Alcohol Actions-Molecular Targets on Brain Proteins

描述（由申请人提供）：乙醇的急性作用是通过与蛋白质上的特定位点结合而介导的。我们建议结合分子、电生理和行为等方法来阐明乙醇对GABAa和甘氨酸受体的作用。目的1将阐明异质GABAa受体中形成乙醇结合位点的氨基酸的具体位置和取向。我们正在进行的甘氨酸受体的研究表明，锌是甘氨酸受体的生理调节剂，对于乙醇对这些受体的作用是重要的，Aim 1也将确定这种相互作用的分子基础。在Aim 2中，我们将使用GABAa和甘氨酸受体敲除和敲入小鼠来指定乙醇对这些受体的作用所产生的行为影响。目的3将评估受体突变对神经元的影响。这将通过对大脑关键区域的基因组分析和对中脑边缘奖赏通路的电生理研究来完成。Aim 3还将确定慢性乙醇消耗对腹侧被盖区和伏隔核（中脑边缘通路的两个关键区域）整体基因表达变化的影响。这些研究将使用野生型小鼠和具有GABAa或甘氨酸受体的小鼠，这些受体被改造成对乙醇作用不敏感。这种新方法将使我们能够将基因表达（和行为，目标2）的变化与酒精对特定受体的作用联系起来。这项工作的长期目标是确定关键的蛋白质位点，这些蛋白位点可以作为缓解酒精强化、依赖和复发的新疗法的靶点。尽管酒精（乙醇）已经被饮用了数千年，但我们对它对大脑产生影响的方式知之甚少。一个重要的进展是识别特定的蛋白质（神经递质受体和离子通道）参与神经元之间的通信作为乙醇的目标。我们将使用不同的技术，从分子水平到行为水平，利用小鼠突变和其他新技术，确定乙醇如何作用于这些蛋白质，最终目标是确定可以作为新疗法靶点的关键蛋白质位点，以减轻酒精成瘾。

DESCRIPTION (provided by applicant): The acute effects of ethanol are mediated by binding to specific sites on proteins. We propose to elucidate the actions of ethanol on the GABAa and glycine receptors, two critical molecular targets for ethanol, by combining molecular, electrophysiological and behavioral methods. Aim 1 will elucidate the specific location and orientation of the amino acids that form the ethanol binding site in heteromeric GABAa receptors. Our ongoing studies of glycine receptors suggest that zinc, a physiological modulator of glycine receptors, is important for ethanol action on these receptors, and Aim 1 will also define the molecular basis of this interaction. In Aim 2, we will use GABAa and glycine receptor knockout and knockin mice to specify the behavioral effects of ethanol that are due to actions on these receptors. Aim 3 will evaluate the neuronal consequences of receptor mutation. This will be accomplished by genomic analysis of key brain regions and by electrophysiological study of the mesolimbic reward pathway. Aim 3 will also define effects of chronic ethanol consumption on global gene expression changes in the ventral tegmental area and nucleus accumbens, the two key areas of the mesolimbic pathway. These studies will use wild type mice and mice with GABAa or glycine receptors that are engineered to be insensitive to ethanol action. This novel approach will allow us to link changes in gene expression (and behavior, Aim 2) to alcohol actions on specific receptors. The long-range goal of this work is to define key protein sites that can serve as targets for new therapies to alleviate alcohol reinforcement, dependence an relapse. PUBLIC HEALTH RELEVANCE Even though alcohol (ethanol) has been consumed for thousands of years, we know remarkably little about the way it produces its effects on the brain. An important advance was the identification of specific proteins (neurotransmitter receptors and ion channels) involved in communication between neurons as a target for ethanol. We will define how ethanol acts on these proteins using different techniques, ranging from the molecular to the behavioral level, using mutations in mice and other new technologies, with the final objective of defining key protein sites that can serve as targets for new therapies to alleviate alcohol addiction.