PHOSPORYLATION EFFECT ON GABAA RECEPTOR FUNCTION

磷酸化对 GABAA 受体功能的影响

• 批准号: 2270126
• 负责人: NANCY J LEIDENHEIMER
• 金额: $ 8.82万
• 依托单位: LOUISIANA STATE UNIV HSC SHREVEPORT
• 依托单位国家: 美国
• 财政年份: 1994
• 资助国家: 美国
• 起止时间: 1994-05-01 至 1999-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2270126
• 关键词: GABA receptor; Xenopus; Xenopus oocyte; allosteric site; barbiturates; benzodiazepines; calcineurin; calmodulin dependent protein kinase; complementary DNA; enzyme activity; gamma aminobutyrate; gene mutation; microinjections; neuropharmacology; phosphoprotein phosphatase; phosphorylation; protein kinase; protein kinase A; protein kinase C; protein structure function; protein tyrosine kinase; receptor binding; recombinant proteins; site directed mutagenesis; voltage /patch clamp

The GABA receptor is a ligand-gated chloride channel which mediates the inhibitory effects of the neurotransmitter GABA (gamma-aminobutyric acid). In addition to binding GABA, the receptor is modulated by several therapeutic agents and endogenous neurosteroids which act through distinct, allosteric binding sites. Over fifteen different receptor subunits have been cloned and sequenced and multiple receptor subtypes exist in vivo. There is evidence that the GABAa receptor is regulated by protein phosphorylation and that is this regulation is complex and involves multiple second messenger system pathways. The kinases and phosphatases which accompany this regulation are not well-characterized. The hypotheses of the present proposal are: 1) multiple kinases and phosphatases participate in the phosphorylation/dephosphorylation of the receptor, 2) the subunit composition of the receptor determines how the receptor is regulated by phosphorylation, and 3) phosphorylation alters the allosteric modulatory interactions of the receptor complex. To test these hypotheses, protein kinases and phosphatases will be microinjected into Xenopus oocytes expressing either recombinant wildtype or mutagenized human GABA receptor subunit cDNAs. The effect of these enzymes on both GABA receptor-gated chloride currents and the allosteric modulation of these currents by benzodiazepines, barbiturates and neurosteroids, will be measured using the two-electrode voltage-clamp technique. Thus, the proposed research adopts a reductionistic strategy to study GABA receptor phosphorylation/dephosphorylation under conditions where the receptor composition is defined and site-directed mutagenesis can be used to identify phosphorylation sites on the receptor. These studies will aid in the understanding of GABA receptor function in the overall scheme of neuronal signaling. The involvement of the GABA receptor in hyperexcitable states such as anxiety disorders and epilepsy, underscores the importance of understanding GABA receptor function.

GABA受体是一种配体门控的氯离子通道