Neuronal Excitability and Motility in Colitis

描述(申请人提供)：伴随炎症性肠病(IBD)的显著运动障碍反映了炎性介质和肠道神经系统之间的动态相互作用。虽然很明显，炎症介质可以触发肠神经反应，但它们的确切靶点和作用机制尚不清楚。这些研究旨在阐明炎症如何调节肠道反射回路的传入成分，即肠嗜铬细胞和AH神经元。我们将验证一种假设，即结肠炎与EC细胞5-羟色胺(5-HT)的可获得性增加和AH神经元的兴奋性增加有关，并且这些变化是由环氧合酶2(COX-2)产生的前列腺素介导的。为了验证这一假设，我们将使用经过彻底验证的三硝基苯磺酸(TNBS)豚鼠结肠炎模型。在第一个具体目标中，我们将通过定量5-羟色胺免疫反应的EC细胞数量、粘膜5-羟色胺水平、刺激诱导的5-羟色胺释放以及炎症结肠粘膜对5-羟色胺的摄取来评估炎症结肠中5-羟色胺细胞的可用性。在特定的目标2中，我们将测试结肠炎是否与AH神经元的兴奋性增加有关，从而导致这些细胞对生理刺激的激活增加。将通过评估其电学特性和对5-羟色胺的敏感性来研究AH神经元的兴奋性。此外，粘膜刺激对AH神经元的激活将通过(1)电生理学、(2)活性标记物FM2-10、(3)通过测量降钙素基因相关肽的释放、(4)通过测量推进运动活动来评估。第三个特定目标将测试炎症期间诱导的EC细胞和AH神经元功能的变化是否由COX-2诱导产生的前列腺素启动。为此，我们将测试前列腺素是否模拟炎症对EC细胞和AH神经元的影响，以及是否通过阻断COX-2来减弱TNBS诱导的结肠炎的神经内分泌效应。这些研究的结果将揭示炎症和前列腺素如何改变肠道神经回路的传入成分的功能。这些信息将促进我们对与IBD相关的神经免疫整合和运动障碍机制的理解，并可能导致将IBD患者的运动功能恢复到正常水平的新的治疗方法。

DESCRIPTION (provided by applicant): The striking motor disturbances that accompany inflammatory bowel disease (IBD) reflect a dynamic interplay between inflammatory mediators and the enteric nervous system. While it is clear that inflammatory mediators trigger enteric neuronal responses, their precise targets and mechanisms of action are unknown. The proposed studies are designed to elucidate how inflammation modulates the afferent components of the reflex circuitry of the bowel, namely the enterochromaffin (EC) cells and AH neurons. We will test the hypothesis that colitis is associated with an increase in the availability of serotonin (5-HT) from EC cells and increased excitability of AH neurons, and that these changes are mediated by prostaglandins generated by cyclooxygenase 2 (COX-2). To test this hypothesis, we will use the thoroughly validated trinitrobenzene sulfonic acid (TNBS) model of colitis in the guinea pig. In the first specific aim, we will evaluate the availability of 5-HT from EC cells in the inflamed colon by quantifying the 5-HT-immunoreactive EC cell population, mucosal 5-HT levels, stimulus-induced 5-HT release, and 5-HT uptake by the mucosa of the inflamed colon. In Specific Aim 2, we will test whether colitis is associated with an increase in the excitability of AH neurons, leading to an increase in the activation of these cells in response to physiological stimuli. The excitability of AH neurons will be investigated by evaluating their electrical properties and their sensitivity to 5-HT. Furthermore, activation of AH neurons by mucosal stimulation will be evaluated (1) electrophysiologically, (2) with the activity marker, FM2-10, (3) by measuring release of calcitonin gene-related peptide, and (4) by measurement of propulsive motor activity. The third specific aim will test whether changes in EC cell and AH neuron function, that are induced during inflammation, are initiated by prostaglandins derived from the induction of COX-2. To accomplish this, we will test whether prostaglandins mimic the effects of inflammation on EC cells and on AH neurons, and we will test whether the neuroendocrine effects of TNBS-induced colitis are attenuated by blockade of COX-2. The results of these studies will reveal how inflammation and prostaglandins alter the function of the afferent components of enteric neural circuits. This information will advance our understanding of the mechanisms of neuro-immune integration and motility disturbances that are associated with IBD, and may lead to novel therapeutic approaches for restoring motor function to normal levels in individuals with IBD.