METABOLIC REGULATION OF CELLULAR POTASSIUM BALANCE

在急性心肌缺血期间，细胞外钾的积聚是主要的 心脏易患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 假说是：新陈代谢敏感的钾通道激活 作为ATP敏感的K(KATP)通道，以及与阴离子外流耦合的K损失 (乳酸盐和/或PI)作为电荷平衡机制。的主要目标 本研究旨在进一步阐明KATP通道在细胞内的作用。 在缺氧和缺血过程中细胞钾丢失，以表征 KATP通道的生物物理、调节和药理学特性 更详细，并评估跨肌膜乳酸的机制 心脏运动及其与阳离子流量的关系。的影响 KATP通道激活对细胞钾丢失的影响将在#年进行研究 兔离体室间隔动脉灌流 43K测量组织钾含量的单向钾流出速率 暴露于KATP通道激动剂。我们的初步调查结果表明 克罗卡林选择性激活KATP通道引起的作用 电位缩短和单向钾流出速率的增加 与低氧相似，但不会造成钾的净损失。在建议的 实验中我们将激活这样的假设：除了 激活KATP通道，需要增强内向电流 发生净K损失。兔鼻中隔的实验结果将 也可以在计算机模型中模拟出心室动作电位 以提供进一步的见解。我们将为调查 KATP通道的生物物理、调节和药理学特性， 膜片钳技术在分离的心肌细胞上的应用。我们会 试图描绘糖酵解优先发生的机制 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--中起着重要的生理作用 KATP通道的敏感性。我们将探讨我们的观察结果，即C-Ca- 严重代谢抑制期间的依赖过程不可逆转 改善KATP通道对ATP的敏感性，并治疗 胰酶和胰酶标记的外膜贴片胞液表面 其他代理人也模仿了这种效果。这些观察结果将是 进一步调查，为渠道监管提供洞察 病理生理条件，并深入了解蛋白质分解是如何 而KATP通道的化学修饰改变了功能。决赛 主要目的是评价跨膜乳酸盐的作用机制。 心脏的运动及其与阳离子(尤其是钾)通量的关系。 我们开发了一种研究跨膜乳酸盐的新方法。 用荧光法研究膜片钳心肌细胞的运动 乳酸对细胞内H、K、Na的监测指标 为此目的而涌入。这些研究应该提供重要的新的 对一种主要致心律失常因素的机制的洞察， 细胞外钾积聚，导致急性猝死 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.