PROPERTIES OF GABA C RECEPTORS ON RETINAL NEURONS

视网膜神经元上 GABA C 受体的特性

• 批准号: 6342662
• 负责人: HAOHUA QIAN
• 金额: $ 14.8万
• 依托单位: UNIVERSITY OF ILLINOIS AT CHICAGO
• 依托单位国家: 美国
• 财政年份: 1998
• 资助国家: 美国
• 起止时间: 1998-01-01 至 2002-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6342662
• 关键词: GABA receptor; X ray; Xenopus oocyte; calmodulin dependent protein kinase; cyclic AMP; electrophysiology; fish; horizontal cell; in situ hybridization; molecular shape; neurotransmitter transport; nucleic acid chemical synthesis; nucleic acid sequence; phosphorylation; picrotoxin; polymerase chain reaction; protein kinase A; protein kinase C; protein sequence; receptor expression; retina; retinal bipolar neuron; second messengers; site directed mutagenesis; voltage /patch clamp

DESCRIPTION (Adapted from applicant's abstract): The application proposes to examine the physiology, pharmacology and kinetics of GABAc receptors on retinal neurons and GABA-rho subunits expressed in Xenopus oocytes. In the first specific aim, the applicant will investigate the five subunits of GABA-rho, which he has cloned from white perch retina. He will examine their electrophysiological and pharmacological properties. He plans to compare the GABA dose-response curves among the five subunits and the pharmacology of antagonists such as I4AA and picrotoxin. In the different subunits, the applicant proposes to compare the kinetics of both the onset and offset of the GABA response using outside-out patch recordings extracted from oocytes. After deciphering these biophysical properties of the individual subunits, he plans to study GABAc receptors expressed in white perch retinal neurons. There will be particular emphasis on bipolar cells and the sensitivity and pharmacology of these cells will be compared to H4 horizontal cells. The applicant will also localize various subunits of the GABAc receptor in white perch retina using in situ hybridization and RT-PCR techniques. The rationale is that a different mix of GABAc receptor subunits in each type of neuron may produce distinct response properties. In a second specific aim, the applicant proposes to study the molecular structure of the GABAc receptor. The rationale is that although the GABAc receptor subunits seem similar to each other, the pharmacology and kinetics are different. Since the amino acid sequence of the subunits are very similar, it may be possible to identify just a few amino acids that account for these differences. The applicant will use site-directed mutagenesis and construction of chimeras to investigate structure/function relationships among these subunits. In the third and final specific aim, the applicant will investigate the modulation of GABAc receptors by second messengers. Based on sequencing data, the applicant has identified cAMP-dependent protein kinase (PKA), protein kinase C (PKC), and calmodulin (CaM)-II kinase consensus sites on the putative cytoplasmic domain of all of these receptor subunits. He will, therefore, investigate the role of phosphorylation in the modulation of each of the five subunits.

描述（根据申请人的摘要改编）：申请提出 检查GABAC受体的生理学，药理学和动力学 视网膜神经元和GABA-RHO亚基在爪蟾卵母细胞中表达。 在 第一个具体目的，申请人将调查的五个亚基 Gaba-rho，他是从White Perch Retina克隆的。 他会检查 它们的电生理和药理特性。 他计划 比较五个亚基之间的GABA剂量响应曲线和 I4AA和PICROTOXIN等拮抗剂的药理学。 在不同的地方 亚基，申请人建议比较两种发作的动力学 并使用提取的外部贴片记录抵消GABA响应的响应 来自卵母细胞。 解密了这些生物物理特性之后 个别亚基，他计划研究以白色表达的GABAC受体 栖息的视网膜神经元。 将特别强调双极细胞 这些细胞的敏感性和药理学将与H4进行比较 水平细胞。 申请人还将定位 使用原位杂交和RT-PCR的白鲈鱼中的GABAC受体 技术。 理由是GABAC受体的不同混合 每种类型的神经元中的亚基可能会产生不同的响应特性。 在第二个特定目的中，申请人建议研究分子 GABAC受体的结构。 理由是尽管GABAC 受体亚基似乎彼此相似，药理学和动力学 是不同的。 由于亚基的氨基酸序列非常 类似，可能只能识别一些氨基酸 对于这些差异。 申请人将使用以网站为导向的诱变和 构建嵌合体以研究结构/功能关系 在这些亚基中。 在第三个也是最后一个特定目标中，申请人将调查 第二使者对GABAC受体的调节。 基于测序 数据，申请人已经确定了cAMP依赖性蛋白激酶（PKA）， 蛋白激酶C（PKC）和钙调蛋白（CAM）-II激酶共识位点 所有这些受体亚基的假定细胞质结构域。 他会， 因此，研究磷酸化在每个调制中的作用 在五个亚基中。