MOLECULAR BIOLOGY OF ETHANOL ACTION ON RECEPTORS

这一研究科学家奖是基于目前正在进行的工作 了解乙醇专一性作用的分子基础 配基门控离子通道。这一提议检验了这样一种假设，即 乙醇的选择性影响一些人对NMDA和GABA的反应， 但不是所有的，神经元是由于特定的NMDA和GABAA的存在 受体亚型。提出用模型方法来研究生物多样性的作用 乙醇对NMDA和GABAA受体的作用是对其研究进展的更新 R01S AA09122和AA10025，定义分子机制的授权(S) 负责乙醇对特定受体亚型的专一性。 关于乙醇对GABAA受体亚型的作用，我们有 证实唑吡坦预测一种GABAA受体亚型对 乙醇。同样，我们已经证明了 芬地尔和乙醇对NMDA反应的拮抗作用。在这份提案中， 每个培养和分离的神经细胞都将在药理学上 以膜片钳电生理学为特征，细胞质将 以及GABAA和NMDA受体亚基的mRNAs 每个单个神经元内的神经元将通过RT/PCR进行鉴定。在……里面 具体目标一，工作最初将集中在假设 α1β2γ2亚单位组合是一种GABAA受体亚型 对乙醇和唑吡坦敏感。由于目前的不确定性， 特别强调的是神经元中哪些Gamma2变体 与这个亚基结合的人对乙醇敏感。一秒钟 研究GABAA受体的部分工作与定义分子有关 乙醇的作用依赖于次级神经的机制 对单元格的输入。最后，我们将确定安定不敏感 含有α4或α6亚基的受体对 乙醇。SPICAL AIM II将鉴定NMDA受体的mRNAs 单个神经元内对乙醇拮抗敏感的亚单位 NMDA。目前的结果表明，部分但不是全部的NMDA受体 对异丙苯地尔敏感的亚型对乙醇敏感。自.以来 依芬地尔与据称含有NMDAR-2b的NMDA受体结合 亚基，可以假设受体亚型对乙醇敏感 将包含这个NMDA受体亚单位，以及适当的 NMDAR-1变种。目前正在进行研究，以证实这一假设。这个 提出模型来定义乙醇作用的分子基础 当应用于所有配体时，影响GABAA和NMDA受体亚型- 门控离子通道，将提供一种识别神经解剖学的手段 酒精最有可能影响大脑的部位 功能。此外，基于对乙醇作用的结构分析 关于受体的蛋白质-蛋白质相互作用，结果可能 最终使我们能够预测其他蛋白质之间的相互作用 受乙醇的影响。

This Research Scientist Award is based upon current work being performed to understand the molecular basis of ethanol's specificity of action on ligand-gated ion channels. This proposal, tests the hypothesis that the selectivity of ethanol to affect responses to NMDA and GABA from some, but not all, neurons is due to the presence of specific NMDA and GABAA receptor subtypes. The model approach proposed to study the action of ethanol on NMDA and GABAA receptors is an update of progress made on R01s AA09122 and AA10025, grants defining the molecular mechanism(s) responsible for ethanol's specificity on specific receptor subtypes. In respect to ethanol's action on GABAA receptor subtypes, we have demonstrated zolpidem predicts one GABAA receptor subtype sensitive to ethanol. Likewise, we have shown a close relationship between ifenprodil and ethanol antagonism of NMDA responses. In this proposal, each cultured and dissociated neurons cell will be pharmacologically characterized with patch-clamp electrophysiology, the cytoplasm will be extracted from the cell, and mRNAs for GABAA and NMDA receptor subunits within each individual neuron will be identified with RT/PCR. In Specific Aim I, work will initially focus on the hypothesis that the alpha1beta2gamma2 subunit combination is one GABAA receptor subtype sensitive to ethanol and zolpidem. Due to present uncertainty, particular emphasis is being placed on which gamma2 variants in neurons with this subunit combination are sensitive to ethanol. A second portion of work on GABAA receptors relates to defining the molecular mechanism by which ethanol's action is dependent upon secondary neural inputs to a cell. Finally, we will determine if diazepam-insensitive receptors, containing alpha4 or alpha6 subunits, are sensitive to ethanol. Specific Aim II will identify the mRNAs for NMDA receptor subunits within individual neurons sensitive to ethanol antagonism of NMDA. Current results suggest that some, but not all, NMDA receptor subtypes sensitive to ifenprodil are sensitive to ethanol. Since ifenprodil binds to NMDA receptors purportedly containing the NMDAR-2b subunit, it can assumed that the receptor subtype sensitive to ethanol will contain this NMDA receptor subunit, as well as an appropriate NMDAR-1 variant. Studies are underway to confirm this hypothesis. The model proposed to define the molecular basis of ethanol's action to affect GABAA and NMDA receptor subtypes, when applied to all ligand- gated ion channels, will provide a means to identify neuroanatomical sites at which ethanol has the greatest probability of affecting brain function. Further, based upon structural analysis of ethanol's action on protein-protein inter-actions for the receptors, results may ultimately allow us to predict other protein-protein interactions influenced by ethanol.