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）气体在包括肠道在内的全身非肾上腺素能非胆碱能（NANC）抑制神经-平滑肌传递中起关键作用。临床上，氮能传递不足已被证明是许多主要胃肠运动障碍的原因。然而，关于氮神经传递的几个重要问题仍未得到解答。例如，神经-平滑肌连接处产生NO的确切位置，包括神经结前神经曲张、结后平滑肌细胞或肌内间质细胞（ICC-IM）仍未明确。一些研究者认为，突触后结构产生的一氧化氮在神经肌肉传递中是必不可少的。目前还不清楚这种高度扩散的气体是如何实现一种需要高度控制释放的神经递质的功能的。经典神经递质的释放是由停靠在质膜上的一小部分分泌颗粒的胞吐调节的。然而，一氧化氮并不储存在分泌颗粒中，而被认为是在神经传递过程中根据需要产生的。含有nNOS的神经静脉曲张也含有肽VIP，但肽VIP在氮神经平滑肌神经传递中的作用尚存争议。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)一种使用荧光染料标记和多光子显微镜的新技术来实时可视化活神经静脉曲张和神经肌肉连接处其他结构中的NO和钙信号；(2)利用多光子显微镜进行免疫细胞学研究，定位可能参与nNOS从催化无活性形式到催化活性形式循环的蛋白质；(3)细胞内氮抑制连接电位的记录。利用上述技术，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)在神经传递过程中是否在神经曲张和结后平滑肌细胞或ICC-IM中产生NO；(2) nNOS的调节，允许NO气体作为神经递质发挥作用；(3) VIP及其独特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氮能神经-平滑肌传递不足引起的临床疾病的生理学、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.