REGULATION OF CHLORIDE ION CONDUCTANCE IN PANCREAS DUCT

囊性纤维化（CE）是一种由囊性纤维化跨膜传导调节蛋白（CFTR）突变引起的多系统疾病。正常情况下，该蛋白在根尖质膜中作为camp调控的c1通道发挥作用。CFTR缺陷与CF病理生理之间的确切联系尚不清楚。一般认为CF是由异常的离子和水在受累上皮间的运动引起的；然而，归因于CFTR的其他调节功能可能导致该疾病。遗传和药理学恢复CF上皮顶端膜c1通透性通常被认为是可行的治疗策略。其中一种方法是激活“交替”的顶端膜，非cftr c1通道。这种方法的基本原理在于观察到Ca2+激活的c1传导（CACC）在人CF气道上皮中被保留，并且CF敲除小鼠的器官特异性疾病严重程度与CACC的活性呈负相关。胰管上皮细胞表达至少4种类型的质膜c1传导：camp激活（CFTR）、camp激活（非CFTR）、CACC和肿胀激活（SACC）。至少有两个，也许多达四个，不同的质膜阴离子通道是这四个电导途径的基础。只有一种（CFTR）的分子特性是已知的。在胰管细胞中，对CACC、camp激活（非cftr）和SACC电导的调节很少受到关注。我们之前的研究表明，Ca2+激活和camp激活（非CFTR）电导存在于胰管细胞的顶细胞膜中，因此代表了规避功能性CFTR丧失的潜在途径。本提案中描述的工作目标是确定这两种传导的单一通道基础，并确定控制通道活动的调节途径。通道功能的电生理测量响应于信号转导通路的操作将被使用。这项工作的长期目标是开发控制盐和水运输的药理学方法，以补偿受囊性纤维化影响的上皮中CFTR功能的丧失。

Cystic Fibrosis (CE) is a multi-system disease caused by mutations in the Cystic Fibrosis Transmembrane conductance Regulator (CFTR) protein. Normally the protein functions as a cAMP-regulated C1-channel in the apical plasma membrane. The precise link between defective CFTR and the pathophysiology of CF is not known. It is generally assumed that CF results from abnormal ion and water movement across affected epithelia; however, other regulatory functions ascribed to CFTR may contribute to the disease. Genetic and pharmacologic restoration of apical membrane C1-permeability in CF epithelia are often cited as a viable therapeutic strategies. One such approach is to activate "alternate" apical membrane, non-CFTR C1-channels. The rationale for this approach rests upon the observation that Ca2+-activated C1-conductance (CACC) is preserved in human CF airway epithelia and that organ-specific disease severity in the CF knockout mouse is inversely related to the activity of CACC. Pancreatic duct epithelial cells express at least 4 types of plasma membrane C1-conductances: cAMP-activated (CFTR), cAMP-activated (non-CFTR), CACC, and swelling activated (SACC). At least two, and perhaps as many as four, different plasma membrane anion channels underlie these four conductance pathways. The molecular identity of only one (CFTR) is known. Regulation of the CACC, cAMP-activated (non-CFTR), and SACC conductances has received little attention in pancreatic duct cells. Our previous studies revealed that the Ca2+-activated and the cAMP-activated (non-CFTR) conductances are present in the apical cell membrane of pancreatic duct cells and thereby represent potential pathways to circumvent the loss of functional CFTR. The goal of the work described in this proposal is to identify the single channel basis for these two conductances and to determine the regulatory pathways that control channel activity. Electrophysiological measurements of channel function in response to manipulation of signal transduction pathways will be used. The long term objective of the work is to develop pharmacologic approaches to control salt and water transport so as to compensate for loss of CFTR function in epithelia affected by cystic fibrosis.