MOLECULAR ANALYSIS OF ENDOGENOUS CANNABINOID TRANSPORT

内源性大麻素运输的分子分析

• 批准号: 6379031
• 负责人: ERIC L BARKER
• 金额: $ 15万
• 依托单位: PURDUE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2000
• 资助国家: 美国
• 起止时间: 2000-09-30 至 2003-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6379031
• 关键词: Mammalia; anandamide; biological transport; fatty acid transport; membrane permeability; neuronal transport; protein structure function; tissue /cell culture; transfection; transport proteins

DESCRIPTION: (Applicant's Abstract) The broad, long-term objective of this R21 application is to advance the overall understanding of molecular mechanisms regulating endogenous cannabinoid signaling by a putative endocannabinoid uptake system. The endogenous cannabinoids such as anandamide are fatty acid-derived molecules whose metabolism and signal termination appear to rely upon transport back into releasing cells for subsequent breakdown by an intracellular amidohydrolase enzyme. Thus, the uptake process would play a critical role in the dynamic regulation of endogenous cannabimimetic activity. The Specific Aims are (1) To isolate and identify the protein(s) responsible for the uptake of endogenous cannabinoids, and (2) To characterize the transport activity associated with anandamide uptake. This project has direct health-relatedness as the endogenous cannabinoid system has been implicated in modulating various physiological and pathological processes including psychosis, motor movement, inflammation, blood pressure, and pain. Based upon data indicating functional similarities between anandamide transport and the uptake of long-chain fatty acids, the Research Design is to explore the potential contributions of various fatty acid transporters in endocannabinoid transport using heterologous expression systems. In addition, the pharmacologic profile of endocannabinoid uptake in native and transfected cells will be determined, and the tissue distribution of the identified anandamide transporter investigated to better understand the role of transport proteins in endocannabinoid signaling. The Methods to be used include molecular cloning techniques, characterization of transport proteins using radiolabeled substrate uptake assays, determinations of intracellular accumulation of endocannabinoid substrates using mass spectrometry, and in situ hybridization to reveal transporter distribution. These studies will provide a basis for future studies examining the molecular mechanisms associated with both structure/function and regulation of endogenous cannabinoid transport.

描述：（申请人的摘要） 该R21应用的广泛长期目标是促进 对调节内源性大麻素的分子机制的总体理解 通过推定的内源性大麻素摄取系统发出信号。内源性 大麻素（例如anandamide）是脂肪酸衍生的分子 新陈代谢和信号终止似乎依赖于运输 释放细胞以随后通过细胞内酰胺水解酶分解 酶。因此，吸收过程将在动态中起关键作用 调节内源性大麻活性。具体目的是（1） 分离并鉴定蛋白质负责摄取内源性的蛋白质 大麻素和（2）表征与 anandamide摄取。该项目具有直接与健康相关性作为内源性 大麻素系统与调节各种生理和 病理过程，包括精神病，运动运动，炎症，血液 压力和痛苦。基于指示功能相似性的数据 该研究 设计是探索各种脂肪酸的潜在贡献 使用异源表达的内源性大麻素运输中的转运蛋白 系统。此外，内源性大麻素摄取的药理学特征 将确定天然和转染的细胞，并将组织分布 已确定的Anandamide运输蛋白调查，以更好地了解 转运蛋白在内源性大麻素信号传导中的作用。要使用的方法 包括分子克隆技术，表征转运蛋白 使用放射标记的底物摄取测定，测定细胞内 使用质谱和原位的内源性大麻素底物积累 杂交以揭示转运蛋白分布。这些研究将提供 未来研究与分子机制相关的研究的基础 内源性大麻素转运的结构/功能和调节。