Ethanol Regulation of GABAalpha receptors

乙醇对 GABAα 受体的调节

• 批准号: 7032457
• 负责人: SANDEEP KUMAR
• 金额: $ 10.15万
• 依托单位: UNIV OF NORTH CAROLINA CHAPEL HILL
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-03-15 至 2010-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7032457
• 关键词: GABA receptor; SDS polyacrylamide gel electrophoresis; alcoholism /alcohol abuse; amygdala; brain electrical activity; cerebral cortex; confocal scanning microscopy; enzyme activity; epilepsy; gene deletion mutation; genetically modified animals; hippocampus; hypothalamus; isozymes; laboratory mouse; laboratory rat; phosphorylation; protein isoforms; protein kinase C; protein protein interaction; receptor expression; western blottings

DESCRIPTION (provided by applicant): The overall goal of this proposal is to provide mentored career development training for a young physician scientist. The primary vehicle of this mentored training involves the development of research to elucidate the molecular mechanisms that underlie alterations in gamma-aminobutyric acid (GABA)A receptor adaptations that influence the development of ethanol dependence. The candidate will receive broad-based training in alcoholism research as well as several methods that will be required to address his research plan including confocal microscopy, transgenic mouse technology, intracerebroventricular (ICV) administration of drugs and measurement of seizure thresholds. In addition, he will be guided to develop new methods to investigate the phosphorylation of GABAA receptors. The research plan will focus on GABAA receptor adaptations since ethanol dependence produces marked increases in CNS excitability and anxiety that involve adaptations in the pharmacological properties of GABAA receptors. Dr. Kumar proposes to focus on the role of receptor trafficking, phosphorylation and PKC activity in mediating GABAA receptor adaptations. He will test the hypothesis that PKC interactions with GABAA receptors may determine receptor subunit adaptations throughout brain. Specific Aim 1 will determine if ethanol dependence alters the association of GABAA receptors with PKC isozymes in cerebral cortex, hypothalamus and amygdala. Next, he will determine if genetic deletion of PKCgamma or PKCepsilon isozymes alter ethanol-induced adaptations in GABAA receptor function and seizure susceptibility. Specific Aim 2 will utilize these mice to determine if PKCy and PKCe differentially alter the effects of ethanol on membrane expression and internalization of specific GABAA receptors. The final aim will investigate the role of PKC'y and PKCe in the phosphorylation state of GABA! receptors, again using mutant mouse models. ICV administration of PKC isoform specific inhibitor/activator will be used to establish a cause and effect relationship between the alterations in PKC and subsequent effects on receptor membrane expression, internalization and function. We predict that these experiments will delineate specific GABAA receptor adaptations involved in ethanol dependence-induced enhancement of seizure susceptibility (bicuculline seizure threshold). These studies will provide important mechanistic information on the molecular basis of ethanol-induced adaptations in GABAA receptors that influence the development of ethanol tolerance and dependence.

描述（由申请人提供）：该提案的总体目标是为年轻的医师科学家提供指导的职业发展培训。 这项指导训练的主要工具涉及研究的发展，以阐明γ-氨基丁酸（GABA）改变的分子机制，一种受体适应，影响乙醇依赖性的发展。候选人将接受基于酒精中毒研究的广泛培训，以及几种解决他的研究计划所需的方法，包括共聚焦显微镜，转基因小鼠技术，药物内脑室内（ICV）给药以及癫痫发作阈值的测量。此外，他将被指导开发新方法来研究GABAA受体的磷酸化。该研究计划将重点关注GABAA受体适应，因为乙醇依赖性在CNS兴奋性和焦虑中产生明显增加，涉及GABAA受体的药理特性的适应性。 Kumar博士建议专注于受体运输，磷酸化和PKC活性在介导GABAA受体适应中的作用。 他将检验以下假设：PKC与GABAA受体的相互作用可能确定整个大脑的受体亚基适应。具体目标1将确定乙醇依赖性是否改变了脑皮质，下丘脑和杏仁核中GABAA受体与PKC同工酶的缔合。接下来，他将确定PKCGAMMA或PKCEPSILON同工酶的遗传缺失是否改变了乙醇诱导的GABAA受体功能和癫痫发作易感性的适应性。具体目标2将利用这些小鼠来确定PKCY和PKCE是否会差异地改变乙醇对特定GABAA受体的膜表达和内在化的影响。最终目标将研究PKC'Y和PKCE在GABA磷酸化状态中的作用！受体，再次使用突变小鼠模型。 PKC同工型特异性抑制剂/激活剂的ICV施用将用于建立PKC改变与随后对受体膜表达，内在化和功能的影响之间的因果关系。我们预测，这些实验将描述参与乙醇依赖性诱导的癫痫易感性增强（双核氨酸癫痫发作阈值）的特定GABAA受体适应。这些研究将提供有关乙醇诱导的GABAA受体适应性的分子基础的重要机械信息，从而影响乙醇耐受性和依赖性的发展。