Neural Control of Large Intestinal Mucosa

肠道神经的神经反射控制着适当的消化功能所必需的分泌和运动模式。机械刺激从肠嗜铬细胞释放5-羟色胺（5- ht），并在反射中与粘膜下传入神经元上的5- ht1p /4受体结合，引起上皮氯离子分泌。总体目标是了解施加于结肠的机械刺激转导成氯离子分泌的机制。将使用三种机械刺激：粘膜抚摸、旋转震动和细胞变形。第一个目标采用综合方法检测豚鼠、人类或核苷酸受体缺陷小鼠肌肉剥离结肠对机械刺激或类癌细胞系BON的5-羟色胺释放和氯离子分泌。将鉴定释放的核苷酸，根据激动剂/拮抗剂效力分类的嘌呤受体亚型，以及通过电子显微镜鉴定被称为小泡的质膜信号站。第二个目的是确定所讨论的嘌呤受体在机械刺激过程中是否依赖于小泡结构来传递适当的信号。第三个目的是确定Gq家族的G蛋白是否与小泡中的嘌呤受体共定位，以介导BON细胞中机械诱发的5-羟色胺释放。对于这些和后续的研究，免疫沉淀和免疫印迹技术将识别在小囊中富集的部分中信号分子的存在。第四个目的是研究单个BON细胞机械变形的机制，并确定磷脂酶C和细胞内钙储存在机械诱发的5-HT释放中的作用。第5个目标是通过检测钙储存和磷脂酶C的磷酸化来确定蛋白激酶C调节机械诱发的5-羟色胺释放的机制。这项研究从检查综合系统到单细胞的机制方法将为正常人类和动物组织中5-羟色胺释放的调节提供新的见解。这将为类癌肿瘤的基本生理和5- HT的释放提供新的信息，可能有利于类癌综合征的治疗。了解5-羟色胺的调节可能是管理与肠色素细胞数量过多或减少相关的功能性肠病的关键。

Neural reflexes within the enteric nervous control secretion and motility patterns necessary for appropriate digestive function. Mechanical stimulation release 5- hydroxytryptamine (5-HT) from enterochromaffin cells and binds to 5-HT1P/4 receptors on submucosal afferent neurons in the reflex to cause epithelial chloride secretion. The overall goal is to understand the mechanisms by which a mechanical stimulus applied to the colon is transduced into chloride secretion. Three mechanical stimuli will be used: mucosal stroking, rotational shaking and cell deformation. The 1st aim uses an integrative approach to examine 5-HT release and chloride secretion in muscle-stripped colon from guinea pig, human or nucleotide- receptor deficient mice n response to mechanical stimulation or in a carcinoid tumor cell line, BON. Nucleotides released will be identified, the purinoreceptor subtype classified by agonist/antagonist potencies and the plasma membrane signaling stations known as caveolae identified by electron microscopy. The 2nd aim is to determine whether the purinoceptor in question is dependent on caveolae structure for appropriate signaling during mechanical stimulation. The 3rd aim is to determine if G proteins of the Gq family co-localize with purinoceptors in caveolae to mediate mechanically-evoked 5-HT release in BON cells. For these and subsequent studies immunoprecipitation and immunoblotting techniques will identify the presence of signaling molecules in fractions enriched in caveolae. The 4th aim examines the mechanism involved in mechanical deformation of single BON cells, and determines the role of phospholipase C and intracellular calcium stores in mechanically-evoked 5-HT release. The 5th aim identifies mechanisms by which protein kinase C in modulates mechanically-evoked 5-HT release by examining calcium stores and phosphorylation of phospholipase C. This study with its mechanistic approach from examining integrated systems to the single cell will provide new insights into the regulation of 5-HT release in normal human and animal tissue. It will provide new information about the basic physiology of carcinoid tumors and 5- HT release that may benefit the management of carcinoid syndrome. Understanding the regulation of 5-HT may be a key to management of functional bowel disease associated with an excess or reduced number of enterochromaffin cells.