G PROTEIN COUPLED ION CHANNELS IN HEART RATE REGULATION

心脏病仍然是世界上发病率和死亡率的主要原因。 美国老年人口。 心率下降， 心律失常随年龄增长而增加。 然而，心率的分子基础 控制仍然不清楚。 超极化激活电流I（f） 和T型钙电流I（Ca-T）是两个重要的贡献者， 舒张期去极化，其设定心脏的速率和节律， 心脏起搏 在斑马鱼（心脏发育的新模型）中， 导致基础心率减慢的原因已经被证明 一（f）.心率的神经控制由G蛋白偶联介导， 受体。 交感神经（通过β-肾上腺素能受体通过 Gs）和副交感神经（通过m2-毒蕈碱受体， 百日咳毒素敏感的Gi/Go）对心脏产生相反的作用， 功能 我们已经产生了三种基因敲除细胞系和动物 在心脏α（i2）α（i3）中表达的Gi/Go α亚基， alpha（0）。我们建议用这些击倒来理解信号 转导途径和这些控制的分子基础 两个离子通道，并利用这些知识来了解 衰老的心脏 结合我最近获得的经验 在电生理学和生物物理学方面有额外的经验， 分子生物学，我会在这个奖项期间获得，我将是 在研究G蛋白调节的机制方面处于独特的地位， 起搏活动从细胞电生理事件到其 功能性后果。 本提案的具体目标是： 肾上腺素能和胆碱能控制舒张期去极化的基础 起搏器活动。 目标2：确定 Gi/Go异源三聚体蛋白介导I（f）的胆碱能调节 和I（Ca-T），以及调节是否是直接的斑块限制或 通过间接途径。 目标3：确定信号 转导途径介导离子通道的控制：3A. 确定一氧化氮是否介导肾上腺素能和胆碱能 调节离子通道和心率。3B.以确定是否G- 蛋白质信号由β-γ或α亚基介导。 目标4： 确定α亚单位基因失活对基础免疫应答的影响。 心率与衰老的关系，并将这些变化与G- 使用WT和敲除小鼠的蛋白水平。 这些实验的结果将增加我们对正常的 信号转导途径和改变G蛋白的影响 心率调节与衰老的关系。

Heart disease remains a major cause of morbidity and mortality in the U.S. older population. Heart rate declines and the incidence arrhythmias increase with age. Yet, the molecular basis for heart rate control remains unclear. The hyperpolarization activated current I(f) and T-type calcium current I(Ca-T) are two important contributors to the diastolic depolarization which sets the rate and rhythm of the cardiac pacemaking. In zebrafish (a ne model of cardiac development) mutations which result in slow basal heart rate have been shown to have altered I(f). The neural control of heart rate is mediated by G-protein coupled receptors. Sympathetic (via beta-adrenergic receptors signaling through Gs) and parasympathetic (via m2-muscarinic receptors that signal through pertussis toxin sensitive Gi/Go) produce opposing effects on cardiac function. We have produced knockout cell lines and animals of the three Gi/Go alpha subunits expressed in heart alpha(i2) alpha(i3) and alpha(0). We propose to use these knockouts to understand the signal transduction pathways and the molecular basis of the control of these two ion channels and to use that knowledge to understand the effects in the aging heart. Combining the experience that I have recently obtained in electrophysiology and biophysics with the additional experience in molecular biology that I would gain during this award period, I will be in an unique position to study the mechanism of G-protein regulation of pacemaking activity from cellular electrophysiological events to their functional consequences. The specific aims for this proposal are: Aim 1: To determine the ionic basis for adrenergic and cholinergic control of diastolic depolarization in pacemaker activity. Aim 2: To determine which Gi/Go heterotrimeric protein mediates the cholinergic regulation of I(f) and I(Ca-T), and whether the regulation is direct patch limited or through an indirect pathway. Aim 3: To determine the signal transduction pathway mediating control of ion channels: 3A. To determine whether nitric oxide mediates adrenergic and cholinergic regulation of ion channels and heart rate. 3B. To determine if the G- protein signal is mediated by beta-gamma or alpha subunits. Aim 4: To determine the effect of the alpha subunit gene inactivations on basal heart rate with aging and correlate these changes with changes in G- protein levels using WT and knockout mice. Results from these experiments will add to our knowledge on the normal signal transduction pathways and the impact of altered G protein expression on heart rate regulation with aging.