NEURAL & PHARMACOLOGICAL CONTROL OF AIRWAY SMOOTH MUSCLE

神经元

基本信息

批准号：
3361051
负责人：
RICHARD W MITCHELL
金额：
$ 10.82万
依托单位：
UNIVERSITY OF CHICAGO
依托单位国家：
美国
项目类别：
财政年份：
1989
资助国家：
美国
起止时间：
1989-08-01 至 1993-07-31
项目状态：
已结题

项目摘要

The purpose of these investigations is to determine the modulating influences of mediators of anaphylaxis on parasympathetic activation in tracheal smooth muscle. Previous investigations have shown that atropine reduced the in vitro sensitivity of ragweed-sensitized tracheal smooth muscle to histamine and high-potassium induced tone. Spontaneous contractions, seen only in sensitized tracheal muscles strips, also were atropine-sensitive and spontaneous phasic contractile activity could be mimicked in muscle strips by addition of small concentrations of acetylcholine or by inhibiting acetylcholinesterase. These studies indicated possible presynaptic augmentation of acetylcholine release and/or reduced cholinesterase activity. We propose to study the parasympathetic innervation of airways with the specific aim of determining the role of histamine, serotonin, and products of arachidonic acid metabolism (prostanoids and leukotrienes) in the release of the neurotransmitter, acetylcholine. It is hypothesized that antigenic sensitization augments tracheal smooth muscle response to acetylcholine. Two mechanism for this effect are hypothesized: 1) increased basal/stimulated release of neurotransmitter and 2) decreased cholinesterase activity resulting from immune-sensitization. All studies will be conducted using tracheal smooth muscle from which the epithelium has been removed to eliminate confounding effects of epithelium-derived relaxing factors. We will 1) determine the normal release characteristics of neurotransmitter (both basal and neurally-activated release of acetylcholine 2) determine whether agents such as histamine, serotonin, prostaglandin F2a and leukotriene D4 augment basal and neurally activated release of acetylcholine 3) compare these date with neurotransmitter release from a sensitized population 4) determine basal acetylcholinesterase activities in homogenates of normal and sensitized tracheal smooth muscle, and 5) determine if agents of anaphylaxis have a modifying effect on cholinesterase activity. We will employ 14C-labeled choline as a precursor of acetylcholine and promote its uptake and turnover by electrical field stimulation in the presence of an incubating concentration of 14C-choline. The Basal efflux of 14C will be measured by liquid scintillation. Augmentation of acetylcholine release will be assessed after incubation with mediators (above) and comparisons will be made between control and sensitized populations. Cholinesterase activities will be measured using spectrophotometric techniques. In addition, total protein and non-collagen protein assays will be performed to enable comparisons to be made between different populations and among different ages. Protein estimations may in themselves reveal differences between normal and sensitized populations. Data derived from these studies will suggest factors influencing modulation of airway smooth muscle tone in vitro and have a direct application to therapeutic interventions in obstructive airways disease and asthma.

这些研究的目的是确定调节过敏反应介质对副交感神经激活的影响气管平滑肌。先前的研究表明阿托品降低豚草致敏气管平滑肌的体外敏感性肌肉以组胺和高钾诱导张力。自发的仅在致敏的气管肌肉条中可见的收缩也阿托品敏感和自发的阶段性收缩活动可能是通过添加小浓度的肌肉条来模仿乙酰胆碱或通过抑制乙酰胆碱酯酶。这些研究表明突触前乙酰胆碱释放可能增加和/或胆碱酯酶活性降低。我们建议研究气道的副交感神经支配，其具体目的是确定组胺、血清素和花生四烯酸产物的作用酸代谢（前列腺素和白三烯）的释放神经递质，乙酰胆碱。据推测，抗原致敏增强气管平滑肌对乙酰胆碱的反应。假设产生这种效应的两种机制：1) 增加神经递质的基础/刺激释放和 2) 减少免疫致敏导致的胆碱酯酶活性。所有研究将使用气管平滑肌进行，上皮细胞来自气管平滑肌已被删除以消除上皮来源的混杂影响放松因素。我们将 1) 确定正常释放特性神经递质（基础释放和神经激活释放）乙酰胆碱 2) 确定组胺、血清素等药物是否前列腺素 F2a 和白三烯 D4 增强基础和神经激活乙酰胆碱的释放 3) 将这些数据与神经递质进行比较敏感人群的释放量 4) 确定基础值正常和致敏匀浆中的乙酰胆碱酯酶活性气管平滑肌，以及 5) 确定过敏反应因子是否具有对胆碱酯酶活性的调节作用。我们将采用 14C 标签胆碱作为乙酰胆碱的前体并促进其吸收和在存在孵化器的情况下通过电场刺激进行周转 14C-胆碱的浓度。 14C 的基础流出量将通过以下方式测量液体闪烁。乙酰胆碱释放增加与介质孵育后进行评估（见上文），并将进行比较在控制人群和敏感人群之间进行。胆碱酯酶将使用分光光度技术测量活性。在此外，还将进行总蛋白和非胶原蛋白检测以便能够在不同人群之间和不同人群之间进行比较不同的年龄。蛋白质估计本身可能揭示差异正常人群和敏感人群之间。数据源自这些研究将提出影响气道平滑调节的因素体外肌张力并直接应用于治疗阻塞性气道疾病和哮喘的干预措施。