The studies described in this proposal address the cellular mechanisms responsible for secretion by cholangiocytes, the epithelial cells that line the lumen of intrahepatic bile ducts and account for approximately 40 percent) of bile volume in man. Previous studies indicate that the secretory capacity of cholangiocytes undergoes dynamic changes in response to varying physiologic demands; and depends on selective) penning of separate Cl- channels in response to cAMP, cytosolic Ca2+, and cell volume. Notably, these channels are present in intracellular vesicles that are highly regulated and are of sufficient capacity to replace 40 percent of the entire cholangiocyte plasma membrane within minutes. Accordingly, the Specific Aims are designed to address the working hypothesis that trafficking (endocytosis and exocytosis) of membrane vesicles containing ion channels provides an early and essential mechanism for modifying the composition and conductance of the cholangiocyte plasma membrane and is essential for the secretory responses to cAMP, Ca2+, and cell volume.
The Specific Aims occurs on 1) characterization of the cellular signals responsible for regulation of endocytosis and exocytosis, 2) evaluation, of the relative roles of vesicular trafficking and direct regulatory phosphorylation by PKCa of SKCa channels essential for modulation of cholangiocyte volume and Juctular secretion; and 3) definition of the cellular strategies for maintaining the multiprotein signaling complex responsible for apical ATP release, P2 receptor distribution and K+ and Cl- secretion. The long-term goal is to define the cellular mechanisms involved in ductular secretion, and to identify the) physiologic factors which contribute to bile formation through effects on duct cells. Thus, the findings ire directly relevant to diagnosis and management of a broad range of cholestatic disorders characterized by impaired cholangiocyte function; and to development of pharmacologic approaches o modify the volume and composition of bile through effects on duct cells.
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