rabbit anti kv1 1  (Alomone Labs)

Journal: Proceedings of the National Academy of Sciences of the United States of America

Article Title: Kv1.1 channelopathy abolishes presynaptic spike width modulation by subthreshold somatic depolarization

doi: 10.1073/pnas.1608763114

Figure Lengend Snippet: Kv1.1 channels determine spike width. ( A ) Example recordings from one neuron before and after 20 nM DTx-K application. This neurotoxin had no effect on somatic spikes but led to broadening of presynaptic action potentials. ( B ) Presynaptic spike widths

Article Snippet: Membranes were immunoblotted with the respective antibodies: rabbit anti-Kv1.1 (APC-161, Alomone Labs; 1:800), anti-Kv1.2 (APC-010, 1:1,500), anti-Kv1.3 (APC-101, 1:1,000), anti-Kv1.4 (APC-167, 1:500), anti-Kv1.6 (APC-003, 1:2,000), and anti-SNAP25 (Abcam ab5666; 1:2,000) at 4 °C overnight.

Techniques:

Journal: Proceedings of the National Academy of Sciences of the United States of America

Article Title: Kv1.1 channelopathy abolishes presynaptic spike width modulation by subthreshold somatic depolarization

doi: 10.1073/pnas.1608763114

Figure Lengend Snippet: Blockade or deletion of Kv1.1 does not prevent analog modulation of presynaptic spike width. ( A ) Trace analysis from one wild-type cell showing bidirectional changes in presynaptic spike width by subthreshold somatic current injections before evoking

Article Snippet: Membranes were immunoblotted with the respective antibodies: rabbit anti-Kv1.1 (APC-161, Alomone Labs; 1:800), anti-Kv1.2 (APC-010, 1:1,500), anti-Kv1.3 (APC-101, 1:1,000), anti-Kv1.4 (APC-167, 1:500), anti-Kv1.6 (APC-003, 1:2,000), and anti-SNAP25 (Abcam ab5666; 1:2,000) at 4 °C overnight.

Techniques:

Journal: BMC Biology

Article Title: The mono-ADP-ribosyltransferase ARTD10 regulates the voltage-gated K+ channel Kv1.1 through protein kinase C delta

doi: 10.1186/s12915-020-00878-1

Figure Lengend Snippet: ARTD10 inhibition reduces the proportion of the inactivating Kv1.1 current and enhances spontaneous excitation in hippocampal neurons. a Left, whole cell currents of mouse hippocampal neurons in the presence of tetrodotoxin with and without an inhibitor of ARTD10 (OUL35). Right, bar graphs representing I steady-state / I peak ( I s / I p ) and peak current amplitudes. b Resting membrane potential (RMP) and spontaneous spikes/s from control and OUL35 treated hippocampal neurons. * p

Article Snippet: For immunoblots, proteins were transferred to PVDF membranes (Roche, Mannheim, Germany), and the membrane was blocked for 1 h at RT in 5% non-fat milk in TBS-T (137 mM NaCl, 2.7 mM KCl, 25 mM Tris, 0.1% Tween-20), and probed overnight at 4 °C with the following primary antibodies: rabbit polyclonal anti-Kv1.1 (Alomone Labs, # APC-161 and APC-009), rabbit polyclonal anti-PKCδ (Santa Cruz, # sc-937), rabbit polyclonal anti-phospho-PKCδ (Tyr311) (Cell Signaling, # 2055), rat monoclonal anti-ARTD10 (Merck, #5H11), or mouse monoclonal anti-acetylated tubulin (Sigma-Aldrich, # T7451).

Techniques: Inhibition

Journal: BMC Biology

Article Title: The mono-ADP-ribosyltransferase ARTD10 regulates the voltage-gated K+ channel Kv1.1 through protein kinase C delta

doi: 10.1186/s12915-020-00878-1

Figure Lengend Snippet: Scheme illustrating the interplay of PKA, PKCδ, and ARTD10 in the regulation of Kv1.1. The regulation of a phosphatase by PKCδ is hypothetical. Current traces above the schemes of the Kv1.1 channels illustrate the typical inactivation pattern of phosphorylated vs. un-phosphorylated Kv1.1α. I/F, IBMX/forskolin; PMA, phorbol-myristate-acetate

Article Snippet: For immunoblots, proteins were transferred to PVDF membranes (Roche, Mannheim, Germany), and the membrane was blocked for 1 h at RT in 5% non-fat milk in TBS-T (137 mM NaCl, 2.7 mM KCl, 25 mM Tris, 0.1% Tween-20), and probed overnight at 4 °C with the following primary antibodies: rabbit polyclonal anti-Kv1.1 (Alomone Labs, # APC-161 and APC-009), rabbit polyclonal anti-PKCδ (Santa Cruz, # sc-937), rabbit polyclonal anti-phospho-PKCδ (Tyr311) (Cell Signaling, # 2055), rat monoclonal anti-ARTD10 (Merck, #5H11), or mouse monoclonal anti-acetylated tubulin (Sigma-Aldrich, # T7451).

Techniques:

Journal: BMC Biology

Article Title: The mono-ADP-ribosyltransferase ARTD10 regulates the voltage-gated K+ channel Kv1.1 through protein kinase C delta

doi: 10.1186/s12915-020-00878-1

Figure Lengend Snippet: ARTD10 inhibition enhances excitability of hippocampal neurons via Kv1.1. a Left, representative current clamp recordings of APs elicited by step current pulses in control neurons and neurons treated with OUL35. Right, bar graphs represent the rheobase and the latency to the first spike. For cells with a RMP more positive than − 60 mV, the membrane potential was adjusted to ~ − 60 mV. b The number of spikes elicited by step current pulses were counted and for stimuli from 10 to 30 pA they were fitted with a linear function. Right, bar graphs summarize the AP amplitude from neurons with and without OUL35 treatment. c Left, bar graphs representing the rheobase. Right, summary of spikes/s with and without the Kv1 inhibitor α-dendrotoxin (DTX) and from neurons with and without OUL35 treatment. * p

Article Snippet: For immunoblots, proteins were transferred to PVDF membranes (Roche, Mannheim, Germany), and the membrane was blocked for 1 h at RT in 5% non-fat milk in TBS-T (137 mM NaCl, 2.7 mM KCl, 25 mM Tris, 0.1% Tween-20), and probed overnight at 4 °C with the following primary antibodies: rabbit polyclonal anti-Kv1.1 (Alomone Labs, # APC-161 and APC-009), rabbit polyclonal anti-PKCδ (Santa Cruz, # sc-937), rabbit polyclonal anti-phospho-PKCδ (Tyr311) (Cell Signaling, # 2055), rat monoclonal anti-ARTD10 (Merck, #5H11), or mouse monoclonal anti-acetylated tubulin (Sigma-Aldrich, # T7451).

Techniques: Inhibition

Journal: BMC Biology

Article Title: The mono-ADP-ribosyltransferase ARTD10 regulates the voltage-gated K+ channel Kv1.1 through protein kinase C delta

doi: 10.1186/s12915-020-00878-1

Figure Lengend Snippet: Inactivation of Kv1.1 is regulated by Kvβ and phosphorylation at S446. a Representative recordings, I steady-state / I peak ( I s / I p ), and peak current amplitudes of Kv1.1 with or without co-expression of Kvβ1.1 in HeLa cells. b Representative recordings and I s / I p before and after the application of either IBMX/forskolin (I/F) or of the phorbol ester PMA. c Representative recordings, I s / I p , and peak current amplitudes of the phosphorylation-deficient mutant Kv1.1 S446A co-expressed with Kvβ1.1. d Representative recordings, I s / I p , and peak current amplitudes of Kv1.1 after 1.5 h pre-incubation with a phosphatase inhibitor cocktail. * p

Article Snippet: For immunoblots, proteins were transferred to PVDF membranes (Roche, Mannheim, Germany), and the membrane was blocked for 1 h at RT in 5% non-fat milk in TBS-T (137 mM NaCl, 2.7 mM KCl, 25 mM Tris, 0.1% Tween-20), and probed overnight at 4 °C with the following primary antibodies: rabbit polyclonal anti-Kv1.1 (Alomone Labs, # APC-161 and APC-009), rabbit polyclonal anti-PKCδ (Santa Cruz, # sc-937), rabbit polyclonal anti-phospho-PKCδ (Tyr311) (Cell Signaling, # 2055), rat monoclonal anti-ARTD10 (Merck, #5H11), or mouse monoclonal anti-acetylated tubulin (Sigma-Aldrich, # T7451).

Techniques: Expressing, Mutagenesis, Incubation

Journal: BMC Biology

Article Title: The mono-ADP-ribosyltransferase ARTD10 regulates the voltage-gated K+ channel Kv1.1 through protein kinase C delta

doi: 10.1186/s12915-020-00878-1

Figure Lengend Snippet: ARTD10 leads to phosphorylation of Kv1.1 at S446. a , b Left, Representative current traces of wild type ( a ) or the phosphorylation-deficient mutant ( b ) of Kv1.1 overexpressed in three different HeLa cell lines. Right, bar graphs representing I steady-state / I peak ( I s / I p ) and peak current amplitudes. * p

Article Snippet: For immunoblots, proteins were transferred to PVDF membranes (Roche, Mannheim, Germany), and the membrane was blocked for 1 h at RT in 5% non-fat milk in TBS-T (137 mM NaCl, 2.7 mM KCl, 25 mM Tris, 0.1% Tween-20), and probed overnight at 4 °C with the following primary antibodies: rabbit polyclonal anti-Kv1.1 (Alomone Labs, # APC-161 and APC-009), rabbit polyclonal anti-PKCδ (Santa Cruz, # sc-937), rabbit polyclonal anti-phospho-PKCδ (Tyr311) (Cell Signaling, # 2055), rat monoclonal anti-ARTD10 (Merck, #5H11), or mouse monoclonal anti-acetylated tubulin (Sigma-Aldrich, # T7451).

Techniques: Mutagenesis

Journal: BMC Biology

Article Title: The mono-ADP-ribosyltransferase ARTD10 regulates the voltage-gated K+ channel Kv1.1 through protein kinase C delta

doi: 10.1186/s12915-020-00878-1

Figure Lengend Snippet: Kv1.1 is regulated by PKCδ in HeLa cells. a , b Wild type Kv1.1 or mutant Kv1.1-S446A were co-expressed with Kvβ1 and either the catalytic domain (CAT) or a dominant negative mutant (DN) of PKCδ. Bar graphs represent I steady-state / I peak ( I s / I p ) and peak current amplitudes

Article Snippet: For immunoblots, proteins were transferred to PVDF membranes (Roche, Mannheim, Germany), and the membrane was blocked for 1 h at RT in 5% non-fat milk in TBS-T (137 mM NaCl, 2.7 mM KCl, 25 mM Tris, 0.1% Tween-20), and probed overnight at 4 °C with the following primary antibodies: rabbit polyclonal anti-Kv1.1 (Alomone Labs, # APC-161 and APC-009), rabbit polyclonal anti-PKCδ (Santa Cruz, # sc-937), rabbit polyclonal anti-phospho-PKCδ (Tyr311) (Cell Signaling, # 2055), rat monoclonal anti-ARTD10 (Merck, #5H11), or mouse monoclonal anti-acetylated tubulin (Sigma-Aldrich, # T7451).

Techniques: Mutagenesis, Dominant Negative Mutation