BRAIN ACETYLCHOLINE STORAGE SYSTEM

脑乙酰胆碱储存系统

• 批准号: 3411860
• 负责人: STANLEY MONROE PARSONS
• 金额: $ 13.31万
• 依托单位: UNIVERSITY OF CALIFORNIA SANTA BARBARA
• 依托单位国家: 美国
• 财政年份: 1988
• 资助国家: 美国
• 起止时间: 1988-04-01 至 1994-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3411860
• 关键词: acetylcholine; active transport; affinity labeling; allosteric site; brain; brain metabolism; cholinergic receptors; depolarizing neuromuscular blocking agent; drug receptors; gel electrophoresis; laboratory rat; neurochemistry; neuropharmacology; neurotransmitter metabolism; neurotransmitter receptor; proteoglycan; radiotracer; receptor binding; second messengers; synaptic vesicles; synaptosomes; tissue /cell culture

This project will characterize biochemical and pharmacological aspects of acetylcholine (ACh) storage by rat brain synaptic vesicles. The storage system is composed of a proton-pumping ATPase and a separate ACh transporter that is associated with an allosteric receptor site for the inhibitory drug vesamicol. The previously characterized ACh transporter- vesamicol receptor (AChT-VR) complex of the electric ray, surprisingly, is strongly associated with synaptic vesicle proteoglycan. This project will determine how similar the rat brain AChT-VR is to the electric ray AChT-VR. Rat brain synaptic vesicles will be prepared either in free form (that is, a P3 preparation) or inside metabolically active synaptosomes. Active transport of radiolabeled ACh and binding of radio-labeled vesamicol or a more tightly binding analog to these preparations will be studied using filter assays. Other biochemical, bioorganic and immunochemical techniques also will be used as appropriate. The specific aims are to (1) study the mechanism and specific ion dependence of the rat brain ACh active transport system, (2) study the relationship of vesamicol analog binding to transport inhibition, (3) test whether the AChT-VR in rat vesicles is linked to proteoglycan, (4) identify the rat brain AChT-VR by photo-affinity labeling with a tritiated high affinity ACh analog, (5) develop an assay for the AchT-VR inside of rat brain synaptosomes based on binding of a high affinity, radioactive analog of vesamicol, and (6) subject the synaptosomes to different pharmacological manipulations that alter the metabolism and that might affect the properties of the AChT-VR. The long term goal of the project is to understand the complete biochemistry of ACh storage by synaptic vesicles in mammals, whether it is subject to regulation, and whether storage and release are coupled. This might lead to development of a pharmacology that would increase ACh storage in and release from compromised cholinergic terminals. Since many disease and toxic states impact on the cholinergic nervous system, this capability could be of significant clinical benefit.

该项目将表征生物化学和药理学方面 大鼠脑突触小泡储存乙酰胆碱（ACh）。 存储 系统由质子泵浦 ATP 酶和单独的 ACh 组成 与变构受体位点相关的转运蛋白 抑制药维沙考。 先前表征的乙酰胆碱转运蛋白- 令人惊讶的是，电射线的维沙考受体（AChT-VR）复合物是 与突触小泡蛋白多糖密切相关。 该项目将 确定大鼠大脑 AChT-VR 与电射线 AChT-VR 的相似程度。 大鼠脑突触小泡将以游离形式（即， P3 制剂）或代谢活跃的突触体内。 积极的 放射性标记的乙酰胆碱的转运和放射性标记的维萨米考或a的结合 将使用这些制剂更紧密结合的类似物进行研究 过滤分析。 其他生化、生物有机和免疫化学技术 也会酌情使用。 具体目标是 (1) 研究 大鼠脑乙酰胆碱主动转运的机制和特异性离子依赖性 系统，（2）研究vesamicol类似物结合与运输的关系 (3) 测试大鼠囊泡中的 AChT-VR 是否与 蛋白多糖，(4)通过光亲和标记识别大鼠大脑AChT-VR 与氚化高亲和力乙酰胆碱类似物一起，(5) 开发一种检测方法 基于高结合力的大鼠脑突触体内部的 AchT-VR 亲和力，维沙考的放射性类似物，以及（6）使突触体受到影响 改变新陈代谢的不同药理操作 这可能会影响 AChT-VR 的特性。 该项目的长期目标是了解完整的 哺乳动物突触小泡储存乙酰胆碱的生物化学，无论是 受监管，以及存储和释放是否耦合。 这 可能会导致增加乙酰胆碱储存的药理学的发展 进入受损的胆碱能末端并从受损的胆碱能末端释放。 由于多种疾病 和有毒状态对胆碱能神经系统的影响，这种能力 可能具有显着的临床益处。