GABAA RECEPTOR BETA-SUBUNIT AND GENERAL ANESTHETICS

A型γ-氨基丁酸（GABAA）受体是一种 是全身麻醉药物重要药理学靶点。最新进展 提示几种全身麻醉药在GABAA中的高度特异性作用 受体;然而，这种机制的细节是未知的。另外 构成GABAA的亚基有许多不同的形式 受体的GABAA受体亚单位组成与 全身麻醉药理学仍然相对未开发， 这种相互作用对神经元功能的重要性是未知的。这 该项目检验了GABAA受体β亚单位亚型 指示依托咪酯调节神经元中GABA诱导的电流。 该研究由一系列精心设计的步骤组成，从生物物理学开始 GABAA时静脉全身麻醉药作用的表征 受体，然后检查b亚单位的作用 基因靶向对全身麻醉药理学的影响，并总结了 产生具有可逆的外部诱导改变的神经元， 对依托咪酯敏感。特别是最近发现的 β-亚基和特定的氨基酸在位置270上， 将通过在HEK 293细胞中表达不同的 β-亚单位同种型和点突变体在A1 BXG 2异聚体组合中。 全细胞膜片钳和快速GABA灌注将用于达到一个 药物作用动力学模型接下来，这个机械模型将被用作 研究β亚基基因靶向对一般 视黄酸诱导的P19神经元的麻醉药理学。P19系统 为研究基因靶向的效果提供了前所未有的机会 而不需要复杂的创造一个完整的动物。常规 b1转基因，b1敲除和条件基因靶向神经元将被 考察在研究项目的下一阶段，最好的基因靶向 在这些研究中发现的策略将用于创建基因靶向小鼠 具有可逆的外部诱导的对一般 麻醉剂 详细了解现有的全身麻醉剂是如何工作的是必不可少的 用于开发改进的麻醉剂， 现有代理商的影响。拟议的实验将增加定量和 全身麻醉药分子药理学和运动机制信息 这一领域的调查更接近真正了解这些如何 临床基本药物起作用。

The type A gamma amino butyric acid (GABAA) receptor is an important pharmacological target for general anesthetic drugs. Recent progress suggests highly specific effects of several general anesthetics at the GABAA receptor; the details of this mechanism, however, are unknown. Additionally there are many different forms of the subunits which make up the GABAA receptor. The interaction between GABAA receptor subunit composition and general anesthetic pharmacology remains relatively unexplored, and the significance of this interaction to the function of neurons is unknown. This project tests the hypothesis that the GABAA receptor beta-subunit isoform dictates etomidate modulation of GABA-induced current in neurons. The study consists of a series of deliberate steps beginning with a biophysical characterization of the effects of intravenous general anesthetics at GABAA receptors, followed by an examination of the effects of b-subunit gene-targeting on general anesthetic pharmacology and concluding with the creating of neurons with a reversible externally-inducible alteration in sensitivity to etomidate. Specifically, the recently discovered critical role of the beta-subunit and the specific amino acid at location 270 on the beta-subunit will be explored through expression, in HEK293 cells, of different beta-subunit isoforms and point-mutants in an a1bxg2 heteromeric combination. Whole cell patch clamp and rapid GABA perfusion will be used to arrive at a kinetic model of drug action. Next this mechanistic model will be used as a tool to investigate the effects of beta-subunit gene-targeting on general anesthetic pharmacology in retinoic-acid-induced P19 neurons. The P19 system offers an unprecedented opportunity to investigate the effect of gene-targeting without the complexities involved in creating a whole animal. Conventional b1-transgenic, b1-knockout, and conditional gene-targeted neurons will be examined. In the next phase of the research project, the best gene-targeting strategy found in these studies will be used to create a gene-targeted mouse with a reversible externally-inducible alteration in sensitivity to general anesthetics. Detailed understanding of how existing general anesthetics work is essential for the development of improved anesthetics without the very significant side effects of existing agents. The experiments proposed will add quantitative and mechanistic information to general anesthetic molecular pharmacology and move this field of investigation closer towards truly understanding how these clinically essential drugs work.