MECHANISMS OF HEPATIC ELECTROLYTE TRANSPORT

这项提案中描述的研究将调查两个相关的 关于激素调节膜离子通透性的假说， 以及最近描述的Na+/HCO3-机理的表征。 肝细胞内的共转运。这些都是基本的，但很差。 了解直接和广泛影响许多人的机制 肝脏的特殊功能，包括糖异生、胆汁 形成和膜溶质运移。拟议的研究将 利用多种方法，但强调对整体的直接测量 采用膜片钳记录技术的细胞和单通道电流 它们以前几乎不适用于这些或其他方面 肝细胞功能。第一个假设的长期目标 是定义荷尔蒙-受体相互作用与 膜K+和其他离子通道活性的调节变化，以及 具体目标是：i)评估膜电位的变化 差异性和离子渗透性表征活性 与钙离子动员和cAMP生成有关的受体；ii) 确定负责cAMP和胰高血糖素的单个K+通道 K+外流的依赖性增加；评估CaM- 通道激活中的依赖蛋白激酶和GTP结合蛋白， 并探讨其抑制作用的机制。 胰岛素；iii)评估刺激和抑制相互作用 钙离子和蛋白激酶C在钙离子通透性阳离子调节中的作用 4)评价肝受体的补体(S) 被细胞外三磷酸腺苷激活。第二个方面的长期目标 评估Na+/HCO3-共转运在调节血管紧张素转换酶中作用的假说 膜酸/碱转运，具体目的是：1)测定 Na+/HCO3-共转运的化学计量学，II)评估 膜电位差与细胞内H、Ca~(2+)和激酶的关系 作为潜在的监管因素；以及iii)评估这些机制 有助于膜电位的Phi依赖的调制 通过测量[H+]对K+开放的影响而得出的差异 和其他渠道。这些机制的异常导致了 直接和重要的是胆汁淤积、细胞肿胀和 伴随大多数肝病的运输和新陈代谢紊乱。

The studies described in this proposal will investigate two related hypotheses regarding hormonal regulation of membrane ion permeability, and characterization of a recently described mechanism for Na+/HCO3- cotransport in hepatocytes. These represent fundamental yet poorly understood mechanisms that directly and broadly influence many specialized functions of the liver including gluconeogenesis, 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 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 hormone-receptor interactions to regulated changes in membrane K+ and other ion channel activity, and the Specific Aims are: i) to evaluate the changes in membrane potential difference and ion permeability which characterize activation of receptors coupled to Ca2+ mobilization and cAMP generation; ii) to identify individual K+ channels responsible for cAMP- and glucagon- dependent increases in K+ efflux; assess the relative roles of cAM- dependent protein kinases and GTP-binding proteins in channel activation, and investigate the mechanisms responsible for the inhibitory effects of insulin; iii) to evaluate stimulatory and inhibitory interactions between Ca2+ and protein kinase C in regulation of Ca2+ permeable cation channels; and iv) to evaluate the complement of hepatic receptor(s) activated by extracellular ATP. The long term objective of the second hypothesis to assess the role of Na+/HCO3-cotransport in regulation of membrane acid/base transport, and the Specific Aims are i) to determine the stoichiometry of Na+/HCO3-cotransport, ii) assess the roles of membrane potential difference and of intracellular H, Ca2+, and kinases as potential regulatory factors; and iii) to evaluate the mechanisms which contribute to pHi-dependent modulation of membrane potential difference through measurement of the effects of [H+] on opening of K+ and other channels. Abnormalities of these mechanisms contribute directly and importantly to the cholestasis, cell swelling, and disordered transport and metabolism that accompany most liver diseases.