METABOLIC REGULATION OF CELLULAR POTASSIUM BALANCE

在急性心肌缺血期间，细胞外K积累是心肌缺血的主要原因。 诱发心脏发生折返性心律失常的因素 和VF。 然而，这一机制仍存在争议。 两大 假设是：激活代谢敏感K通道， 作为ATP敏感性K（KATP）通道，以及与阴离子流出偶联的K损失 （乳酸盐和/或Pi）作为电荷平衡机制。 的主要目标 本项目旨在进一步阐明KATP通道在 缺氧和缺血期间细胞K损失，以表征 KATP通道的生物物理、调节和药理学特性 更详细，并评估经肌膜乳酸的机制 运动及其与心脏阳离子流的关系。 的影响 KATP通道对细胞钾流失的激活将在 离体动脉灌注兔室间隔， 43 K测量组织钾含量的单向钾流出率， 暴露于KATP通道激动剂。 我们的初步调查结果显示 用色满卡林选择性激活KATP通道， 潜在缩短和单向钾流出率增加 类似于缺氧，但不引起净钾损失。 拟议 实验中，我们将激活的假设，除了 激活KATP通道，需要增强内向电流 导致钾的净流失。 在兔隔中的实验结果将 也可以在心室动作电位的计算机模型中模拟 提供更多的见解。 我们将协助调查 KATP通道的生物物理、调节和药理学特性， 在离体心室肌细胞中使用膜片钳技术。 我们将 试图描述糖酵解优先 调节KATP通道活性。 我们将测试一个新的假设， 表面电荷在调节ATP中起重要的生理作用， KATP通道的敏感性。 我们将探索我们的观察，c Ca- 严重代谢抑制过程中的依赖过程 改变了KATP通道的ATP敏感性， 用胰蛋白酶对切除的由内而外的膜贴片的胞质表面进行检测， 其他代理人模仿这种效果。 这些观察将是 进一步研究，以了解 病理生理条件，并深入了解如何蛋白水解 KATP通道的化学修饰改变了功能。 最终 主要目的是评估跨膜乳酸的机制 心脏运动及其与阳离子（特别是K）通量的关系。 我们开发了一种研究跨膜乳酸的新方法 用荧光法观察离体膜片钳心肌细胞的运动 用于监测细胞内H、K和Na对乳酸的响应的指示剂 为达到这一目的。 这些研究将提供重要的新 深入了解一个主要致瘤因素的机制， 细胞外钾积累，导致急性期猝死 心肌缺血

During acute myocardial ischemia, extracellular K accumulation is a major factor predisposing the heart to the development of reentrant arrhythmias and VF. The mechanism, however, remains controversial. Two major hypotheses are: activation of metabolically-sensitive K channels such as ATP-sensitive K(KATP) channels, and K loss coupled to anion efflux (lactate and/or Pi) as a charge-balancing mechanism. The major goals of this project are to further elucidate the role of KATP channels in cellular K loss during hypoxia and ischemia, to characterize the biophysical, regulatory and pharmacologic properties of KATP channels in greater detail, and to evaluate mechanisms of transsarcolemmal lactate movement and its relationship to cation fluxes in heart. The effects of activation of KATP channels on cellular K loss will be studied in isolated arterially perfused rabbit interventricular septa loaded with 43K to measure unidirectional K efflux rate of tissue K content during exposure to KATP channel agonists. Our preliminary findings indicate that selective activation of KATP channels with cromakalim caused action potential shortening and an increase in unidirectional K efflux rate similar to hypoxia, but did not cause net K loss. In the proposed experiments we will activate the hypothesis that in addition to activation of KATP channels, enhancement of inward currents is required for net K loss to occur. Experimental findings in the rabbit septum will also be simulated in a computer model of the ventricular action potential to provide further insights. We will contribute to investigate the biophysical, regulatory and pharmacologic properties of KATP channels, using patch clamp techniques in isolated ventricular myocytes. We will attempt to delineate the mechanism by which glycolysis preferentially regulates KATP channel activity. We will test a novel hypothesis that surface charge plays an important physiologic role in regulating the ATP- sensitivity of KATP channels. We will explore our observation that c Ca- dependent process during severe metabolic inhibition irreversibly modified the ATP-sensitivity of KATP channels, and that treatment of the cytosolic surface of excised inside-out membrane patches with trypsin and other agents mimicked this effect. These observations will be investigated further to provide insight into channel regulation under pathophysiological conditions, and to gain insight into how proteolysis and chemical modification of KATP channels alters function. The final major goal is to evaluate the mechanisms of transmembrane lactate movement in heart and its relationship to cation (particularly K) fluxes. We have developed a novel method for studying transmembrane lactate movement in isolated patch-clamped cardiac myocytes using fluorescent indicators to monitor intracellular H, K and Na in response to lactate influx for this purpose. These studies should provide important new insights into the mechanisms of a major arrhythmogenic factor, extracellular K accumulation, contributing to sudden death during acute myocardial ischemia.