Nitrergic Neuro-smooth Muscle Transmission in the Gut

描述（由申请人提供）：大量证据表明，一氧化氮（NO）气体在全身（包括肠道）的非肾上腺素能非胆碱能（NANC）抑制性神经平滑肌传递中发挥关键作用。临床上，氮能传递的缺陷已被证明是许多主要胃肠动力障碍的原因。然而，关于氮能神经传递的几个重要问题仍然没有答案。例如，在神经-平滑肌连接处（包括连接前神经静脉曲张、连接后平滑肌细胞或肌内型Coal间质细胞（ICC-IM））产生NO的精确位点仍然不确定。一些研究者认为，在神经肌肉传递中，连接后结构产生的NO是必不可少的。目前还不清楚这种高度扩散的气体如何实现需要高度控制释放的神经递质的功能。一种经典神经递质的释放受停靠在质膜上的一小部分分泌颗粒的胞吐作用调节。然而，NO不储存在分泌颗粒中，并且被认为是在神经传递期间按需产生的。含有nNOS的神经静脉曲张也含有VIP肽，然而，VIP在氮能神经平滑肌神经传递中的作用是有争议的。我们提出了nNOS的调节模型和VIP在氮能神经传递中的中介作用。为了测试这些模型，我们计划利用：（1）一种新的技术，使用荧光染料标记物和多光子显微镜，以实时显示在活神经静脉曲张和其他结构的神经肌肉接头处的NO和钙信号;（2）免疫细胞学研究，使用多光子显微镜定位可能参与nNOS从催化失活到催化活性形式的循环的蛋白质;（3）氮能抑制性连接电位的细胞内记录。使用上述技术，我们计划研究药理学操作的影响，以测试所提出的模型。这些研究将有助于确定：（1）在神经传递过程中，NO是否在神经静脉曲张和连接后平滑肌细胞或ICC-IM中产生;（2）允许NO气体作为神经递质发挥作用的nNOS的调节;（3）VIP及其独特的受体亚型在氮能神经传递中的中介作用。这些信息可能有助于了解生理学，以及由于缺乏氮能神经平滑肌传递引起的临床疾病的发病机制和合理治疗。

DESCRIPTION (provided by applicant): A large body of evidence suggests that nitric oxide (NO) gas play a key role in nonadrenergic noncholinergic (NANC) inhibitory neuro-smooth muscle transmission throughout the body including the gut. Clinically, deficiency of nitrergic transmission has been shown to be responsible for many major gastrointestinal motility disorders. However, several important questions regarding nitrergic neurotransmission remain unanswered. For example, the precise site(s) of production of NO at the neuro-smooth muscle junctions, including the prejunctional nerve varicosities, the postjunctional smooth muscle cells or the intramuscular type of interstitial cells of Coal (ICC-IM) remain undefined. Some investigators believe that NO produced in the postjunctional structures is essential in the neuromuscular transmission. It is also unclear how this highly diffusible gas can fulfill the function of a neurotransmitter that requires highly controlled release. The release of a classical neurotransmitter is regulated by exocytosis of a small fraction of the secretory granules that are docked onto the plasma membrane. However, NO is not stored in the secretory granules and is thought to be produced on demand during the neurotransmission. The nerve varicosities that contain nNOS also contain peptide VIP, however, the role VIP in nitrergic neuro smooth muscle neurotransmission is controversial. We propose models of regulation of nNOS and an intermediary role of VIP in nitrergic neurotransmission. To test these models we plan to utilize: (1) a novel technique using fluorescent dye markers and multi-photon microscopy to visualize NO and calcium signals in real-time in live nerve varicosities and other structures at the neuromuscular junctions; (2) immunocytological studies using multi-photon microscopy to localize the proteins that may be involved in cycling of nNOS from catalytically inactive to catalytically active forms; (3) intracellular recordings of the nitrergic inhibitory junction potentials. Using the above techniques, we plan to study the effects of pharmacological manipulations to test the proposed models. These studies will help define:(1) whether during the neurotransmission NO is produced in the nerve varicosities and also in the postjunctional smooth muscle cells or ICC-IM; (2) the regulation of nNOS that permits NO gas to function as a neurotransmitter; (3) the intermediary role of VIP and its unique receptor subtype in nitrergic neurotransmission. This information may help in understanding physiology, and the pathogenesis and rational treatment of clinical disorders due to deficiency of nitrergic neuro-smooth muscle transmission.