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Enhancement effects of martentoxin on glioma BK channel and BK channel (?+?1) subtypes.

ABSTRACT: BACKGROUND: BK channels are usually activated by membrane depolarization and cytoplasmic Ca(2+). Especially,the activity of BK channel (?+?4) can be modulated by martentoxin, a 37 residues peptide, with Ca(2+)-dependent manner. gBK channel (glioma BK channel) and BK channel (?+?1) possessed higher Ca(2+) sensitivity than other known BK channel subtypes. METHODOLOGY AND PRINCIPAL FINDINGS: The present study investigated the modulatory characteristics of martentoxin on these two BK channel subtypes by electrophysiological recordings, cell proliferation and Ca(2+) imaging. In the presence of cytoplasmic Ca(2+), martentoxin could enhance the activities of both gBK and BK channel (?+?1) subtypes in dose-dependent manner with EC(50) of 46.7 nM and 495 nM respectively, while not shift the steady-state activation of these channels. The enhancement ratio of martentoxin on gBK and BK channel (?+?1) was unrelated to the quantitative change of cytoplasmic Ca(2+) concentrations though the interaction between martentoxin and BK channel (?+?1) was accelerated under higher cytoplasmic Ca(2+). The selective BK pore blocker iberiotoxin could fully abolish the enhancement of these two BK subtypes induced by martentoxin, suggesting that the auxiliary ? subunit might contribute to the docking for martentoxin. However, in the absence of cytoplasmic Ca(2+), the activity of gBK channel would be surprisingly inhibited by martentoxin while BK channel (?+?1) couldn't be affected by the toxin. CONCLUSIONS AND SIGNIFICANCE: Thus, the results shown here provide the novel evidence that martentoxin could increase the two Ca(2+)-hypersensitive BK channel subtypes activities in a new manner and indicate that ? subunit of these BK channels plays a vital role in this enhancement by martentoxin.

SUBMITTER: Tao J

PROVIDER: S-EPMC3060806 | biostudies-literature | 2011

REPOSITORIES: biostudies-literature

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Enhancement effects of martentoxin on glioma BK channel and BK channel (α+β1) subtypes.

Tao Jie J Shi Jian J Yan Li L Chen Ying Y Duan Yan Hong YH Ye Pin P Feng Qi Q Zhang Jian Wei JW Shu Xue Qin XQ Ji Yong Hua YH

PloS one 20110318 3

<h4>Background</h4>BK channels are usually activated by membrane depolarization and cytoplasmic Ca(2+). Especially,the activity of BK channel (α+β4) can be modulated by martentoxin, a 37 residues peptide, with Ca(2+)-dependent manner. gBK channel (glioma BK channel) and BK channel (α+β1) possessed higher Ca(2+) sensitivity than other known BK channel subtypes.<h4>Methodology and principal findings</h4>The present study investigated the modulatory characteristics of martentoxin on these two BK ch ...[more]

PMID: 21445248

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Project description:BackgroundThe large conductance Ca(2+) - and voltage-activated K(+) channel (BK) is an important player in molecular and behavioral alcohol tolerance. Trafficking and surface expression of ion channels contribute to the development of addictive behaviors. We have previously reported that internalization of the BK channel is a component of molecular tolerance to ethanol (EtOH).MethodsUsing primary cultures of hippocampal neurons, we combine total internal reflection fluorescence microscopy, electrophysiology, and biochemical techniques to explore how exposure to EtOH affects the expression and subcellular localization of endogenous BK channels over time.ResultsExposure to EtOH changed the expression of endogenous BK channels in a time-dependent manner at the perimembrane area (plasma membrane and/or the area adjacent to it), while total protein levels of BK remain unchanged. These results suggest a redistribution of the channel within the neurons rather than changes in synthesis or degradation rates. Our results showed a temporally nonlinear effect of EtOH on perimembrane expression of BK. First, there was an increase in BK perimembrane expression after 10 minutes of EtOH exposure that remained evident after 3 hours, although not correlated to increases in functional channel expression. In contrast, after 6 hours of EtOH exposure, we observed a significant decrease in both BK perimembrane expression and functional channel expression. Furthermore, after 24 hours of EtOH exposure, perimembrane levels of BK had returned to baseline.ConclusionsWe report a complex time-dependent pattern in the effect of EtOH on BK channel trafficking, including successive increases and decreases in perimembrane expression and a reduction in active BK channels after 3 and 6 hours of EtOH exposure. Possible mechanisms underlying this multiphasic trafficking are discussed. As molecular tolerance necessarily underlies behavioral tolerance, the time-dependent alterations we see at the level of the channel may be relevant to the influence of drinking patterns on the development of behavioral tolerance.

Project description:KCNMA1, encoding the voltage- and calcium-activated potassium channel, has a pivotal role in brain physiology. Mutations in KCNMA1 are associated with epilepsy and/or dyskinesia (PNKD3). Two KCNMA1 mutations correlated with these phenotypes, D434G and N999S, were previously identified as producing gain-of-function (GOF) effects on BK channel activity. Three new patients have been reported harboring N999S, one carrying a second mutation, R1128W, but the effects of these mutations have not yet been reported under physiological K+ conditions or compared to D434G. In this study, we characterize N999S, the novel N999S/R1128W double mutation, and D434G in a brain BK channel splice variant, comparing the effects on BK current properties under a physiological K+ gradient with action potential voltage commands. N999S, N999S/R1128W, and D434G cDNAs were expressed in HEK293T cells and characterized by patch-clamp electrophysiology. N999S BK currents were shifted to negative potentials, with faster activation and slower deactivation compared with wild type (WT) and D434G. The double mutation N999S/R1128W did not show any additional changes in current properties compared with N999S alone. The antiepileptic drug acetazolamide was assessed for its ability to directly modulate WT and N999S channels. Neither the WT nor N999S channels were sensitive to the antiepileptic drug acetazolamide, but both were sensitive to the inhibitor paxilline. We conclude that N999S is a strong GOF mutation that surpasses the D434G phenotype, without mitigation by R1128W. Acetazolamide has no direct modulatory action on either WT or N999S channels, indicating that its use may not be contraindicated in patients harboring GOF KCNMA1 mutations.NEW & NOTEWORTHY KCNMA1-linked channelopathy is a new neurological disorder characterized by mutations in the BK voltage- and calcium-activated potassium channel. The epilepsy- and dyskinesia-associated gain-of-function mutations N999S and D434G comprise the largest number of patients in the cohort. This study provides the first direct comparison between D434G and N999S BK channel properties as well as a novel double mutation, N999S/R1128W, from another patient, defining the functional effects during an action potential stimulus.

Dataset Information

Enhancement effects of martentoxin on glioma BK channel and BK channel (?+?1) subtypes.

Publications

Enhancement effects of martentoxin on glioma BK channel and BK channel (α+β1) subtypes.

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