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.

颈动脉小体是外周化学感受器。他们转译成 动脉血氧、二氧化碳和氢离子水平的变化 浓缩成动作电位编码神经信号。这 转导过程被认为是通过一个复杂的机制发生的， 由生物胺等多种神经化学物质介导， 神经肽和气体分子，比如一氧化氮。我们对生物多样性的研究 颈动脉小体中神经肽的代谢显示中性 内肽酶(NEP)在神经肽的调节中起重要作用 比如颈动脉小体中的速激肽和脑啡肽。另外，我们还发现 颈动脉小体中的NEP以两种活跃形式出现，即较少的 常见的胞质结合形式和常规膜结合形式接近相等 比例。此外，体内NEP活性出现抑制。 增强颈动脉小体对低氧的反应 这两种NEP形式可能发挥迄今未定义的重要功能作用 在颈动脉小体。这项调查，特别是针对 颈动脉小体和颈总动脉两种NEP类型的分子特征 它们在颈动脉小体功能中的重要性(S)。 我们的假设是，像缺氧这样的生理刺激改变了 NEP的活性和/或合成。NEP中的这些变化反过来又调节 兴奋性和调节性神经递质的浓度和作用 抑制神经肽，从而决定化学感觉的大小 颈动脉小体对急性和/或慢性缺氧的反应。 这一假设将通过生物化学和细胞生物学进行检验。 免疫学和免疫细胞化学相结合的技术 颈动脉小体NEP的两种形态研究。大多数 实验将使用从猫和大鼠身上获得的颈动脉体 明确的气体挑战。具体目标1中概述的实验将 解决了可溶性和膜的物理化学表征- 颈动脉小体NEP的结合形式及结构评价 两种NEP表格之间的关系。中提出的动力学实验 具体目标2将审查NEP形成的可能性 颈动脉小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定位与速激肽、脑啡肽定位的关系 颈动脉小体内的心钠素。潜力 不同气体对颈动脉小体NEP形态的影响 这些改变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 尸体。这些研究也可能产生关于发展的信息 以酶及其抑制物(S)为基础的药物治疗慢性粒细胞白血病 低氧血症是慢性阻塞性肺的主要问题 疾病和高原适应。