Structural Determinants of GABA-A receptor function

描述（由申请人提供）：γ-氨基丁酸（GABA）是哺乳动物中枢神经系统中的主要抑制性神经递质，大脑中最快的抑制性神经传递由GABAA受体（GABAR）介导。GABAR亚基有许多家族，具有多种亚型。α亚基家族是最大的，有六种不同的亚型（α 1-α 6）。亚型的表达受区域性和发育性调节，并且很可能每个α亚型执行独特的生理作用。亚型表达也会因病理状况如癫痫而改变。受体的亚基组成影响GABAR的许多特征，但关于α亚型的身份对通道动力学性质的影响知之甚少。这项工作的长期目标是了解GABAR结构异质性的功能意义，并确定亚基之间的结构差异是其不同功能特性的基础。拟议的工作将检查这个问题，通过比较含有不同的α亚型的重组受体在单通道，macropatch和全细胞水平的属性。这些结果的组合将允许与每个α亚型相关的动力学特性的完整描述，并可以预测包含这些亚型的突触受体的行为。负责不同的动力学特性的结构差异将详细检查的α 1和α 6亚型，这是最功能多样的α亚基家族。为了分离负责与每个亚型相关的功能特征的一般结构域，将检查六个嵌合α 1-α 6亚基。设计这些嵌合体以分离由四个跨膜结构域中的每一个所做出的功能贡献。一旦一般结构域被分离，负责这些特征的特定残基将通过定点诱变来鉴定。GABAR在调节神经元活动中起着不可或缺的作用。GABAR是通常用作镇静剂、抗焦虑剂和抗癫痫剂的药物以及成瘾药物（包括巴比妥酸盐和酒精）的重要靶标。许多这些调节剂的活性取决于受体的α亚型组成。所描述的项目将解决的问题，如何在α亚型之间的结构差异有助于通道的功能特性。这项工作的结果将有助于我们了解天然GABARs的α亚基组成的变化如何影响神经元的特性及其对GABA的反应。

DESCRIPTION (provided by applicant): Gamma-aminobutyric acid (GABA) is the primary inhibitory neurotransmitter in the mammalian central nervous system, and most fast inhibitory neurotransmission in the brain is mediated by the GABAA receptors (GABARs). There are many families of GABAR subunits with multiple subtypes. The alpha subunit family is the largest, with six different subtypes (alpha1-alpha6). The expression of the subtypes is regulated regionally and developmentally and it is likely that each alpha subtype performs a unique physiological role. Subtype expression is also altered by pathological conditions, such as epilepsy. The subunit composition of the receptor affects many characteristics of the GABAR, but little is known about the effect of the identity of the alpha subtype on the channel's kinetic properties. The long-term goal of this work is to understand the functional significance of the structural heterogeneity of the GABAR and to determine which structural differences among the subunits underlie their different functional properties. The proposed work will examine this question by comparing the properties of recombinant receptors containing different alpha subtypes at the single channel, macropatch and whole-cell levels. The combination of these results will allow a complete description of the kinetic properties associated with each alpha subtype and may predict the behavior of synaptic receptors containing these subtypes. The structural differences responsible for distinct kinetic properties will be examined in detail for the alpha1and alpha 6 subtypes, which are the most functionally diverse in the alpha subunit family. To isolate the general structural domains responsible for the functional characteristics associated with each subtype, six chimeric alpha1 - alpha6 subunits will be examined. These chimeras were designed to isolate the functional contribution made from each of the four transmembrane domains. Once the general structural domains are isolated, the specific residues responsible for these characteristics will be identified through site-directed mutagenesis. The GABARs play an integral role in regulating neuronal activity. GABARs are important targets for drugs commonly used as sedatives, anxiolytics and anti-epileptics as well as for drugs of addiction, including barbiturates and alcohol. The activity of many of these modulators depends upon the alpha subtype composition of the receptor. The projects described will address the question of how structural differences among the alpha subtypes contribute to the functional properties of the channel. The results of this work will help us to understand how variations in the alpha subunit composition of native GABARs can affect the properties of neurons and their responses to GABA.