ETHANOL AND PHOSPHORYLATION OF THE 5-HT3 RECEPTOR

5-羟色胺3（5-HT 3）受体的功能通过以下方式增强： 醇类，但作用机制尚不清楚。 此外，本发明还提供了一种方法， 通过翻译后修饰事件调节5-HT 3受体， 例如磷酸化，还没有得到很好的表征。 迄今为止，一 已经克隆了5-HT 3受体的亚基，并且它具有多个 蛋白激酶依赖性磷酸化的共有序列位点。 以前的工作与其他配体门控离子通道，即GABA A和 NMDA受体，表明蛋白激酶C可能参与了 介导这些受体对乙醇的反应。 因此有 合理推测激酶可能调节 5-HT 3受体对醇类的反应。 在本提案中， 5-HT 3受体的研究将使用非洲爪蟾卵母细胞作为 表达系统 双电极电压钳电生理 将记录表达的野生型或诱变的5-HT 3受体， 进行了 首先，5-HT 3受体对酒精的敏感性将被降低。 测定了 接下来，研究丝氨酸/苏氨酸激酶改变5-HT 3的能力。 将讨论受体功能。 确定哪些氨基酸是 这些激酶的底物，候选丝氨酸的共有序列， 受体中的蛋白激酶依赖性磷酸化将突变为 非磷酸化残基。 改变5-HT 3受体功能的激酶 将检查他们改变这种敏感性的能力， 酒精受体 另一种假设是， 活性是5-HT 3受体对酒精敏感性的基础， 通过测量5-HT 3受体的酒精调节进行测试， 磷酸化的共有序列位点。 因此， 该计划旨在确定磷酸化事件是否改变了 5-HT 3受体及其由醇类调节的研究进展 更好的 了解5-HT 3受体的调节是重要的， 这种受体可能参与介导的奖励行动， 滥用药物，如乙醇。

The function of the 5-hydroxytryptamine3 (5-HT3) receptor is enhanced by alcohols, but the mechanism(s) of action remains unknown. In addition, regulation of the 5-HT3 receptor by post-translational modification events, such as phosphorylation, has not been well characterized. To date, one subunit of the 5-HT3 receptor has been cloned, and it possesses multiple consensus sequence sites for protein kinase dependent phosphorylaiton. Previous work with other ligand-gated ion channels, namely the GABA A and NMDA receptors, has suggested that protein kinase C may be involved in mediating these receptors' responsiveness to ethanol. Therefore, it is reasonable to speculate that kinases may modulate the responsiveness of the 5-HT 3 receptor to alcohols. In the present proposal, functional studies of the 5-HT3 receptor will be performed using Xenopus laevis oocytes as an expression system. Two-electrode voltage clamp electrophysiological recordings of expressed wild-type or mutagenized 5-HT3 receptors will be made. First, the sensitivity of the 5-HT3 receptor to alcohols will be measured. Next, the ability of serine/threonine kinases to alter 5-HT3 receptor function will be addressed. To determine which amino acid(s) are substrates for these kinases, candidate serines in consensus sequences for protein kinase dependent phosphorylation in the receptor will be mutated to nonphosphorylatable residues. Kinases which alter 5-HT3 receptor function will be examined for their ability to change the sensitivity of this receptor to alcohols. The alternative hypothesis, that tonic kinase activity underlies the 5-HT3 receptor's sensitivity to alcohols, will be tested by measuring alcohol modulation of 5-HT3 receptors with mutagenized consensus sequence sites for phosphorylation. Thus, the ultimate goal of the proposal is to determine if phosphorylation events alter the function of the 5-HT3 receptor and its modulation by alcohols. A better understanding of the regulation of the 5-HT3 receptor is important, given that this receptor may be involved in mediating the rewarding actions of drugs of abuse such as ethanol.