Nitrergic Neuro-smooth Muscle Transmission in the Gut

描述（申请人提供）：一氧化氮（NO）是胃肠平滑肌重要的抑制性神经递质。NO作为一种神经递质，在蠕动和括约肌的生理松弛中起关键作用，从而促进食物通过胃肠道的运输。一氧化氮相关神经传递的丧失已被证明会导致吞咽困难、胃瘀、消化不良、肠瘀和便秘等疾病。一氧化氮神经传递缺陷也与糖尿病性胃轻瘫和肠易激综合征、胆道和胰腺括约肌痉挛等肠道功能障碍有关。一氧化氮与其他经典神经递质有着独特的区别。与传统的神经递质不同，一氧化氮是一种高度扩散的气体，它是由一氧化氮的Ca-CaM依赖性激活重新产生的。然而，NO生成和氮神经传递的调控尚不清楚。我们最近的研究表明，从小鼠肠道分离的氮能静脉曲张含有非活性和活性的nNOS池。催化活性nNOS1的动态调控负责氮能神经传递的调控。本研究的总体目的是扩展我们对氮氧变异中催化活性nNOS调控的研究，并确定nNOS如何被运输到膜上，附着在膜上并被调节产生NO。我们将使用这些信息来识别可能损害氮能神经传递的步骤中的异常。这些研究将有助于确定没有解剖证据表明氮能神经丢失或损伤的氮能神经传递受损的病理生理学。研究目的有四：1)研究PIN/LC8和肌球蛋白Va在静脉曲张膜靶向nNOS中的作用，研究肌球蛋白Va缺乏小鼠的氮能神经传递。2)研究PSD蛋白在nNOS1二聚体的膜结合中的作用，研究PSD去棕榈酰化和PSD 95缺乏小鼠的氮能神经传递。3)研究nNOS- psd复合物与n型钙通道的聚类及nNOS去磷酸化或磷酸化酶的作用，并探讨这些酶的抑制剂对氮能神经传递的影响。4)探讨糖尿病动物模型（NOD小鼠）抑制性神经传递降低的机制。

DESCRIPTION (provided by applicant): Nitric oxide (NO) is an important inhibitory neurotransmitter for the gastrointestinal smooth muscles. As a neurotransmitter, NO plays a key role in peristalsis and physiological relaxation of the sphincters and thus facilitates transport of food through the gastrointestinal tract. Loss of NO-related neurotransmission has been shown to cause diseases causing difficulty in swallowing, gastric stasis, dyspepsia, intestinal stasis and constipation. Defective NO neurotransmission is also implicated in diabetic gastroparesis and functional disorders of the gut such as irritable bowel syndrome and spasm of biliary and pancreatic sphincters. Nitric oxide is uniquely different from other classical neurotransmitters. Unlike the classical neurotransmitters, NO is a highly diffusible gas that is produced de novo on demand from Ca-CaM dependent activation of nNOS. However, regulation of NO generation and nitrergic neurotransmission is not well understood. We have recently shown that isolated nitrergic varicosities from mice gut contain inactive and active pools of nNOS. Dynamic regulation of the catalytically active nNOS1 is responsible for regulation of nitrergic neurotransmission. Overall purpose of the proposed studies is to extend our studies of regulation of catalytically active nNOS in the nitrergic varicosities and determine how nNOS is transported to the membrane, attached there and regulated to produce NO. We will use this information to identify abnormities in the steps that may impair nitrergic neurotransmission. These studies will help define pathophysiology of impaired nitrergic neurotransmission that have no anatomical evidence of loss or damage to nitrergic nerves. There are four specific aims: 1) To examine the role of PIN/LC8 and myosin Va in targeting nNOS to varicosity membrane and to investigate nitrergic neurotransmission in mice lacking myosin Va. 2) To examine the role of PSD proteins in membrane association of nNOS1 dimer and to investigate nitrergic neurotransmission with depalmitoylation of PSD and in mice lacking PSD 95. 3) To investigate the clustering of nNOS-PSD complex with N-type calcium channels and the enzymes that dephosphorylate or phosphorylate nNOS and to investigate the effects of inhibitors of these enzymes on nitrergic neurotransmission. 4) To investigate the mechanism of reduced inhibitory neurotransmission in an animal model of diabetes mellitus (NOD mice). PUBLIC HEALTH RELEVANCE: Gastrointestinal motility disorders result in a variety of diseases that cause difficulty in swallowing, heartburn, dyspepsia, constipation and chronic abdominal pain. This proposal is to understand how these disorders are caused and how better treatments for these disorders can be developed.