunofluorescence studies and confocal microscopy were performed in high-density ( E ) or low-density ( F ) isolated cortical neurons. E , Top, Confocal sections of 0.772 μm in the z -plane showing immunofluorescence of NP1, VGLUT1, Kv7.2, and negative control (omitting primary antibodies). Bottom, Colocalization (in white) of the excitatory presynaptic marker VGLUT1 (blue) with NP1 (green) and Kv7.2 (red) is shown in a single section with the corresponding orthogonal views of the stack of confocal sections. White arrows indicate sites of colocalization. F , NP1 (green) and KV7.2 (magenta) immunofluorescence and DIC images of an isolated cortical cultured neuron (1×) with its corresponding axonal growth cone highlighted in a white square box, shown in a confocal section of 0.772 μm in the z -plane at higher (5×) magnification. The negative control for primary antibodies is shown in another growth cone on the right. The image in the bottom is the merge of NP1 and Kv7.2 immunofluorescence images in the single confocal section obtained at 5× showing colocalization (white) of NP1 and Kv7.2 in the growth cone, with the corresponding orthogonal views from its respective stack of confocal sections. Images were acquired using restricted spectral emission wavelength ranges chosen to avoid crosstalk or bleed-through between the three different channels. Scale bar, 5 μm.

    Journal: The Journal of Neuroscience

    Article Title: Neuronal Pentraxin 1 Negatively Regulates Excitatory Synapse Density and Synaptic Plasticity

    doi: 10.1523/JNEUROSCI.2548-14.2015

    Figure Lengend Snippet: NP1 interacts and colocalizes with Kv7.2 at presynaptic terminals of excitatory synapses and axonal growth cones. A – D , Representative Western blots of immunoprecipitation eluates separated in 6% Tris-Glycine (for high molecular weight proteins) and 4–12% Bis-Tris gels (for low molecular weight proteins). TL, Total protein lysate; Syx, syntaxin. A , Both native Kv7.2 and syntaxin 1A, but not Kv7.3, coprecipitate with NP1 in total brain extracts. B , Both native Kv7.2 and NP1 coprecipitate with syntaxin in total brain extracts. C , D , Recombinant NP1 coprecipitates Kv7.2, but not syntaxin or Kv7.3, in 293T cells transfected with NP1, 5Myc-Kv7.2, 2HA-Kv7.3, and syntaxin 1A cDNAs. Kv7.2 and Kv7.3 were immunoprecipitated with antibodies against their respective Myc and HA tags. E , F , Immunofluorescence studies and confocal microscopy were performed in high-density ( E ) or low-density ( F ) isolated cortical neurons. E , Top, Confocal sections of 0.772 μm in the z -plane showing immunofluorescence of NP1, VGLUT1, Kv7.2, and negative control (omitting primary antibodies). Bottom, Colocalization (in white) of the excitatory presynaptic marker VGLUT1 (blue) with NP1 (green) and Kv7.2 (red) is shown in a single section with the corresponding orthogonal views of the stack of confocal sections. White arrows indicate sites of colocalization. F , NP1 (green) and KV7.2 (magenta) immunofluorescence and DIC images of an isolated cortical cultured neuron (1×) with its corresponding axonal growth cone highlighted in a white square box, shown in a confocal section of 0.772 μm in the z -plane at higher (5×) magnification. The negative control for primary antibodies is shown in another growth cone on the right. The image in the bottom is the merge of NP1 and Kv7.2 immunofluorescence images in the single confocal section obtained at 5× showing colocalization (white) of NP1 and Kv7.2 in the growth cone, with the corresponding orthogonal views from its respective stack of confocal sections. Images were acquired using restricted spectral emission wavelength ranges chosen to avoid crosstalk or bleed-through between the three different channels. Scale bar, 5 μm.

    Article Snippet: We used the following antibodies: mouse anti-NP1 antibody (1:1000; Becton Dickinson), mouse anti-synaptophysin, clone SY38 (1:1000), mouse anti-PSD95 (1:2000; both from Millipore Bioscience Research Reagents), rabbit anti-Kv7.2 (1:200; Alomone Labs), mouse anti-syntaxin 1A (HPC-1 clone, 1:2000; Sigma-Aldrich), mouse anti-HA (1:3000, Covance), and mouse anti-Myc (9E10 clone, 1:5000; Sigma-Aldrich).

    Techniques: Western Blot, Immunoprecipitation, Molecular Weight, Recombinant, Transfection, Immunofluorescence, Confocal Microscopy, Isolation, Negative Control, Marker, Cell Culture

    Knockdown of NP1 reduces Kv7.2 surface expression and M-current. A , B , The proportion of Kv7.2 at the neuronal surface was determined by biotinylation (30 min at 4°C) of all surface proteins in cultured cortical neurons. The ratio of the surface over the total amount of receptor was determined by densitometric analysis and expressed as a percentage of control values. Only samples of biotinylated proteins lacking immunoreactivity to actin, which indicates contamination with intracellular proteins, were used for analyses. A , Representative Western blots of Kv7.2, Na-KATPase, and actin from total (T) and biotinylated (B) proteins. B , Quantitative analysis of the effect of gain and loss of NP1 on the surface/total levels of the Kv7.2 subunit of the voltage-gated Kv7/M K + channels. Values are mean ± SEM of shRdm ( n = 11), shNP1 ( n = 11), and pWPI-NP1 ( n = 5), from at least three independent experiments; * p

    Journal: The Journal of Neuroscience

    Article Title: Neuronal Pentraxin 1 Negatively Regulates Excitatory Synapse Density and Synaptic Plasticity

    doi: 10.1523/JNEUROSCI.2548-14.2015

    Figure Lengend Snippet: Knockdown of NP1 reduces Kv7.2 surface expression and M-current. A , B , The proportion of Kv7.2 at the neuronal surface was determined by biotinylation (30 min at 4°C) of all surface proteins in cultured cortical neurons. The ratio of the surface over the total amount of receptor was determined by densitometric analysis and expressed as a percentage of control values. Only samples of biotinylated proteins lacking immunoreactivity to actin, which indicates contamination with intracellular proteins, were used for analyses. A , Representative Western blots of Kv7.2, Na-KATPase, and actin from total (T) and biotinylated (B) proteins. B , Quantitative analysis of the effect of gain and loss of NP1 on the surface/total levels of the Kv7.2 subunit of the voltage-gated Kv7/M K + channels. Values are mean ± SEM of shRdm ( n = 11), shNP1 ( n = 11), and pWPI-NP1 ( n = 5), from at least three independent experiments; * p

    Article Snippet: We used the following antibodies: mouse anti-NP1 antibody (1:1000; Becton Dickinson), mouse anti-synaptophysin, clone SY38 (1:1000), mouse anti-PSD95 (1:2000; both from Millipore Bioscience Research Reagents), rabbit anti-Kv7.2 (1:200; Alomone Labs), mouse anti-syntaxin 1A (HPC-1 clone, 1:2000; Sigma-Aldrich), mouse anti-HA (1:3000, Covance), and mouse anti-Myc (9E10 clone, 1:5000; Sigma-Aldrich).

    Techniques: Expressing, Cell Culture, Western Blot