Intrahepatic ducts contribute importantly to the volume and composition of bile secreted by the liver. The studies described in this proposal address the specialized functions of cholangiocytes, the epithelial cells that line the lumen of these ducts and are responsible for bile formation. Opening of Cl- channels in the apical membrane of cholangiocytes represents one point for regulation of secretion and involves Cl- efflux through CFTR, the protein product of the cystic fibrosis gene. Isolated cholangiocytes also express a rich array of other channels that are regulated by both receptor-dependent (secretin, ATP) and -independent (bile acids, cell volume) pathways. The proposed studies will evaluate the working hypothesis that transepithelial transport of Cl- ions represents an important driving force for secretion across intrahepatic duct epithelium; and that both local (autocrine, paracrine) factors and systemic (hormonal) factors work in concert to modulate the volume and composition of bile through direct effects on duct cells. Patch clamp and other techniques will be utilized to evaluate transport currents in isolated cholangiocytes and polarized models of biliary cells which have had little prior application to investigation of these issues.
The specific aims are 1) to localize membrane ion channels to the apical or basal domains and critically evaluate the role of Cl- transport as a driving force for ductular secretion; 2) to assess the complementary roles of Ca2+/calmodulin-dependent kinases in regulation of secretion, and protein kinase C in regulation of cell volume; and to evaluate 3) extracellular ATP and 4) bile acids as local regulatory factors which modulate membrane Cl-permeability in response to changing physiologic demands. The long-term goal of these studies is to define the cellular mechanisms involved in regulation of ductular secretion. Collectively, the findings are directly relevant to the diagnosis and management of a broad range of disorders characterized by impaired cholangiocyte transport, and to the development of new therapeutic approaches aiming to modulate the volume and composition of bile through direct effects on duct cells.
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