Mechanisms of Neurotransmitter-gated Ion Channels

描述（由申请人提供）：该提案的重点是肌肉烟碱乙酰胆碱受体（AChR），即在脊椎动物神经肌肉接头（NMJ）介导快速突触传递的神经递质门控离子通道。这个建议的主要目的是了解这种受体通道的结构如何产生其功能，以及这种功能如何根据脊椎动物NMJ的生理需求进行精心定制。多年来，对AChR的研究为离子通道的物理、化学和生物学的基本问题提供了答案，这些问题（可能）无法用任何其他通道来解决。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），蛋白质工程和定量思维。我们的四个具体目标是：I）确定AChR门的位置和物理性质，以及在通道打开和关闭时的重排，II）理解AChR选择性过滤器中结构和功能之间的关系，以及离子占据和门控/脱敏之间的耦合，III）估计野生型AChR的未配体门控平衡常数的值，和IV）确定脱敏对AChR的突触反应的贡献。值得注意的是，除了它们在神经元到肌肉信号的传递中的明显相关性之外，ACh、AChR和NMJ通常还充当神经递质、受体和突触的模型。因此，从拟议的实验中获得的知识有望对我们对突触后受体和快速突触传递的基本理解产生广泛的影响。还应该强调的是，乙酰胆碱受体介导的神经传递不仅支持运动，而且还支持自主神经和认知功能，因此，它不仅在肌无力症中受到损害，而且在心血管疾病和神经退行性疾病如阿尔茨海默病中也受到损害。此外，AChR还在气道、角质形成细胞、淋巴细胞和内皮细胞中表达，它们是细胞分化、增殖和抗细胞凋亡以及癌症的介导剂。事实上，越来越清楚的是，尼古丁成瘾（其由尼古丁与大脑的行为控制区域中的AChR的相互作用引起）增加了患肺癌和口腔癌的风险，这是由于吸烟时吸入的尼古丁和尼古丁衍生的亚硝胺过度激活这些非肌肉/非神经元AChR。

DESCRIPTION (provided by applicant): 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 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 quantitative 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 wild-type 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 disease. Furthermore, AChRs are also expressed in the airways, keratinocytes, lymphocytes, and endothelial cells where they are mediators of cell differentiation, proliferation, and resistance to apoptosis and, thus, of cancer. Indeed, it is becoming increasingly clear that nicotine addiction (which is caused by the interaction of nicotine with AChRs in behavior- controlling areas of the brain) increases the risk of developing lung and oral cancer owing to the excessive activation of these non-muscle/non-neuronal AChRs by inhaled nicotine and nicotine-derived nitrosamines while smoking.