DIABETIC NEUROPATHY--RECEPTORS AND SIGNALING MECHANISM

糖尿病神经病——受体和信号传导机制

• 批准号: 3464493
• 负责人: LILIANA N BERTI-MATTERA
• 金额: $ 9.89万
• 依托单位: CASE WESTERN RESERVE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1992
• 资助国家: 美国
• 起止时间: 1992-09-30 至 1997-09-29
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3464493
• 关键词: G protein; SDS polyacrylamide gel electrophoresis; Schwann cells; adenosinetriphosphatase; adenylate cyclase; adrenergic receptor; biological signal transduction; bradykinin; cell line; cyclic AMP; diabetic neuropathy; glucose; immunochemistry; laboratory rat; membrane transport proteins; muscarinic receptor; neuropeptide receptor; phospholipase C; potassium; receptor coupling; sodium; stimulant /agonist; tissue /cell culture; western blottings

Diabetic neuropathy, one of the major complications of chronic diabetes is characterized by several biochemical alterations including a reduction in myo-inositol levels, decrease in Na+K+-ATPase activity and abnormalities in phosphoinositide (PPI) metabolism and adenylyl cyclase activity. The current hypothesis regarding the pathophysiology of diabetic neuropathy assign a central role to the decrease in myo-inositol uptake-dependent phosphoinositide turnover and hence availability of second messengers, like diacylglycerol which modulate Na+K+-ATPase activity through activation of protein kinase C. While it is generally accepted that the reduction in Na+K+-ATPase plays a key role in the development of neuropathy, its precise regulation is far from clear. Recent evidence from our and other labs suggests that peripheral nerve possesses muscarinic, bradikynin, purinergic and beta-adrenergic receptors which seem to be linked to stimulation of the breakdown of PPI or adenylyl cyclase. We have recently identified several types of guanine-nucleotide binding (G) proteins, that are known to be involved in other tissues in coupling between the agonist-receptor complexes and the adenylyl cyclase or phospholipase C systems. In nerve and myelin membranes from experimental diabetic animals we demonstrated that some of these signalling pathways are altered. With this evidence, we propose to: 1) characterize the relative levels of G-proteins in total and subcellular fractions of sciatic nerve from diabetic rats by ADP- ribosylation with bacterial toxins and immunoblotting; 2) investigate the receptor-linked signalling pathways that modulate the activity of phospholipase C and adenylyl cyclase and their influence on Na+K+-ATPase activity in sciatic nerve slices or membranes by: a)activation of phospholipase C through muscarinic, purinergic and bradikynin receptors b) modulation of adenylyl cyclase activity through activation of beta- adrenergic and muscarinic receptors: 3) evaluate how the alterations in Na+K+-ATPase in nerves from diabetic animals correlate with changes in the different components of the above mentioned signalling pathway and 4) determine if in cultured Schwann cells the above mentioned agonists are linked to cAMP and phospholipase C and if these pathways are altered under conditions that mimic diabetes like high glucose and myo-inositol free culture conditions.

糖尿病神经病变，慢性糖尿病的主要并发症之一 以几种生化变化为特征，包括减少 在肌醇水平上，Na+K+-ATPase活性降低和 磷脂酰肌醇(PPI)代谢异常与腺苷环化酶 活动。目前关于心绞痛的病理生理学假说 糖尿病神经病变在肌醇减少中起中心作用 依赖摄取的磷脂酰肌醇周转率和因此可利用性 第二信使，如调节Na+K+-ATPase的甘油二酯 通过激活蛋白激酶C的活性，而它通常是 认为Na+K+-ATPase的降低在 神经病变的发生、发展，其确切的调控机制还远不清楚。 来自我们和其他实验室的最新证据表明，周围神经 具有毒扁豆碱、布拉吉宁、嘌呤能和β-肾上腺素能 似乎与刺激PPI分解有关的受体 或者是腺苷环化酶。我们最近发现了几种类型的 鸟嘌呤核苷酸结合(G)蛋白，已知参与 在激动剂-受体复合体之间耦合的其他组织中 腺酰环化酶或磷脂酶C系统。在神经和髓鞘中 来自实验性糖尿病动物的膜我们证明了一些 这些信号通路中的一部分被改变了。有了这个证据，我们建议 为：1)表征G-蛋白在总和 ADP法测定糖尿病大鼠坐骨神经亚细胞组分 细菌毒素的核糖化和免疫印迹；2)研究 受体连接的信号通路，调节细胞的活性 磷脂酶C和腺苷环化酶及其对Na~+-K~+-ATPase的影响 坐骨神经薄片或神经膜的活动通过：a)激活 磷脂酶C通过M受体、嘌呤能受体和BRadikynin受体 B)通过激活β-β-来调节腺苷环化酶的活性 肾上腺素能和毒扁豆碱能受体：3)评估 糖尿病动物神经中Na+K+-ATPase与 上述信号通路的不同组成部分和 4)测定培养的雪旺细胞中上述激动剂是否 与cAMP和磷脂酶C有关，如果这些途径发生改变 在类似糖尿病的条件下，如高血糖和肌醇 自由培养条件。