Identification of structures within GABAA receptor α subunits that regulate the agonist action of pentobarbital
Identification of structures within GABAA receptor α subunits that regulate the agonist action of pentobarbital
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DOI：
10.1124/jpet.106.104844
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发表时间：
2006-09-01
影响因子：
3.5
通讯作者：
Fisher, Janet L.
中科院分区：
文献类型：
--
作者：
Drafts, Brandon C.;Fisher, Janet L.
sBarbiturates通过三种不同的机制作用于GABA - A受体（gabar），导致正变构调节、直接激活和抑制。在不同浓度下观察到这些效应，并且受某些突变和受体亚基组成的不同影响。哺乳动物gabar可以由16种不同亚基亚型的组合形成。虽然巴比妥类药物的作用主要取决于β亚基，但它们的激动剂活性在很大程度上受α亚基亚型的影响。戊巴比妥仅在受体含有α 6亚基时才比GABA更有效。嵌合α 1/ α 6亚基的结果表明，细胞外n端结构域的结构差异是造成这种特征的原因。在这个区域内，我们检查了α 6亚型特有的15个氨基酸残基。这些位点中的每个都在α 6亚基中单独突变为α 1亚基的相应残基。突变对戊巴比妥直接激活的影响通过全细胞电生理记录来确定。我们的研究结果表明，这些突变中只有α 6（T69K）一种会改变戊巴比妥的疗效。这种单一突变将戊巴比妥的反应降低到野生型α 1 β 1 γ 2L和α 6 β 1 γ 2L亚型的中间水平。这种突变不影响受体对GABA的敏感性，但确实降低了另一种与戊巴比妥活性相似的静脉麻醉剂依托咪酯的疗效。α 1亚基（K70T）的反向突变没有改变对戊巴比妥的反应。这是首次鉴定出调节巴比妥类药物直接激活的GABAR α亚型的结构差异。
sBarbiturates act on GABA A receptors (GABARs) through three distinct mechanisms, resulting in positive allosteric modulation, direct activation, and inhibition. These effects are observed at different concentrations and are differentially affected by some mutations and by the receptor's subunit composition. Mammalian GABARs can be formed from a combination of 16 different subunit subtypes. Although the effect of barbiturates depends largely on the beta subunit, their agonist activity is substantially influenced by the alpha subunit subtype. Pentobarbital is a more effective agonist than GABA only when receptors contain an alpha 6 subunit. Results from chimeric alpha 1/alpha 6 subunits suggested that structural differences within the extracellular N-terminal domain were responsible for this characteristic. Within this domain, we examined 15 amino acid residues unique to the alpha 6 subtype. Each of these sites was individually mutated in the alpha 6 subunit to the corresponding residue of the alpha 1 subunit. The effect of the mutation on direct activation by pentobarbital was determined with whole-cell electrophysiological recordings. Our results indicate that only one of these mutations, alpha 6( T69K), altered pentobarbital efficacy. This single mutation reduced the response to pentobarbital to a level intermediate to the wild-type alpha 1 beta 1 gamma 2L and alpha 6 beta 1 gamma 2L isoforms. The mutation did not affect the sensitivity of the receptor to GABA but did reduce the efficacy of etomidate, another i.v. anesthetic with activity similar to pentobarbital. The reverse mutation in the alpha 1 subunit (K70T) did not alter the response to pentobarbital. This is the first identification of a structural difference in GABAR alpha subtypes that regulates direct activation by barbiturates.