REGULATION OF CARDIAC K+ CHANNEL FUNCTION

K通道在心脏细胞功能中起重要作用。长期 本研究的目的是了解细胞过程， 哪些心脏K通道被激活和脱敏 神经递质，激素和药物，以及这些过程如何影响 心率、力量和节律。在这项建议中，将研究 聚焦于心房肌氨酸门控K通道， 乙酰胆碱（ACh）通过抑制性GTP结合蛋白（Gi）。虽然 激活的机制， 毒蕈碱钾电流是已知的， 细胞过程的后续快速和缓慢阶段的K 目前的脱敏是完全未知的。使用全细胞电压- 钳和膜片钳方法，拟议的研究将首先测试 假设K电流脱敏是由 乙酰胆碱激活的钾通道通过磷酸化开放的动力学 和去磷酸化。 K电流脱敏将完全 其特征在于存在和不存在抑制或 激活特定的蛋白激酶或磷酸酶。 由于乙酰胆碱激活 磷脂酶，我们还将检查是否某些细胞内第二 通过磷脂代谢产生的信使（如花生四烯酸 酸或其代谢物）参与K电流脱敏。使用 激动剂如腺苷、ATP、内皮素、生长抑素和降钙素 基因相关肽，其通过以下途径激活心房毒蕈碱K电流 单独的，非毒蕈碱受体，我们将研究是否乙酰胆碱- 诱导的K电流脱敏是同源的或异源的 过程最后，毒蕈碱钾电流在乙酰胆碱诱导的 负变时性及其脱敏将在 心房细胞自发收缩，通过测量 上述修改K通道功能的各种条件。 这些研究将为细胞生物学提供重要的新知识， 毒蕈碱钾电流的快速和缓慢脱敏机制。 这些研究将了解磷酸化的作用， 蛋白激酶和磷酸酶的去磷酸化作用以及 K通道磷脂酶介导的第二信使 脱敏 毒蕈碱钾电流在乙酰胆碱引起的变化中的作用 收缩行为的影响。

K channels play an important role in cardiac cell function. The long-tern objective of this research is to understand the cellular processes by which cardiac K channels are activated and desensitized by neurotransmitters, hormones and drugs, and how these processes affect cardiac rate, force, and rhythm. In this proposal, studies will be focussed on the atrial muscarinic-gated K channel that is activated by acetylcholine (ACh) via the inhibitory GTP binding protein (Gi). Although the mechanism of activation of the muscarinic K current is known, the cellular processes underlying the subsequent rapid and slow phases of K current desensitization are completely unknown. Using whole-cell voltage- clamp and patch-clamp methods, the proposed studies will first test the hypothesis that the K current desensitization is produced by changes in the kinetics of the ACh-activated K channel opening via phosphorylation and dephosphorylation. The K current desensitization will be fully characterized in the presence and absence of agents that inhibit or activate specific protein kinases or phosphatases. Since ACh activates phospholipases, we will also examine whether certain intracellular second messengers generated via phospholipid metabolism (such as arachidonic acid or its metabolites) are involved in K current desensitization. Using agonists such as adenosine, ATP, endothelin, somatostatin and calcitonin gene-related peptide which activate the atrial muscarinic K current via separate, non-muscarinic receptors, we will investigate whether the ACh- induced K current desensitization is a homologous or a heterologous process. Finally, the role of muscarinic K current in the ACh-induced negative chronotropy and its desensitization will be studied in spontaneously contracting atrial cells by measuring beating rates under various conditions described above that modifies the K channel function. These studies will provide important new knowledge on the cellular mechanisms of rapid and slow desensitization of the muscarinic K current. The studies will understand the role of phosphorylation and dephosphorylation by protein kinases and phosphatases and the role of second messengers generated via phospholipases in K channel desensitization. The role of muscarinic K current in ACh-induced changes in contractile behavior will be identified.