Control of motility in the internal anal sphincter

描述(申请人提供)：肛门内括约肌(IAS)保持70%的肛门静息张力，这是维持大便控制的关键。收缩松弛是排便的一个基本特征，这是由抑制性神经输入调节的。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IAS正常收缩的机制的了解有限。最近的研究强调了Rho激酶通路的作用，但只有4篇报道研究了膜电位(Em)在这一临床上重要的胃肠道区域调节中的作用。我们的初步数据表明，受Em调节的L型钙通道(Cav1.2)对钙离子向收缩机构的输送和决定IAS的收缩活动模式是基本的。初步实验还表明，Rho激酶对收缩的调节至少有一部分可能涉及Em的变化。我们对IAS的免疫组织化学研究显示，Cajal间质细胞(ICC)密集分布于IAS的肌层。然而，这些细胞对IAS的Em和电节律的贡献尚不清楚。因此，我们对这些研究的总体假设是，由于Em对Cav1.2活动的影响，IAS的收缩和抑制性运动神经引起的变化主要由Em调节，ICC通过调节相邻SMC中的Em在这一过程中发挥重要作用。为了验证这一假设，我们将在猴子和小鼠IAS的单独节段中测量Em和收缩。ICC和SMC也将从这些组织中分散出来，以评估离子通道的活性，并使用荧光激活细胞分类(FACS)进行分选，以评估基因表达。Rho激酶途径将使用生化技术进行评估，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确定。还将使用新开发的转基因小鼠模型的新方法来研究和消除ICC。IAS的功能失调导致了严重的社会孤立、失业和制度化，并在美国榨取了数十亿美元的医疗费用。尽管存在这一巨大的卫生保健问题，但IAS仍是胃肠道研究最少的地区。这项拟议的研究将为控制肌肉收缩从而控制肛门压力的机制提供新的见解。这些信息对于制定新的战略以帮助预防以及诊断和治疗IAS功能障碍至关重要。 公共卫生相关性： 肛门内括约肌(IAS)维持70%的静息肛门压力。肛门静息压降低和增加分别导致大便失禁和排便障碍。IAS的功能失调导致了严重的社会孤立、失业和制度化，并在美国榨取了数十亿美元的医疗费用。尽管存在这一巨大的卫生保健问题，但IAS仍是胃肠道研究最少的地区。这项拟议的研究将为控制肌肉收缩从而控制肛门压力的机制提供新的见解。这些信息对于制定新的战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. PUBLIC HEALTH RELEVANCE: The internal anal sphincter (IAS) maintains 70% of resting anal pressure. Reduced and increased anal resting pressure contributes to fecal incontinence and defecatory disorders respectively. 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.