ENDOGENOUS LIGANDS FOR GLUTAMATE RECEPTORS IN BRAIN

L谷氨酸(Glu)和L天冬氨酸(Asp)被认为是重要的 脊椎动物神经系统中的兴奋性氨基酸。然而， 它们作为真正的内源性配体的身份被很好地表征 EAA受体的亚型尚不确定。这些研究 在本提案中描述的是用来测试假设在 添加(或代替)带有C-末端谷氨酸的小肽Glu和Asp 和天冬氨酸分别可能存在于大脑中，并可能作为 兴奋性神经递质或调节剂。这一假设是基于 初步研究表明，我们制备的两种抗Glu的抗血清 和Asp分别识别。(1)谷氨酸和天冬氨酸；(2)C-二肽 末端谷氨酸和天冬氨酸残基，(3)可以 区分EAA受体的亚类。因此，Quisqualate(Quis) 用谷氨酸抗血清海人酸(KA)选择性阻断免疫染色 用天冬氨酸抗血清阻断这种作用，而N-甲基-D-天冬氨酸(NMDA)不能 被任何一种抗血清识别(一种新的NMDA抗血清在 准备)。我们研究的主要目标是使用这些抗血清 鉴定和鉴定可能的内源性配体 大鼠EAA受体的三种主要亚型(Quis、KA和NMDA) 大脑。首先，抗血清将进一步表征在以下方面 最佳抗原识别的分子要求，使用合成的 二肽以及受体激动剂和拮抗剂。第二， 这些抗血清识别的内源性化合物将从 用抗血清中的免疫球蛋白组份制备的脑上亲和层析柱。 最后是层析。酶免疫分析。光和电子 显微免疫细胞化学和体外受体激活研究 将用于监测和补充这些程序。 预计这些研究的结果将提供基本的新的 对EAA神经传递的分子机制的洞察。 鉴定EAA受体的新内源性配体将允许 更好地描述了使用这些配体的神经元通路， 将有助于为EAA设计新的特定激动剂和拮抗剂 并将为理解和治疗这种疾病提供线索 疾病的过程是中风和低血糖后的脑损伤， 癫痫、亨廷顿舞蹈症和阿尔茨海默病。

L-Glutamate (Glu) and L-aspartate (Asp) are regarded as important excitatory amino acids (EAA) in the vertebrate nervous systems. However, their identity as the true endogenous ligands for the well characterized subtypes of EAA receptors is not established with certainty. The studies described in this proposal are designed to test the hypothesis that in addition to (or instead of) Glu and Asp, small peptides with C-terminal Glu and Asp, respectively, may be present in the brain and may act as excitatory neurotransmitters or modulators. This hypothesis is based on preliminary studies indicating that two antisera we have produced to Glu and Asp recognize,respectively. (1)Glu and Asp, (2) dipeptides with C- terminal Glu and Asp residues, (3) nonendogenous compounds which can discriminate between subclasses of EAA receptors. Thus, quisqualate (Quis) selectively blocks the immunostaining with the Glu antiserum, kainate (KA) blocks that with the Asp antiserum, while N-methyl-D-Asp (NMDA) is not recognized by either of the antisersa (a new antiserum to NMDA is in preparation). The principal goal of our studies is to use these antisera to identify and characterize putative endogenous ligands specific for the three major subtypes (Quis, KA, and NMDA) of EAA receptors in the rat brain. First, the antisera will be further characterized in terms of the molecular requirements for optimal antigen recognition, using synthetic dipeptides as well as receptor agonists and antagonists. Second, the endogenous compounds recognized by these antisera will be extracted from brain on affinity columns prepared with IgG fractions from the antisera. Finally, chromatography. Enzyme immunoassays. light and electron microscopic immunocytochemistry, and in vitro receptor activation studies will be used to monitor and complement these procedures. The results of these studies are expected to provide fundamental new insights into the molecular mechanisms of EAA neurotransmission. Identification of new endogenous ligands for EAA receptors will allow a better characterization of the neuronal pathways that use these ligands, will help in the design of new specific agonists and antagonists for EAA receptors and will provide clues for the understanding and therapy of such disease processes as brain damage following stroke and hypoglycemia, epilepsy, Huntington's chorea, and Alzheimer's disease.