Mechanisms of Neurotransmitter-gated Ion Channels

这项提议的重点是肌肉烟碱型乙酰胆碱受体(AChR)，这是一种神经递质门控 在脊椎动物神经肌肉接头(NMJ)介导快速突触传递的离子通道。这个 这一建议的广泛目标是了解这种受体通道的结构如何引起其 功能，以及这种功能是如何精致地为脊椎动物NMJ的生理需求量身定做的。 多年来，对AChR的研究已经回答了关于 离子通道的物理、化学和生物学，这些离子通道不能用(可能)任何其他方法来解决 频道。我们的主要工具是单通道和宏观电生理学(使用体外和 原位方法)、蛋白质工程和量化思维。我们的四个具体目标是：i) 确定ACHR大门(S)的位置和物理性质，以及在打开和关闭时的重新排列 关闭通道，ii)了解AChR的结构和功能之间的关系 选择性滤光片，以及离子占有率和门控/减敏之间的耦合，iii)估计 AChR的非连接门控平衡常数的值，以及IV)以定义 对AChR的突触反应脱敏。值得注意的是，除了明显的 在神经元到肌肉信号的传递中，ACh、AChR和NMJ一直是 神经递质、受体和突触。因此，从拟议的 实验将对我们对突触后受体的基本理解产生广泛的影响 快速的突触传输。还应该强调的是，AChR介导的神经传递支持 不仅是运动，还有自主神经和认知功能，因此，它不仅在 重症肌无力，但也有心血管疾病和神经退行性疾病，如阿尔茨海默氏症 疾病。

The focus of this proposal is the muscle nicotinic acetylcholine receptor (AChR), the neurotransmitter-gated ion channel that mediates fast synaptic transmission at the vertebrate neuromuscular junction (NMJ). The broad objective of this proposal is to understand how the structure of this receptor-channel gives rise to its function, and how this function is exquisitely tailored to the physiological needs of the the vertebrate NMJ. Throughout the years, research on the AChR has provided answers to fundamental questions on the physics, chemistry, and biology of ion channels that cannot be addressed with (probably) any other channel. Our main tools are single-channel and macroscopic electrophysiology (using both in-vitro and in-situ approaches), protein engineering, and quantitatitive thinking. Our four specific aims are: I) To determine the location and physical nature of the AChR's gate(s), and its rearrangement upon opening and shutting of the channel, II) To understand the relationship between structure and function in the AChR's selectivity filter, and the coupling between ion-occupancy and gating/desensitization, III) To estimate the value of the AChR's unliganded-gating equilibrium constant, and IV) To define the contribution of desensitization to the synaptic response of the AChR . It is worth noting that, in addition to their obvious relevance in the relay of neuron-to-muscle signals, ACh, the AChR, and the NMJ have served as models of neurotransmitters, receptors, and synapses in general. Hence, the knowledge derived from the proposed experiments is poised to have a broad impact on our basic understanding of postsynaptic receptors and fast synaptic transmission. It should also be emphasized that AChR-mediated neurotransmission supports not only motor but, also, autonomic and cognitive function and, therefore, that it is compromised not only in myasthenias but, also, in cardiovascular diseases and neurodegenerative disorders such as Alzheimer's disease.