MECHANISM OF FORCE GENERATION & MAINTENANCE IN BLADDER--OULET OBSTRUCTION

尿路梗阻导致尿路的许多变化 有损其储存和排空特性的膀胱。详细的 导致这些功能变化的分子机制相关 有梗阻所致的重塑，但尚未清楚描绘。 这项研究项目是基于一种假设，即抑郁症患者 梗阻性膀胱的功能是两者改变的结果。 兴奋-收缩偶联(钙可利用性)与收缩 逼尿肌中的装置(钙敏感性和钙依赖性活动) 肌肉。为了验证这一假设，将实现以下具体目标 使用取自正常人的逼尿肌条， 失代偿和代偿兔膀胱：目标1：相互关联 完整和带皮逼尿肌力-速度长度关系的研究 与膀胱梗阻及其逆转有关。这一目标将 确定组织、细胞和组织的力学改变的机制 跨桥水平。经过化学处理的肌肉条带可以精确地 以及对收缩周围环境的直接控制 装置，绕过正常的兴奋-收缩通路。目标2：实现 确定激动剂浓度与胞浆[Ca~(2+)]的关系 以及收缩的时间进程。笼状化合物的激光光解 将被用来启动收缩，同时监测细胞内 [Ca2+]与Indo-1。这一目标将决定改变的收缩能力是否 是由于钙离子动员的改变。目标3：确定时间 肌球蛋白轻链磷酸化、[Ca~(2+)]与等长力的变化 在完整和肌球蛋白轻链磷酸化中，肌动蛋白激活的肌球蛋白 通透性组织中的ATPase活性和等长力 各种刺激。这一目标将测试在联轴器的变化 Ca~(2+)信号转导收缩激活。目标4：确定 其他调控信号通路的意义。蛋白激酶C， 丝裂原活化蛋白激酶、钙调蛋白磷酸化级联反应将 被测量。这一目标将决定维持收缩的损失是否 失代偿膀胱中的力是由于细丝的变化所致 监管。这些研究的结果，连同在 项目1将阐明兴奋-收缩的具体步骤 与膀胱功能障碍相关的偶联。此外， 这些研究的结果还将确定哪些变化 收缩或调节蛋白，与项目2合作，是 与膀胱重塑相关，并将提供更完整的 对细胞和分子机制的理解 与出口梗阻相关的膀胱功能低落。

Obstruction of the urethra results in numerous alterations in the urinary bladder that impair both its storage and emptying properties. The detailed molecular mechanisms responsible for these changes in function associated with obstruction-induced remodeling that have not been clearly delineated. This research project is based upon the hypothesis that the depressed function of the obstructed bladder is the result of alterations in both excitation-contraction coupling (Ca2+-availability) and the contractile apparatus (Ca2+-sensitivity and Ca2+-dependent activity) in detrusor muscle. To test this hypothesis, the following specific aims will be addressed using strips of detrusor muscle obtained from normal, decompensated, and compensated rabbit bladder: Aim 1: To correlate the force-velocity length relations of intact and skinned detrusor muscle associated with bladder obstruction and its reversal. This aim will determine the mechanism of altered mechanics at the tissue, cell and cross-bridge level. Chemically skinned muscle strips allow for the precise and direct control of the environment surrounding the contractile apparatus, bypassing the normal excitation-contraction pathway. Aim 2: To determine the relationship among agonist concentration, cytoplasmic [Ca2+] and the time course of contraction. Laser photolysis of caged-compounds will be used to initiate contraction while monitoring the intracellular [Ca2+] with Indo-1. This aim will determine if the altered contractility is due to altered calcium mobilization. Aim 3: To determine the time course of myosin light chain phosphorylation, [Ca2+], and isometric force in intact and myosin light chain phosphorylation, actin-activated myosin ATPase activity and isometric force in permeabilized tissues during various stimuli. This aim will test for alteration in the coupling of the Ca2+ signal to contractile activation. Aim 4: To determine the significance of other regulatory signaling pathways. The protein kinase C, mitogen-activated protein kinase, caldesmon phosphorylation cascade will be measured. This aim will determine if the loss of maintained contractile force in the decompensated bladder is due to alterations in thin filament regulation. The results of these studies, in conjunction with those in Project 1, will elucidate the specific steps of excitation-contraction coupling that are associated with bladder dysfunction. Moreover, the results of these studies will also determine which alterations in contractile or regulatory proteins, in collaboration with Project 2, are associated with bladder remodeling and will provide a more complete understanding of the cellular and molecular mechanisms responsible for the depressed bladder function associated with outlet obstruction.