MECHANISMS OF HEPATIC ELECTROLYTE TRANSPORT

这项提案中描述的研究将调查两个相关的 关于激素调节细胞膜K+通透性的假说； 最近描述的一种内向Na+偶联机制的表征 HCO3-在肝细胞中的转运。这些都是基本的，但很差。 了解直接和广泛影响专业人员的机制 肝脏的功能，包括糖异生、小管胆汁 形成和膜溶质运移。拟议的研究将 利用各种方法，但强调对整个细胞的直接测量 以及具有膜片钳记录技术的单通道电流 以前几乎没有应用于这些或其他方面 肝细胞功能。第一个假设的长期目标 就是定义连接某些激素受体的机制 细胞表面对膜K+(和其他)变化的相互作用 渠道活动，具体目标是：i)进一步评估 CAMP类似物激活膜特异钾电流的机制 特别强调K+的调节磷酸化的作用 通过cAMP依赖的蛋白激酶(PKA)；ii)确定 胰高血糖素和其他与腺苷环化酶连接的激素激活膜 K+电流是否完全通过cAMP或cAMP非依赖性机制 也参与其中；以及iii)调查负责 胰岛素的抑制作用和钙离子的调节作用 和/或钙依赖的钾通道。的长期目标是 第二个假设是确定导致 膜H~+-HCO3-转运与细胞内维持有关 PH和胆汁HCO3-分泌。具体目标是：i)确定 Na+偶联HCO3-内向转运的化学计量及其调控 膜电位差对HCO3-内流的影响 研究这一机制可能的变构调节。 细胞内H+、激酶和某些胆汁酸；以及iii)确定 抑制作用的功能意义和机制 膜上K+通道受H+影响。肝病这些机制的异常 电解质转运直接导致胆汁淤积，细胞 最常见的是肿胀、运输和代谢紊乱 肝病。

The studies described in this proposal will investigate two related hypotheses regarding hormonal regulation of membrane K+ permeability; and characterization of a recently described mechanism for inward Na+-coupled HCO3-transport in hepatocytes. These represent fundamental yet poorly understood mechanisms that directly and broadly influence specialized functions of the liver including gluconeogenesis, canalicular bile formation, and membrane solute transport. The proposed studies will utilize a variety of methods but emphasize direct measurement of whole cell and single channel currents with patch clamp recording techniques which have had little prior application to these or other aspects of hepatocellular function. The long term objective of the first hypothesis is to define the mechanisms which link certain hormone-receptor interactions at the cell surface to changes in membrane K+ (and other) channel activity, and the specific aims are: i) to further evaluate the mechanisms for activation of specific membrane K+ currents by cAMP analogs with particular emphasis on the role of regulatory phosphorylation of K+ channels by cAMP-dependent protein kinases (PKA); ii) to determine whether glucagon and other hormones linked to adenylate cyclase activate membrane K+ currents exclusively through cAMP or whether cAMP-independent mechanisms are also involved; and iii) to investigate the mechanisms responsible for the inhibitory effects of insulin; and the modulatory effects of Ca2+ and/or Ca2+ -dependent kinases on K+ currents. The long term objective of the second hypothesis is to determine the mechanisms responsible for membrane H+HCO3- transport a they pertain to maintenance of intracellular pH and biliary HCO3- secretion. The specific aims are i) to determine the stoichiometry of inward Na+-coupled HCO3- transport and the regulatory influence of membrane potential difference on HCO3- influx; ii) to investigate possible allosteric regulation of this mechanism by intracellular H+, kinases, and certain bile acids; and iii) to determine the functional significance and mechanisms responsible for inhibition of membrane K+ channels by H+. Abnormalities of these mechanisms of hepatic electrolyte transport directly contribute to the cholestasis, cell swelling, and disordered transport and metabolism that accompanies most liver disease.