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ION TRANSPORT IN NERVE AND FIBROBLASTS

神经和成纤维细胞中的离子传输

基本信息

批准号：
6213630
负责人：
JOHN M RUSSELL
金额：
--
依托单位：
SYRACUSE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
1998
资助国家：
美国
起止时间：
1998-11-11 至 2002-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/6213630
关键词：
Xenopus oocyte anions bicarbonates cell morphology cytomegalovirus fibroblasts ion transport membrane transport proteins neurons potassium protein kinase sodium squid virus replication voltage /patch clamp

项目摘要

In this proposal we will study membrane ion transport processes, with particular emphasis on mechanisms involving the chloride anion. We will study not only basic, mechanistic issues related to such ion transport processes, but also issues that relate these ion transport processes to human health and disease. Using the internally dialyzed squid giant axon, which gives direct access to both the inner and outer faces of the axolemma, several important mechanistic issues related to the operation of the Na-K-Cl cotransporter will be addressed. Radioisotopic fluxes and voltage clamping will be used to address these issues. We will test (a) whether high [Cl-]i acts to change the apparent affinity of external and internal binding sites for the cotransported ions; (b) whether the effect of elevated [Cl-]i is due to inhibition of a protein kinase. An examination of possible effects of intracellular K+ on Na-K-Cl cotransporter function will be made. This ion occupies a key spot in the putative order of binding and release. Therefore, a careful examination of its effects will shed considerable light on the actual mechanism of ion binding and release by this cotransporter. Activation by cell shrinkage of this cotransporter will be studied. Voltage-clamping and isotopic fluxes will be combined to examine possible voltage sensitivity of cotransport fluxes. A new direction for our lab is to examine the role of ion transport mechanisms in the development of human fibroblast cell swelling (cytomegaly) in response to infection with the human cytomegalovirus. In particular, we will address the possible role of Na/H exchange and Cl/HCO3 in the development of this pathology. Infection appears to cause the endogenous Na/H exchanger in MRC-5 fibroblasts to become sensitive to cell shrinkage. We will follow up a preliminary finding that suggests a fall of [Cl-]i plays a role in this effect. The effects of the viral infection on the MRC-5 cell's ability to volume regulate will be studied. These studies involve the use of fluorescent probes for ion concentrations and cell volume as well as the use of radioisotopes.

在本提案中，我们将研究膜离子传输过程，其中特别强调涉及氯离子的机制。我们将不仅研究与此类离子传输相关的基本机械问题过程，以及与这些离子传输过程相关的问题人类健康和疾病。使用内部透析的鱿鱼巨轴突，可直接进入对于轴突的内表面和外表面，有几个重要的与Na-K-Cl协同转运蛋白运行相关的机械问题将得到解决。将使用放射性同位素通量和电压钳位来解决这些问题。我们将测试 (a) 高 [Cl-]i 是否会导致改变外部和内部结合位点的表观亲和力共转运离子； (b) [Cl-]i 升高的影响是否是由于抑制蛋白激酶。检查可能产生的影响细胞内 K+ 会发挥 Na-K-Cl 协同转运蛋白的功能。该离子在假定的绑定和释放顺序中占据着关键位置。因此，仔细检查其影响将得出相当大的结论。揭示离子结合和释放的实际机制协同转运蛋白。该协同转运蛋白的细胞收缩将被激活研究过。电压钳位和同位素通量将结合起来检查共转运通量可能的电压敏感性。我们实验室的一个新方向是研究离子传输的作用人成纤维细胞肿胀发生的机制（细胞肿大）对人类巨细胞病毒感染的反应。在特别是，我们将讨论 Na/H 交换和 Cl/HCO3 的可能作用在这种病理学的发展过程中。感染似乎会导致 MRC-5 成纤维细胞中的内源性 Na/H 交换剂对细胞变得敏感收缩。我们将跟进一项表明下降的初步调查结果 [Cl-]i 在这种效应中发挥了作用。病毒感染对人的影响将研究 MRC-5 细胞的容量调节能力。这些研究涉及使用荧光探针来检测离子浓度和细胞体积以及放射性同位素的使用。