MECHANISMS OF DEPENDENCE PATHOGENESIS

依赖性发病机制

• 批准号: 6712919
• 负责人: A LESLIE MORROW
• 金额: $ 16.96万
• 依托单位: UNIV OF NORTH CAROLINA CHAPEL HILL
• 依托单位国家: 美国
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-12-27 至 2007-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6712919
• 关键词: GABA receptor; SDS polyacrylamide gel electrophoresis; alcoholic beverage consumption; amygdala; anxiety; behavioral /social science research tag; cerebral cortex; drug addiction; drug tolerance; drug withdrawal; epilepsy; ethanol; gene targeting; genetically modified animals; hypothalamus; immunoprecipitation; laboratory mouse; laboratory rat; phosphorylation; protein kinase C; protein protein interaction; receptor expression; western blottings

The overall goal of this proposal is to elucidate the molecular mechanisms that underlie alterations in gamma-aminobutyric acid (GABA)A receptor adaptations that influence the development of ethanol dependence. Ethanol has several sites of action in the brain, but direct or indirect modulation of GABA/A receptors may behavioral actions of ethanol. Moreover, prolonged ethanol consumption results in the development of tolerance and dependence upon ethanol. Withdrawal from ethanol, and particularly repeated withdrawals from ethanol, produce marked increases in CNS excitability and anxiety. Substantial evidence suggests that these behavioral and neural adaptations involve marked adaptations in the pharmacological properties of GABA/A receptors. Furthermore, research over the previous funding period has established that GABA/A receptor submit adaptations accompany these changes and differ markedly across brain regions. We plan to focus on the role of PKCgamma and PKCepsilon in mediating GABA/A receptor adaptations. We hypothesize that PKC interactions with GABA/A receptors may determine receptor subunit adaptations and may underlie the regional differences in these differences in these adaptations. Specific Aim 1 will determine if ethanol dependence alters the association of GABA/A receptors with PKC isozymes in alter ethanol-induced adaptatins in GABA/A receptor function and seizure susceptibility. Specific Aim 2 will utilize these mice to determine if PKCgamma and PKCepsilon differentially alter the effects of ethanol on membrane expression and internalization to alter specific GABA/A receptors. The final aim will investigate the role of PKCgamma and PKCepsilon in the phosphorylation state of GABA/A receptors, again using mutant mouse models. Vector-mediated gene delivery for tissue specific rescue of PKCgamma or PKCepsilon in vivo 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. Tissue specific rescue of PKCgamma or PKCepsilon as well as pharmacological challenge with PKC antagonists will also be used to control for the possibility that adaptations of other proteins contribute to the effects of genetic deletion of PKCgamma or PKCepsilon. We predict that these experiments will delineate specific GABA/A receptor adaptations involved in ethanol dependence-induced enhancement of seizure susceptibility (bicuculline seizure threshold) and ethanol self-administration (in collaboration with Clyde Hodge). These studies will provide important mechanistic information on the molecular basis of ethanol-induced adaptations in GABA/A receptors that influence the development of ethanol tolerance and dependence.

该提案的总体目标是阐明γ-氨基丁酸（GABA）一种受体适应的分子机制，这些受体适应会影响乙醇依赖性的发展。 乙醇在大脑中具有多种作用部位，但是GABA/A受体的直接或间接调节可能的行为作用可能是乙醇的行为作用。 此外，延长的乙醇消耗会导致耐受性和对乙醇的依赖性的发展。 从乙醇中提取，尤其是从乙醇中反复退出，在中枢神经系统兴奋性和焦虑中产生明显增加。 大量证据表明，这些行为和神经适应性涉及GABA/A受体的药理特性的明显适应。 此外，在以前的资金期间的研究表明，GABA/A受体提交适应性伴随着这些变化，并且在大脑区域之间明显不同。 我们计划专注于Pkcgamma和Pkcepsilon在介导GABA/A受体适应性中的作用。 我们假设PKC与GABA/A受体的相互作用可能决定受体亚基适应，并可能是这些适应性差异的区域差异的基础。 具体目标1将确定乙醇依赖性是否改变了GABA/A受体与PKC同工酶在Alter乙醇诱导的适应蛋白中的相关性，在GABA/A受体功能和癫痫发作易感性中。 具体的目标2将利用这些小鼠来确定pkcgamma和pkcepsilon是否会差异地改变乙醇对膜表达和内在化的影响以改变特定的GABA/A受体。 最终目的将再次使用突变小鼠模型研究pkcgamma和pkcepsilon在GABA/A受体的磷酸化状态中的作用。 载体介导的基因递送用于组织特异性的PKCGAMMA或PKCEPSILON体内的基因递送，用于在PKC的改变与受体膜表达，内在化和功能的影响之间建立原因和作用关系。 PKCGAMMA或PKCEPSILON的组织特异性营救以及对PKC拮抗剂的药理挑战也将用于控制其他蛋白质的适应性可能导致PKCGAMMA或PKCEPSILON遗传缺失的影响。 我们预测，这些实验将描绘出参与乙醇依赖性诱导的癫痫发作易感性（双瓜氨酸癫痫发作阈值）和乙醇自我给药（与Clyde Hodge的协作）的特定GABA/A受体适应。 这些研究将提供有关影响乙醇耐受性和依赖性发展的GABA/A受体中乙醇诱导适应的分子基础的重要机械信息。