Alcohol actions on NMDA receptor gating domains

描述(申请人提供)：大脑中的N-甲基-D-天冬氨酸(NMDA)谷氨酸受体离子通道是酒精行为效应的重要介体。虽然已知酒精通过影响离子通道门控来抑制NMDA受体，但其分子作用部位和这种作用的机制尚未确定。以前的研究已经指出，神经递质门控离子通道的跨膜结构域在酒精的作用中发挥了作用。本实验室的结果表明，NMDA受体NR2A亚单位第四膜相关(M)结构域中保守的蛋氨酸残基(Met823)深刻影响离子通道门控。本实验室最近获得的证据表明，NR2A(Met823)也以与酒精作用部位一致的方式影响受体的酒精敏感性，相邻残基也可能影响离子通道门控和酒精敏感性。在拟议的研究中要检验的主要假设是，这个残基和邻近的残基也参与离子通道门控，形成酒精作用于NMDA受体-离子通道的关键部位。这些研究将结合全细胞和单通道膜片钳电生理记录来研究下列特定目的：1)NR2A(Met823)邻近结构域在调节NMDA受体离子通道门控和酒精敏感性中的作用；2)这些区域是否构成酒精对NMDA受体作用的关键部位；3)酒精与这些部位相互作用改变NMDA受体离子通道门控的精确动力学机制；以及4)在非神经细胞系中是否也观察到NMDA受体突变对门动力学和酒精调制的影响。这些研究的结果将为酒精调节NMDA受体的分子机制提供重要信息，对NMDA受体-离子通道的结构和功能有新的见解，并更好地理解酒精和类似分子与蛋白质相互作用的一般方式。由于NMDA受体在认知、运动功能和记忆中具有重要和广泛的作用，并且在酒精的主观效应和中枢神经系统抑制效应中都起着重要的作用，这些研究中获得的信息将代表着理解酒精对神经系统影响的重要一步。

DESCRIPTION (provided by applicant): N-methyl-D-aspartate (NMDA) glutamate receptor-ion channels in the brain are important mediators of the behavioral effects of alcohol. Although alcohol is known to inhibit NMDA receptors by influencing ion channel gating, its molecular site of action and the mechanism underlying this effect have not been established. Previous studies have pointed to a role of the membrane-spanning domains of neurotransmitter-gated ion channels in the actions of alcohol. Results from this laboratory have shown that a conserved methionine residue (Met823) in the fourth membrane-associated (M) domain of the NMDA receptor NR2A subunit profoundly influences ion channel gating. Recent evidence obtained in this laboratory has shown that NR2A(Met823) also affects alcohol sensitivity of the receptor in a manner that is consistent with a site of alcohol action, and that adjacent residues may also influence both ion channel gating and alcohol sensitivity. The main hypothesis to be tested in the proposed studies is that this residue and adjacent residues that are also involved in ion channel gating form a key site of alcohol action on the NMDA receptor-ion channel. These studies will incorporate whole-cell and single-channel patch-clamp electrophysiological recording to investigate the following specific aims: 1) the role of domains adjacent to NR2A(Met823) in the regulation of NMDA receptor ion channel gating and alcohol sensitivity; 2) whether these regions constitute a key site of alcohol action on the NMDA receptor; 3) the precise kinetic mechanism by which alcohol interacts with these sites to alter NMDA receptor ion channel gating; and 4) whether effects of NMDA receptor mutations on gating kinetics and alcohol modulation observed in a non-neuronal cell line are also observed in CNS neurons. The results of these studies will yield important information about the molecular mechanism of alcohol modulation of the NMDA receptor, new insights into the structure and function of the NMDA receptor-ion channel, and a better understanding of the ways in which alcohols and similar molecules interact with proteins in general. Because NMDA receptors have vital and widespread roles in cognition, motor function, and memory, and contribute importantly to both the subjective effects of alcohol as well as its CNS depressant effects, the information gained in these studies will represent an important step forward in the understanding of alcohol effects on the nervous system.