NEUTRAL ENDOPEPTIDASE AND CAROTID BODY FUNCTIONS

中性内肽酶和颈动脉体功能

• 批准号: 2222945
• 负责人: Ganesh K Kumar
• 金额: $ 17.43万
• 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 1998-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2222945
• 关键词: acetylcholinesterase; carotid body; cats; chemical structure function; endopeptidases; enkephalins; enzyme activity; hemorrhagic shock; high performance liquid chromatography; hypercapnia; hyperoxia; hypoxia; immunoelectron microscopy; laboratory rat; microspectrophotometry; molecular weight; 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器官。 他们转换 动脉氧、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 据信转导过程是通过复杂的机制发生的， 由多种神经化学物质介导，例如生物胺， 神经肽和气体分子，如一氧化氮。 我们的研究关于 颈动脉体中神经肽的代谢表明，中性 肽链内切酶（NEP）在神经肽的调节中发挥着重要作用 例如颈动脉体中的速激肽和脑啡肽。 另外，我们还发现 颈动脉体中的 NEP 以两种活跃形式发生，即，一种较少 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 活性的抑制似乎 增强颈动脉体对缺氧的反应表明 这两种 NEP 形式可能发挥迄今为止尚未明确的重要功能作用 在颈动脉体中。 这项调查特别针对 颈动脉体两种 NEP 形式的分子特征 它们在颈动脉体功能中的重要性。 我们的假设是缺氧等生理刺激会改变 NEP的活性和/或合成。 NEP 的这些变化反过来又调节 浓度并调节兴奋性和 抑制性神经肽，从而决定化学感应的强度 颈动脉体对急性和/或慢性缺氧的反应。 该假设将使用生化和细胞生物学进行检验 结合免疫学和免疫细胞化学技术 颈动脉体两种形式 NEP 的研究。 大部分的 实验将使用从接触过的猫和老鼠身上获得的颈动脉体 明确的天然气挑战。 具体目标 1 中概述的实验将 解决可溶性和膜性物质的物理化学特征 颈动脉体 NEP 的结合形式并评估结构 两种 NEP 形式之间的关系。 中提出的动力学实验 具体目标 2 将检验 NEP 形成的可能性 颈动脉体具有不同的酶学特性。 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、脑啡肽的位置相关 和颈动脉体中的心房钠尿肽。 潜力 颈动脉体 NEP 形式响应不同气体的变化 这些改变涉及的挑战和可能的机制将是 具体目标 4 中提出。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。 这些研究还可能为开发提供信息 基于酶及其抑制剂的治疗药物 低氧血症是与慢性阻塞性肺相关的一个主要问题 疾病和高海拔适应。