GLUTAMATE TRANSPORTERS IN THE CNS

中枢神经系统中的谷氨酸转运蛋白

基本信息

批准号：
6363905
负责人：
Michael Byrne Robinson
金额：
$ 35.12万
依托单位：
CHILDREN'S HOSP OF PHILADELPHIA
依托单位国家：
美国
项目类别：
财政年份：
1997
资助国家：
美国
起止时间：
1997-05-01 至 2002-07-31
项目状态：
已结题

项目摘要

DESCRIPTION: The acidic amino acids are the predominant excitatory neurotransmitters in the mammalian central nervous system (CNS). Most evidence indicates that Na+-dependent high affinity transporters are responsible for the clearance and regulation of extracellular excitatory amino acids (EAA). Pharmacological, biochemical, and molecular biological studies indicate that there are several subtypes of Na+-dependent glutamate (Glu) transporters that are expressed in different brain regions and on different cell types, including the astroglial transporters GLT-1 and GLAST, and the neuronal transporters EAAC1 and EAAT4. This project represents a collaborative effort between two laboratories who have defined the biochemical, pharmacological and pathophysiological properties of Glu transport. The goal of the investigators is to increase understanding of the predominant mechanism that normally protects the brain from excitotoxicity. This information should improve our understanding of the failure of these systems during acute and chronic insults to the CNS, including stroke and head trauma. The investigators will use in vitro and in vivo preparations to test the functional significance of our observations. The investigators have previously established the pharmacology of transporter activity in vitro and in vivo preparations. The investigators hypothesize that the cloned transporters recapitulate the properties observed in these preparations. This hypothesis will be tested by expressing the individual transporters in xenopus oocytes. In addition to this correlative strategy, the investigators propose to use antisense oligonucleotides to "knock-out" subtypes of the transporters to examine the relative contributions of each transporter to activity observed in astrocyte-enriched cultures and in synaptosomes prepared from different brain regions. The investigators hypothesize different transporters contribute to the regulation of extracellular Glu in the pre- and early post-natal period. The investigators propose to study the level of expression and ultrastructural localization of the transporter subtypes during development. Based on the preliminary data, the investigators hypothesize that neurons regulate expression of glial Glu transporters. In vivo and in vitro systems will be used to examine this regulation. The investigators propose that cAMP through activation of protein kinase A is one of the factors that controls expression of the GLT-1 subtype of glial transporter. This will be tested in astrocyte-enriched cultures. The effects of selective antisense "knock-out" of subtypes of transporters on excitotoxicity and the accumulation of extracellular Glu will be examined in vitro. The investigators also propose to study the effects of excitotoxic insults on the expression of subtypes of transporters.

描述：酸性氨基酸是主要的兴奋性哺乳动物中枢神经系统（CNS）中的神经递质。最多证据表明Na+依赖性高亲和力转运蛋白是负责清除和调节细胞外兴奋性氨基酸（EAA）。药理，生化和分子生物学研究表明，Na+依赖性谷氨酸有几种亚型（GLU）在不同大脑区域和上表达的转运蛋白不同的细胞类型，包括星形胶质细胞转运蛋白GLT-1和GLAST，以及神经元转运蛋白EAAC1和EAAT4。这个项目代表两个定义的实验室之间的合作努力 GLU的生化，药理和病理生理特性运输。调查人员的目的是增加对通常保护大脑免受的主要机制兴奋性毒性。这些信息应提高我们对这些系统在急性和长期侮辱中枢神经系统期间的失败，包括中风和头部创伤。调查人员将在体外使用体内准备测试我们的功能意义的准备观察。调查人员以前已经建立了转运蛋白活性在体外和体内制剂的药理学。这研究人员假设克隆的转运蛋白概括了在这些制剂中观察到的特性。该假设将进行检验通过表达异爪蟾卵母细胞中的单个转运蛋白。此外对于这种相关策略，研究人员建议使用反义寡核苷酸以“敲除”转运蛋白的亚型来检查每个转运蛋白对观察到的活性的相对贡献富含星形胶质细胞的培养物和从不同的突触体中大脑区域。研究人员假设不同的转运蛋白有助于调节前后早期细胞外GLU 产后期。调查人员建议研究转运蛋白亚型的表达和超微结构定位在开发过程中。根据初步数据，调查人员假设神经元调节神经胶质Glu转运蛋白的表达。在体内和体外系统将用于检查该调节。这研究人员建议通过激活蛋白激酶A IS扎营控制神经胶质GLT-1亚型表达的因素之一转运蛋白。这将在富含星形胶质细胞的文化中进行测试。这转运蛋白亚型的选择性反义“敲除”对兴奋性毒性和细胞外GLU的积累将在体外。研究人员还建议研究兴奋毒性的影响侮辱转运蛋白亚型的表达。