Control of motility in the internal anal sphincter

描述（由申请人提供）：肛门内括约肌（IAS）维持70%的肛门静息张力，这对维持粪便蠕动至关重要。收缩的放松是排便的基本特征，这是由抑制神经的输入调节的。肛门张力降低和增加分别导致大便失禁和排便障碍。我们对维持正常IAS收缩的机制的理解是有限的。最近的研究强调了Rho激酶途径的作用，但只有4篇报道研究了膜电位（Em）在胃肠道这一临床重要区域的调节中的作用。我们的初步数据表明，L-型钙通道（Cav1.2），由EM调节，是根本的钙输送到收缩装置，并确定在IAS的收缩活动的模式。初步的实验还表明，至少有一部分的调节收缩的Rho激酶可能涉及的变化。免疫组化研究表明，Cajal间质细胞（ICC）密集分布在整个肌层的IAS。然而，这些细胞的贡献，EM和电节律的IAS是未知的。因此，我们对这些研究的总体假设是，IAS的收缩和抑制性运动神经引起的变化主要由Em调节，这是由于其对Cav1.2活性的影响，ICC通过调节相邻SMC中的Em在此过程中发挥重要作用。为了检验这一假设，将在猴和小鼠IAS的分离节段中测量Em和收缩。还将从这些组织中分散ICC和平滑肌细胞（SMC）以评价离子通道活性，并使用荧光激活细胞分选（FACS）进行分选以评价基因表达。Rho激酶通路将使用生物化学技术进行评估，氮能神经的分布及其与ICC和SMC的关系将使用免疫组织化学和免疫细胞化学技术进行确定。涉及新开发的转基因小鼠模型的新方法也将用于研究和消除ICC。IAS的功能失调导致了大量的社会孤立，失业和制度化，以及在美国提取数十亿美元的医疗保健费用。尽管有这个大的卫生保健问题，IAS是胃肠道研究最少的区域。这项研究将为控制肌肉收缩和肛门压力的机制提供新的见解。这些信息对于制定新的策略以帮助预防以及诊断和治疗IAS功能障碍至关重要。

DESCRIPTION (provided by applicant): The internal anal sphincter (IAS) maintains 70% of anal resting tone, which is critical for maintaining fecal continence. Relaxation of contraction is an essential feature of defecation, and this is regulated by input from inhibitory nerves. Reduced and increased anal tone contribute to fecal incontinence and defecatory disorders respectively. Our understanding of the mechanisms which maintain normal IAS contraction is limited. Recent studies have emphasized the role of the Rho kinase pathway, but there are only 4 reports which have investigated the role of membrane potential (Em) in the regulation of this clinically important region of the GI tract. Our preliminary data suggest that L-type Ca2+ channels (Cav1.2), regulated by Em, are fundamental to the delivery of Ca2+ to the contractile apparatus and for determining the pattern of contractile activity in the IAS. Preliminary experiments also suggest that at least a portion of the regulation of contraction by Rho kinase may involve changes in Em. Our immunohistochemical studies of the IAS have revealed that interstitial cells of Cajal (ICC) are densely distributed throughout the muscularis of the IAS. However, the contribution of these cells to Em and electrical rhythmicity in the IAS is unknown. Thus, our overall hypothesis for these studies is that contraction in the IAS and the changes elicited by inhibitory motor nerves is primarily regulated by Em due to its effects upon Cav1.2 activity and that ICC play an important role in this process by modulating Em in adjacent SMCs. To examine this hypothesis Em and contraction will be measured in isolated segments of the monkey and mouse IAS. ICC and smooth muscle cells (SMC) will also be dispersed from these tissues to evaluate ion channel activity and sorted using fluorescence activated cell sorting (FACS) to evaluate gene expression. The Rho kinase pathway will be assessed using biochemical techniques and the distribution of nitrergic nerves and their relationship to ICC and SMC will be determined using immmunohistochemical and immunocytochemical techniques. Novel approaches involving newly developed transgenic mice models will also be used to study and eliminate ICC. Dysfunction of the IAS leads to substantial social isolation, loss of employment, and institutionalization as well as extracting billions of dollars in health care costs in the United States. In spite of this large health care problem the IAS is the least studied region of the GI tract. The proposed study will provide new insights into the mechanisms which control muscle contraction and hence anal pressure. Such information is critical for developing new strategies to help prevent as well as diagnose and treat IAS dysfunction.