Structural Determinants of GABA-A receptor function

描述（申请人提供）：γ -氨基丁酸（GABA）是哺乳动物中枢神经系统中主要的抑制性神经递质，大脑中最快速的抑制性神经传递是由GABAA受体（gabar）介导的。GABAR亚基有许多家族，具有多种亚型。α亚基家族是最大的，有六个不同的亚型（alpha1-alpha6）。这些亚型的表达受区域和发育的调控，很可能每个α亚型都发挥着独特的生理作用。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的许多特性，但对α亚型的身份对通道动力学特性的影响知之甚少。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。本研究将通过比较含有不同α亚型的重组受体在单通道、大补丁和全细胞水平上的特性来研究这个问题。这些结果的结合将允许完整描述与每个α亚型相关的动力学特性，并可能预测含有这些亚型的突触受体的行为。对于α亚基家族中功能最多样化的α 1和α 6亚型，我们将详细研究导致不同动力学性质的结构差异。为了分离负责与每个亚型相关的功能特征的一般结构域，将检查六个嵌合阿尔法1 -阿尔法6亚基。这些嵌合体被设计用来分离四个跨膜结构域的功能贡献。一旦一般结构域被分离，负责这些特征的特定残基将通过位点定向诱变被识别。gabar在调节神经元活动中起着不可或缺的作用。gabar是常用的镇静药、抗焦虑药和抗癫痫药以及成瘾药物（包括巴比妥酸盐和酒精）的重要靶点。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α亚型之间的结构差异如何影响通道的功能特性的问题。这项工作的结果将有助于我们了解天然gabar的α亚基组成的变化如何影响神经元的性质及其对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.