NEUTRAL ENDOPEPTIDASE AND CAROTID BODY FUNCTIONS

中性内肽酶和颈动脉体功能

• 批准号: 2392674
• 负责人: Ganesh K Kumar
• 金额: $ 19.1万
• 依托单位: CASE WESTERN RESERVE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1995
• 资助国家: 美国
• 起止时间: 1995-06-07 至 1998-07-06
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2392674
• 关键词: acetylcholinesterase; carotid body; cats; chemical structure function; enkephalins; enzyme activity; hemorrhagic shock; high performance liquid chromatography; hypercapnia; hyperoxia; hypoxia; immunoelectron microscopy; laboratory rat; microspectrophotometry; molecular weight; neprilysin; radioimmunoassay; superior cervical ganglion; sympathectomy; western blottings

The carotid bodies are peripheral chemoreceptor organs. They transduce changes in the levels of arterial oxygen, carbon dioxide, and hydrogen ion concentrations into action potential encoded nerve signals. This transduction process is believed to occur through a complex mechanism, mediated by a variety of neurochemicals such as biogenic amines, neuropeptides and gas molecules like nitric oxide. Our studies on the metabolism of neuropeptides in the carotid body have shown that neutral endopeptidase (NEP) plays a major role in the regulation of neuropeptides like tachykinins and enkephalins in the carotid body. Also, we have found that NEP in the carotid body occurs in two active forms viz., a less common cytosolic and conventional membrane-bound form in near equal proportions. Furthermore, inhibition of the NEP activity in vivo appears to potentiate the response of the carotid body to hypoxia suggesting that the two NEP forms may play hitherto undefined important functional roles in the carotid body. This investigation, specifically, addresses the molecular characterization of the two NEP forms of the carotid body and their importance in the function(s) of the carotid body. Our hypothesis is that physiological stimulus such as hypoxia alters the activity and/or synthesis of NEP. These changes in NEP, in turn, regulate the concentrations and modulate the actions of both the excitatory and inhibitory neuropeptides and thus determine the magnitude of chemosensory responses of the carotid body to either acute and/or chronic hypoxia. This hypothesis will be tested using biochemical and cellular biological techniques in combination with immunological and immunocytochemical studies on the two forms of NEP of the carotid body. Most of the experiments will use carotid bodies obtained from cats and rats exposed to defined gas challenges. Experiments outlined in Specific Aim 1 will address the physico-chemical characterization of the soluble and membrane- bound forms of NEP of the carotid body and evaluate the structural relationship among the two NEP forms. The kinetic experiments proposed in Specific Aim 2 will examine the possibility that the NEP forms of the carotid body have different enzymological properties. In Specific Aim 3, using immunocytochemical approaches, we will determine the cellular location of NEP in relation to the location of tachykinins, enkephalins and atrial natriuretic peptides in the carotid body. The potential alterations in NEP forms of the carotid body in response to different gas challenges and possible mechanisms involved in these alterations will be addressed in Specific Aim 4. It is anticipated that the proposed studies will provide clues as to the role of NEP in the functions of the carotid body. These studies may also yield information in the development of enzyme and its inhibitor(s) based therapeutic agents in the treatment of hypoxemia, a major problem associated with chronic obstructive lung diseases and high altitude adaptation.

颈动脉体是外周化学感受器。 他们转换 动脉血氧、二氧化碳和氢离子水平的变化 浓度转化为动作电位编码的神经信号。 这 转导过程被认为通过复杂的机制发生， 由多种神经化学物质如生物胺介导， 神经肽和气体分子如一氧化氮。 我们的研究 颈动脉体中神经肽的代谢已经表明， 内肽酶（NEP）在神经肽的调节中起主要作用 如颈动脉体中的速激肽和脑啡肽。 同时，我们发现 颈动脉体中的NEP以两种活性形式发生，一个不那么 常见的胞质和传统的膜结合形式几乎相等 比例。 此外，体内NEP活性的抑制出现 增强颈动脉体对缺氧的反应， 这两种形式的新经济政策可能发挥着迄今为止尚未明确的重要功能作用 在颈动脉体。 这项调查，具体来说，解决了 颈动脉体的两种NEP形式的分子表征， 它们在颈动脉体功能中的重要性。 我们的假设是，生理刺激，如缺氧，改变了 NEP的活性和/或合成。 NEP的这些变化反过来又调节了 浓度和调节兴奋性和 抑制性神经肽，从而决定化学感受的大小 颈动脉体对急性和/或慢性缺氧的反应。 这一假设将使用生物化学和细胞生物学方法进行检验。 免疫学和免疫细胞化学联合技术 颈动脉体两种形式NEP的研究。 大部分 实验将使用从猫和大鼠中获得的颈动脉体， 天然气挑战 具体目标1中概述的实验将 解决可溶性和膜的物理化学表征- 颈动脉体NEP的结合形式，并评估结构 两种新经济政策的关系。 提出的动力学实验 具体目标2将审查新经济政策形式的可能性， 颈动脉体具有不同的酶学性质。 在具体目标3中， 使用免疫细胞化学方法，我们将确定细胞 NEP的位置与速激肽、脑啡肽 和心钠素。 的潜在 颈动脉体对不同气体的反应性NEP形式的改变 这些变化所涉及的挑战和可能的机制将是 具体目标4。 预计拟议的研究 将为NEP在颈动脉功能中的作用提供线索 身体 这些研究也可能在发展中产生信息， 基于酶及其抑制剂的治疗剂 低氧血症是慢性阻塞性肺的主要问题 疾病和高海拔适应。