GENERAL ANESTHETIC SITE(S) OF ACTION ON GABA A RECEPTOR

γ-氨基丁酸（GABA）是主要的抑制性神经递质 这种神经递质的一种亚型 受体，GABAA受体是一个可能的药理学目标， 全身麻醉药很明显，全身麻醉剂作用于GABAA 受体以高度特异性的方式。然而， GABAA受体多肽内的全身麻醉作用，或 “行动地点”不详。 确定全身麻醉药的作用部位只能通过 全身麻醉药与 表达的GABAA受体由已知的亚基组成， 氨基酸序列是已知的。GABAA受体仅由 在非洲爪蟾卵母细胞中表达的β 3或rho 1亚基完全显示出 对GABAA和全身麻醉剂的电生理反应相反， 因此，是分子解剖的极好模型系统， 行动地点全身麻醉剂诱导电流，但GABA没有 对β 3亚单位受体的影响。相反，rho 1亚基 受体对GABA有反应，但对全身麻醉药无反应。的目标 本研究旨在验证全身麻醉药相互作用的假设 与GABAA受体以结构域特异性方式在不同于 GABA识别位点。 这将通过 对重组天然的、突变的或 嵌合GABAA受体在非洲爪蟾卵母细胞中表达。epitope-tag 免疫组化和Western印迹技术 分析，将用于区分非功能性和非表达性 受体。通过对氨基酸相似性的系统研究， 不同亚基之间的差异， 多肽所必需的直接开放通道的一般 麻醉剂，以及增强GABA诱导的 电流将被定义。接下来，这些的生理相关性 将在GABAA受体亚单位中检测已确定的作用位点 最有可能存在于大脑中的组合。 了解全身麻醉剂的工作场所对未来至关重要 开发无副作用的全身麻醉药， 目前导致临床发病率和死亡率。此外，委员会认为， 一旦确定了作用地点。位点特异性逆转剂可 可以开发。这种药理学上的补充 临床麻醉医师可用的设备是巨大的， 重要性，也许只是次要的发现一般 麻醉剂本身。

Gamma-amino-butyric-acid (GABA) is the major inhibitory neurotransmitter in the vertebrate brain, and one subtype of this neurotransmitter receptor, the GABAA receptor is a likely pharmacological target for general anesthetics. It is clear that general anesthetics act on the GABAA receptor in a highly specific manner. However the precise loci of the general anesthetic action within the GABAA receptor polypeptide, or the "sites-of-action", is unknown. Defining the sites-of-action of general anesthetics is only possible by a systematic study of the interactions between general anesthetics and expressed GABAA receptors composed of known subunits where the precise amino acid sequences are known. GABAA receptors consisting of only the beta3 or the rho1 subunits expressed in Xenopus oocytes show diametrically opposite electrophysiological responses to GABAA and general anesthetics, and therefore, are excellent model systems for the molecular dissection of the sites-of-action. General anesthetics induce currents, but GABA has no effect on the beta3 subunit receptors. Conversely, the rho1 subunit receptors respond to GABA, but not to general anesthetics. The goal of this study is to test the hypothesis that general anesthetics interact with the GABAA receptor in a domain specific manner at sites distinct from the GABA recognition site. This will be accomplished by electrophysiological experiments on recombinant native, mutated, or chimeric GABAA receptors expressed in Xenopus oocytes. The epitope-tag technique, in combination with immunohistochemistry and Western blot analysis, will be used to distinguish non-functional from non-expressed receptors. Through a systematic study of the amino acid similarities and differences between different subunits, the sites on the receptor polypeptide necessary for the direct opening of channels by general anesthetics, and the sites necessary for the potentiation of GABA induced currents, will be defined. Next, the physiological relevance of these identified sites-of-action will be tested in GABAA receptor subunit combinations most likely to be present in the brain. Understanding where general anesthetics work is essential for future development of general anesthetics without the side-effects which presently contribute to clinical morbidity and mortality. Furthermore, once the sites-of-action are defined. site specific reversal agents could possibly be developed. Such an addition to the pharmacological armamentarium available to clinical anesthesiologists is of immense significance, perhaps only secondary to the discovery of general anesthetics itself.