Ethanol Sensitivity of Native and Cloned NMDA Receptors

描述（由申请人提供）：了解酒精对大脑功能的影响对于开发更好的酒精相关问题治疗方法至关重要。在一个正在进行的研究项目的更新中，我们专注于了解酒精如何影响神经元表达的关键离子通道的功能。该通道，即N-甲基-D-天冬氨酸（NMDA）受体，在兴奋性神经递质传递中起主要作用，并关键性地参与作为学习、记忆和其他高级认知过程基础的复杂过程。在上一个资助期内进行的先前研究确定，酒精对NMDA受体的抑制可以通过受体上的离散结构域进行调节，并确定了蛋白质跨膜结构域内可能定义酒精作用位点的关键残基。在这个应用中，我们提出了一系列的具体目标，旨在建立这些领域如何控制受体对酒精的敏感性，以及如何在体内表达乙醇不敏感受体影响酒精诱导的行为。目的1将检验NMDA受体亚结构域内的特定氨基酸决定乙醇抑制受体功能的能力的假设。在这个目标的实验将使用重组表达和记录技术来测试如何定点突变的关键跨膜结构域中的残基改变酒精对通道功能的影响。目的2将测试的假设，酒精抑制NMDA受体也修改了新的NR 3亚基的表达，并通过磷酸化的NR 1和NR 2亚基的胞内结构域上的关键残基。这方面的实验将利用表达和记录技术，再加上使用转基因动物。目的3将测试这一假设，即对酒精的行为反应可以通过表现出改变的乙醇敏感性的突变型NMDA受体的表达来改变。这一目标的实验将使用目前正在开发的新型NMDA基因敲入小鼠和病毒过表达技术来改变离散脑区中NMDA亚基的表达。总的来说，这些研究的结果有望导致对NMDA受体在介导酒精对大脑作用中的作用的更完整的理解。 公共卫生相关性：影响个体对酒精致醉作用的敏感性的过程是未来酒精问题的重要预测因素。本提案中进行的研究将确定影响大脑离子通道酒精敏感性的因素，该通道在调节大脑活动中发挥着关键作用。

DESCRIPTION (provided by applicant): Understanding the effects of alcohol on brain function is critical for developing better treatments for alcohol-related problems. In this renewal of an ongoing research project, we focus on understanding how alcohol affects the function of a key ion channel expressed by neurons. This channel, the N- methyl-D-aspartate (NMDA) receptor, plays a major role in excitatory glutamatergic transmission and is critically involved in complex processes that underlie learning, memory and other higher cognitive processes. Previous studies carried out during the previous funding period established that alcohol's inhibition of the NMDA receptor could be modulated by discrete domains on the receptor and identified key residues within transmembrane domains of the protein that may define an alcohol site of action. In this application, we propose a series of specific aims designed to establish how these domains control the receptor's sensitivity to alcohol and how expression of ethanol-insensitive receptors in vivo affects alcohol-induced behaviors. Aim 1 will test the hypothesis that specific amino acids within sub-domains of the NMDA receptor determine the ability of ethanol to inhibit receptor function. Experiments in this aim will use recombinant expression and recording techniques to test how site-directed mutagenesis of residues in key transmembrane domains alters the effects of alcohol on channel function. Aim 2 will test the hypothesis that alcohol inhibition of NMDA receptors is also modified by expression of the novel NR3 subunit and through phosphorylation of key residues on the intracellular domain of the NR1 and NR2 subunits. Experiments in this aim will utilize expression and recording techniques coupled with the use of genetically modified animals. Aim 3 will test the hypothesis that the behavioral responses to alcohol can be modified by expression of mutant NMDA receptors that show altered ethanol sensitivity. Experiments in this aim will use a novel NMDA knock-in mouse that is currently under development and viral over-expression techniques to alter NMDA subunit expression in discrete brain areas. Overall, results from these studies are expected to lead to a more complete understanding of the role of NMDA receptors in mediating alcohol's action of the brain. PUBLIC HEALTH RELEVANCE: Processes that affect an individual's sensitivity to the intoxicating effects of alcohol are an important predictor of future alcohol problems. Research to be carried out in this proposal will determine the factors that influence the alcohol sensitivity of a brain ion channel that is critically involved in regulating brain activity.