Mechanisms of fusion and transmitter release in neurosecretion

神经分泌中的融合和递质释放机制

• 批准号: 7647152
• 负责人: Manfred LINDAU
• 金额: $ 31.8万
• 依托单位: CORNELL UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2000
• 资助国家: 美国
• 起止时间: 2000-04-20 至 2012-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7647152
• 关键词: Affect; Amino Acids; Anions; Bacteria; Behavior; Binding; Brain; C-terminal; Catecholamines; Cations; Cell Size; Cell membrane; Cell physiology; Cells; Charge; Chromaffin Cells; Chromaffin granule; Complex; Disease; Electric Capacitance; Electric Stimulation; Emotional; Endocrine; Event; Exocytosis; Extracellular Space; Health; Hormones; Human body; Immune response; Individual; Invaded; Ions; Lead; Location; Measurement; Measures; Mediating; Membrane; Modeling; Modification; Molecular; Molecular Structure; Mutate; Mutation; Nerve; Neuromodulator; Neurons; Neuropeptides; Neurosecretion; Neurotransmitters; Organelles; Organism; Parasites; Physiological; Physiological Processes; Play; Point Mutation; Positioning Attribute; Process; Property; Proteins; Role; SNAP receptor; Secretory Vesicles; Stimulus; Structure; Synaptic Transmission; Testing; Transmembrane Domain; VAMP-2; Vesicle; Viral; base; carbon fiber; insight; interest; mutant; neurotransmitter release; overexpression; patch clamp; public health relevance; receptor; research study; response; syntaxin; vesicle-associated membrane protein

DESCRIPTION (provided by applicant): The principle long-term objective of this project is to provide a detailed biophysical and molecular understanding of exocytotic vesicle fusion and transmitter release in endocrine cells and nerve terminals. Upon electrical stimulation nerve terminals and endocrine cells release a variety of neurotransmitters and neuropeptides by an exocytotic mechanism. This process of neurosecretion is of outstanding importance in a broad range of physiological functions in the human body. It allows for synaptic transmission as well as release of hormones and neuromodulators and is thus an essential event mediating brain function, emotional response, behavior and various other physiological processes. A detailed understanding of the exocytotic event is of particular interest for diseases where neurosecretion is impaired. It will also advance our understanding of many other cellular functions in health and disease because exocytotic fusion events play a central role in the immune response against bacteria or parasites invading a host organism and because many other fusion events that occur inside the cell or the fusion events during viral entry appear to use closely related mechanisms. Upon stimulation the contents of the secretory vesicles are released through a fusion pore that connects the vesicular lumen to the extracellular space. The mechanism by which the fusion pore is formed and the molecular components forming the fusion pore are therefore central to understanding the mechanisms of exocytosis. The SNARE (Soluble NSF Attachment REceptor) complex, composed of the proteins synaptobrevin, syntaxin and SNAP-25 forming a coiled coil, a parallel four-helix bundle, plays a key role in exocytosis and may be responsible for tight binding and fusion between vesicle and plasma membrane. This proposal is based on the hypothesis that the fusion pore is opened and expanded by SNARE proteins but that additional proteins participate in fusion pore formation and expansion. We study exocytosis of single vesicles in chromaffin cells combining electrophysiological measurements of fusion pore conductance with amperometric measurements of catecholamine release through the fusion pore. We perform a detailed characterization of the fusion pore selectivity and fusion pore dynamics to obtain a clear understanding of the mechanisms by which the fusion pore properties regulate the release of transmitter from individual vesicles. Molecular manipulations of the SNARE proteins SNAP-25 and VAMP2 are performed in conjunction with measurements of fusion pore opening, fusion pore conductance, lifetime, fluctuations, selectivity and transmitter release to determine molecular basis of fusion pore formation, fusion pore structure, fusion pore dynamics and the mechanism of transmitter release through the fusion pore. PUBLIC HEALTH RELEVANCE: Neurotransmitters and hormones as well as various other compounds are stored at high concentration in membrane-bound organelles, called secretory vesicles, inside the cell. Upon stimulation a fusion pore is formed connecting the vesicular lumen to the extracellular space, thereby releasing the stored molecules. This project is aimed at elucidating the molecular structure and dynamics of the fusion pore and mechanism by which transmitters are released through the fusion pore.

描述（由申请人提供）：该项目的主要长期目标是在内分泌细胞和神经末端提供详细的生物物理和分子理解和胞外囊泡融合和递质释放。电刺激后，神经末端和内分泌细胞通过胞吐机制释放了多种神经递质和神经肽。在人体的广泛生理功能中，这种神经分泌的过程至关重要。它允许突触传播以及激素和神经调节剂的释放，因此是介导大脑功能，情绪反应，行为以及各种其他生理过程的重要事件。对胞吐事件的详细理解对于神经分泌受损的疾病特别感兴趣。这也将提高我们对健康和疾病中许多其他细胞功能的理解，因为胞外融合事件在对细菌或寄生虫的免疫反应中起着核心作用，因为入侵了宿主有机体，以及在病毒进入过程中发生的许多其他融合事件，似乎使用了密切相关的机制。刺激后，分泌囊泡的含量将通过将囊泡腔连接到细胞外空间的融合孔释放。因此，形成融合孔和形成融合孔的分子成分的机制对于理解胞吐作用的机制至关重要。由蛋白质蛋白突触，语法素和SNAP-25组成的SNARE（可溶性NSF附着受体）复合物，形成盘绕的线圈（平行的四旋螺旋束）在胞外胞病症中起关键作用，并可能负责Vesicle和Plasmambrane之间的紧密结合和融合。该提议基于以下假设：融合孔由圈圈蛋白打开和扩展，但额外的蛋白质参与融合孔的形成和膨胀。我们研究了通过融合孔通过融合孔的融合孔电导率的电生理测量和对儿茶酚胺释放的安培测量的电生理测量结果的单囊泡的胞吐作用。我们对融合孔的选择性和融合孔动力学进行详细的表征，以获得对融合孔特性调节从单个囊泡释放发射机的机制的清晰理解。 SNARE蛋白SNAP-25和VAMP2的分子操作是通过测量融合孔口的测量，融合孔电导，寿命，波动，选择性，选择性和递质释放的，以确定融合孔形成的分子基础，融合孔结构，融合孔动力学和融合式的机制，融合孔结构，融合孔动力学和融合机制。公共卫生相关性：神经递质和激素以及其他各种化合物在细胞内部的膜结合细胞器中储存在膜结合的细胞器中，称为分泌囊泡。刺激后，形成了融合孔，将囊泡腔连接到细胞外空间，从而释放储存的分子。该项目旨在阐明融合孔的分子结构和动力学和通过融合孔释放发射机的机制的动力学。