PKC modulation of calcium current and anesthetic action

PKC 调节钙电流和麻醉作用

• 批准号: 7101698
• 负责人: GANESAN Lenin KAMATCHI
• 金额: $ 21.06万
• 依托单位: UNIVERSITY OF VIRGINIA
• 依托单位国家: 美国
• 财政年份: 2003
• 资助国家: 美国
• 起止时间: 2003-08-01 至 2008-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7101698
• 关键词: SDS polyacrylamide gel electrophoresis; Xenopus oocyte; active sites; anesthetics; calcium channel; drug interactions; electrophysiology; enzyme induction /repression; halothane; immunocytochemistry; immunoprecipitation; isoflurane; isozymes; microinjections; neural transmission; pharmacokinetics; phosphorylation; polymerase chain reaction; protein kinase C; receptor expression; site directed mutagenesis; voltage gated channel; western blottings

DESCRIPTION (provided by applicant): Voltage-gated Ca2+-channels (Cav) and protein kinase C (PKC) can both be regulated by volatile anesthetics, but the nature of the effect has been controversial. We hypothesize that anesthetic inhibition of Cav channels and synaptic transmission are mediated by specific PKC isozymes. Although potential PKC phosphorylation sites are present in the pore-forming alpha1 and the auxiliary beta and alpha2/delta subunits of Cav channels; alpha1subunits seem to play the major role in PKC-induced regulation of these channels. In Aim I of this proposal, we will identify phosphorylation sites in the a1 subunits of Cav channels by direct (with phorbol 12-myristate 13-acetate; PMA) or receptor-stimulated (with muscarinic M1) activation of PKC in Xenopus oocytes coexpressing these receptors and Cav channels. Possible PKC phosphorylation sites will be mutated to Ala or Glu and whole-cell Ba2+ (a substitute for Ca2+)-current in response to two types of PKC activation will be investigated using voltage-clamp measurements. In Aim II we will identify PKC isozymes involved in the direct or receptor-induced modulation of PKC-sensitive Cav channels expressed in Xenopus oocytes. This will be examined using experiments involving down-regulation of PKC, activation-induced translocation, selective loss/inhibition of individual PKC isozymes and 'add-back' studies. In Aim III we will identify the PKC isozymes modulated by volatile anesthetics, halothane or isoflurane in Xenopus oocytes by employing the methods mentioned in Aim II. Based on the results of our studies, isozyme selective activators or inhibitors of PKC may be used along with the volatile general anesthetics to i) potentiate the action of anesthetics and ii) to reduce the side effects of these agents caused by their non-specific actions.

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