NA+-K+-2CL- COTRANSPORTERS IN CEREBRAL ISCHEMIA

脑缺血中的 NA -K -2CL- 协同转运蛋白

• 批准号: 6499424
• 负责人: Dandan Sun
• 金额: $ 19.74万
• 依托单位: UNIVERSITY OF WISCONSIN-MADISON
• 依托单位国家: 美国
• 财政年份: 2000
• 资助国家: 美国
• 起止时间: 2000-02-01 至 2003-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6499424
• 关键词: astrocytes; bumetanide; calcium flux; cerebral ischemia /hypoxia; chloride ion; glutamates; ion transport; laboratory rat; membrane transport proteins; neurotoxicology; potassium ion; sodium ion; tissue /cell culture

nA+-k+-2Cl-cotransporters (NKCC) are important in renal salt reabsorption and secretion by reabsorptive and secretory epithelia. The NKCC also function in maintenance and regulation of cell volume in both epithelial and non-epithelial cells. However, the role of the NKCC in the central nervous system (CNS) have not been defined. The long-term goal of this research is to understand function and regulation of the NKCC in the CNS under physiological as well as pathological conditions such as ischemia. Our pilot studies described in this proposal suggest that the NKCC are essential in maintenance of K+ homeostasis in astrocytes and they also appear to be involved in glutamate (Glu)-mediated neurotoxicity. There are four Specific Aims to test our hpotheses. Aim 1: Characterize high [K+]o-mediated stimulation of the NKCC activity in primary astrocyte cultures. The objectives are: 1). to verify that high [K+]o causes stimulation of the NKCC; 2). to determine whether the high [K+]o mediated stimulation of the NKCC is due to an increase in intracellular [Ca++]. Aim 2: Determine whether high [K+]o- mediated stimulation of the NKCC causes astrocyte swelling and non-vesicular release of Glu. The objectives are to investigate: 1). whether the NKCC contributes to accumulation of intracellular Na+, C1 and cell swelling under [K+]o; 2). whether inhibition of the NKCC activity could significantly increase [3H]-L-Glu uptake, while blocking of high [K+]-evoked release of Glu from astrocytes. Aim 3: Investigate whether stimulation of the NKCC contributes to Glu-mediated neuronal toxicity. The objectives are to investigate: 1). whether the NKCC contributes to Glu- mediated increase in [C1-]i and neuronal swelling; 2). whether inhibition of the NKCC activity could protect cells from Glu- induced damage. Aim 4: Determine whether ischemia-induced cell damage is reduced when the NKCC activity is transiently inhibited by a local administration of bumetanide in brain. This study will be performed in an in vivo cerebral ischemic model.

nA+-k+- 2cl共转运体（NKCC）在肾盐的重吸收和分泌中起重要作用。NKCC还在上皮细胞和非上皮细胞的细胞体积的维持和调节中起作用。然而，NKCC在中枢神经系统（CNS）中的作用尚未明确。本研究的长期目标是了解NKCC在生理和病理条件下（如缺血）在中枢神经系统中的功能和调控。我们在本提案中描述的初步研究表明，NKCC在维持星形胶质细胞中的K+稳态中是必不可少的，它们似乎也参与谷氨酸（Glu）介导的神经毒性。有四个具体目标来检验我们的假设。目的1：表征原代星形胶质细胞培养中高[K+]o介导的NKCC活性刺激。目标是：1)验证高[K+]o引起NKCC的刺激；2).确定高[K+]o介导的NKCC刺激是否是由于细胞内[Ca++]的增加。目的2：确定高[K+] 0介导的NKCC刺激是否会导致星形胶质细胞肿胀和非水疱性谷氨酸释放。目的是研究：1)NKCC是否有助于[K+] 0下细胞内Na+、C1的积累和细胞肿胀；2)抑制NKCC活性是否能显著增加[3H]-L-Glu摄取，同时阻断[K+]高诱发的星形胶质细胞谷氨酸释放。目的3：研究NKCC的刺激是否有助于葡萄糖介导的神经元毒性。目的是研究：1)NKCC是否参与Glu介导的[C1-]i升高和神经元肿胀；2)抑制NKCC活性是否能保护细胞免受Glu诱导的损伤。目的4：确定当局部给药布美他尼暂时抑制NKCC活性时，脑缺血诱导的细胞损伤是否减少。本研究将在体内脑缺血模型中进行。