REGULATION OF CHLORIDE ION CONDUCTANCE IN PANCREAS DUCT

囊性纤维化是一种由囊性纤维化跨膜电导调节蛋白突变引起的多系统疾病。正常情况下，该蛋白在顶端质膜中作为cAMP调节的c1-通道发挥作用。CFTR缺陷与CF的病理生理学之间的确切联系尚不清楚。一般认为，CFT是由于异常的离子和水跨受影响的上皮细胞运动所致；然而，被归因于CFTR的其他调节功能也可能与该病有关。遗传和药物恢复根尖膜在CF上皮细胞的通透性常被认为是一种可行的治疗策略。其中一种方法是激活“交替”的根尖膜，即非CFTRc1通道。这种方法的理论基础是观察到，人CF呼吸道上皮细胞中保留了钙激活的C1-电导(CACC)，并且在CF基因敲除小鼠中，器官特异性疾病的严重程度与CACC的活性成反比。胰腺管上皮细胞至少表达4种质膜电导：cAMP激活(CFTR)、cAMP激活(非CFTR)、CACC和肿胀激活(SACC)。至少有两个，甚至多达四个不同的质膜阴离子通道构成了这四条电导通路的基础。只有一种(CFTR)的分子同一性是已知的。在胰管细胞中，CACC、cAMP激活的(非CFTR)和SACC电导的调节几乎没有受到关注。我们以前的研究表明，在胰管细胞的顶端细胞膜上存在钙激活和cAMP激活的(非cftr)电导，因此是规避功能性cftr丢失的潜在途径。本提案中描述的工作的目标是确定这两个电导的单通道基础，并确定控制通道活动的调节路径。将使用响应于信号转导通路操纵的通道功能的电生理学测量。这项工作的长期目标是开发药物方法来控制盐和水的运输，以补偿囊性纤维化影响的上皮细胞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.