HUMAN GABA RECEPTORS

人类 GABA 受体

基本信息

批准号：
6187886
负责人：
ROBERT A DAVIDOFF
金额：
$ 24.55万
依托单位：
UNIVERSITY OF MIAMI SCHOOL OF MEDICINE
依托单位国家：
美国
项目类别：
财政年份：
1998
资助国家：
美国
起止时间：
1998-07-15 至 2002-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/6187886
关键词：
GABA receptor Xenopus oocyte afferent nerve electrophysiology embryo /fetus tissue /cell culture gamma aminobutyrate human fetus tissue human genetic material tag laboratory rat neurons neuropharmacology neurotransmitter antagonist picrotoxin spinal ganglion voltage /patch clamp

项目摘要

DESCRIPTION: This proposal aims to investigate the electrophysiological/ pharmacological characteristics and subunit composition of GABA receptors located on human primary afferent neurons. The proposed research stems from intriguing results of recent investigations which establish that cultured human primary afferent neurons express GABA receptors with unique, and hitherto undescribed, pharmacological properties. An association with both a large transplant center and an internationally recognized spinal cord trauma unit has provided the applicants with access to human neurons. Three separate laboratories with expertise in: (1) neuronal cell culturing; (2) electrophysiology and pharmacology of GABA receptors; and (3) molecular biology of neurotransmitter receptors will participate in these studies. The overall intent is to clarify the properties of these novel human GABA receptors by using a combination of sophisticated electrophysiological techniques, pharmacological probes, and molecular biological methods so as to define the channel/molecular properties and to determine the GABA receptor subunits in human sensory neurons. The proposed investigations are important to neurobiology, implicated in disease states (e.g., spinal spasticity, pathophysiology of pain). The results will be relevant to understanding how spinal neuron excitability is controlled by therapeutic agents that alter afferent transmission by acting at GABA receptors, and hence should foster eventual development of more efficacious pharmacological treatment of spinal cord injuries, pain states, and other pathologies. Whole-cell, cell-attached, and outside-out patch clamp recordings from cultured embryonic and adult human dorsal root ganglion (DRG) neurons will be used to define the electrophysiological/pharmacological properties of GABA receptors. The results of these investigations will be compared with data from molecular biological experiments designed to determine the subset of GABA receptor subunit-encoding mRNAs expressed by cultured embryonic and adult human DRG neurons. RT-PCR on cultures of DRG neurons will reveal the potential array of involved subunits. Based on conclusions drawn from these data, the candidate subunit combinations reproducing the properties of the novel GABA receptor will be identified using exogenous expression systems and antisense oligonucleotides. The reduced number of subunits potentially responsible for the observed properties reduced in this manner to a manageable number will allow performance of single-neuron RT-PCR to correlate expression of functional properties with expression of particular subunit mRNAs. These investigations will allow the applicants to specify the properties of a defined human GABA receptor with unique pharmacological properties within a native neuronal environment.

描述：本提案旨在研究电生理学/ GABA受体的药理特征和亚基组成位于人类初级传入神经元上。拟议的研究源于最近的调查令人感兴趣的结果表明，文化人类初级传入神经元表达具有独特的 GABA 受体，并且迄今为止尚未描述的药理学特性。与两者都有关联大型移植中心和国际认可的脊髓创伤科为申请人提供了接触人类神经元的机会。三拥有以下专业知识的独立实验室：(1) 神经元细胞培养； (2) GABA受体的电生理学和药理学； (3)分子神经递质受体生物学将参与这些研究。总体目的是阐明这些新型人类 GABA 的特性受体通过使用复杂的电生理学的组合技术、药理学探针和分子生物学方法，以便定义通道/分子特性并确定 GABA 人类感觉神经元中的受体亚基。拟议的研究对神经生物学很重要，涉及疾病状态（例如，脊柱痉挛、疼痛的病理生理学）。这结果将有助于了解脊髓神经元的兴奋性如何由治疗剂控制，通过作用改变传入传递 GABA受体，因此应该促进更多的最终发展有效的药物治疗脊髓损伤、疼痛状态、和其他病症。全细胞、细胞贴壁和外向膜片钳记录培养的胚胎和成人背根神经节（DRG）神经元将用于定义电生理学/药理学特性 GABA 受体。这些调查的结果将与来自分子生物学实验的数据，旨在确定子集培养胚胎表达的 GABA 受体亚基编码 mRNA 成人 DRG 神经元。对 DRG 神经元培养物进行 RT-PCR 将揭示涉及的亚基的潜在阵列。基于这些得出的结论数据，候选子单元组合再现了新型 GABA 受体将使用外源表达系统进行鉴定和反义寡核苷酸。潜在的亚基数量减少负责以这种方式减少观察到的属性可控的数量将允许单神经元 RT-PCR 的性能将功能特性的表达与特定的表达相关联亚基 mRNA。这些调查将使申请人能够具体说明具有独特药理作用的特定人类 GABA 受体的特性天然神经元环境中的特性。