Central Nicotinic Receptors and Epileptic Seizures

中枢烟碱受体和癫痫发作

基本信息

批准号：
6529791
负责人：
BRUCE N COHEN
金额：
$ 19.75万
依托单位：
CALIFORNIA INSTITUTE OF TECHNOLOGY
依托单位国家：
美国
项目类别：
财政年份：
2001
资助国家：
美国
起止时间：
2001-09-04 至 2004-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/6529791
关键词：
Xenopus oocyte acetylcholine calcium flux calcium indicator epilepsy gene mutation genetic disorder membrane potentials neural transmission neurotransmitter transport nicotinic receptors protein structure function receptor expression sleep tissue /cell culture voltage /patch clamp

项目摘要

Genetic defects cause 20-40 percent of all epilepsies. Several types of inherited epilepsy appear to be linked to nicotinic receptors. However, autosomal dominant nocturnal frontal lobe epilepsy (ADNFLE) is the only one that has been linked to specific nicotinic mutations -alpha4(S248F), alpha4(777ins3), and alpha4(S252L). These mutations produce brief, repetitive seizures that occur primarily during phase 2 sleep. The mechanism of seizure generation has not been established. We hypothesize that a reduction in Ca2+-induced potentiation of the acetylcholine (ACH) response causes ADNFLE seizures. Our preliminary data show that all three presently identified ADNFLE mutations reduce Ca2+ potentiation of the ACH response. However, previous studies show that the alpha4(S248F) mutation also reduces the Ca2+ permeability of the receptor relative to Na+ (Pca/PNa) and, that the alpha4(S248F) and alpha4(777ins3) mutations enhance ACh-induced receptor desensitization. Aim 1 is to determine whether reduced Ca2+ potentiation is the only common functional effect of the ADNFLE mutations. We will express rat versions (S252F, +L264, S256L) of the human ADNFLE mutations with wild-type (WT) rat beta2 subunits in Xenopus oocytes and examine their effects on desensitization, surface expression, receptor turnover, and choline potentiation of the ACH response. Aim 2 is to determine whether all three ADNFLE mutations reduce the Pca/PNa and Ca2+ influx through the receptors. Aim 3 is to demonstrate that the ADNFLE mutations reduce Ca2+ potentiation by enhancing Ca2+ block of the receptor rather than by reducing Ca2+ potentiation per se. Aim 4 is to characterize the mechanism of mutant-induced reductions in Ca2+ potentiation at the single-channel level. Our results should elucidate the connection between central nicotinic receptors and epileptic seizures.

20-40% 的癫痫症是由遗传缺陷引起的。几种类型的遗传性癫痫似乎与烟碱受体有关。然而，常染色体显性遗传性夜间额叶癫痫 (ADNFLE) 是唯一一种与特定烟碱突变 -alpha4(S248F)、alpha4(777ins3) 和 alpha4(S252L) 有关的疾病。这些突变会产生短暂、重复的癫痫发作，主要发生在第二阶段睡眠期间。癫痫发作的发生机制尚未确定。我们假设 Ca2+ 诱导的乙酰胆碱 (ACH) 反应增强的减少导致 ADNFLE 癫痫发作。我们的初步数据表明，目前发现的所有三种 ADNFLE 突变都会降低 ACH 反应的 Ca2+ 增强作用。然而，先前的研究表明，α4(S248F) 突变还降低了受体相对于 Na+ (Pca/PNa) 的 Ca2+ 通透性，并且 α4(S248F) 和 α4(777ins3) 突变增强了 ACh 诱导的受体脱敏。目标 1 是确定 Ca2+ 增强作用降低是否是 ADNFLE 突变的唯一常见功能效应。我们将在爪蟾卵母细胞中表达人类 ADNFLE 突变的大鼠版本（S252F、+L264、S256L）与野生型（WT）大鼠 β2 亚基，并检查它们对 ACH 脱敏、表面表达、受体周转和胆碱增强的影响回复。目标 2 是确定所有三种 ADNFLE 突变是否都会减少 Pca/PNa 和 Ca2+ 通过受体的流入。目标 3 是证明 ADNFLE 突变通过增强受体的 Ca2+ 阻断而不是降低 Ca2+ 增强本身来降低 Ca2+ 增强。目标 4 是在单通道水平上表征突变体诱导的 Ca2+ 增强降低的机制。我们的结果应该阐明中枢烟碱受体与癫痫发作之间的联系。