MECHANISMS OF CELL VOLUME REGULATION AND LIVER FUNCTION

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DESCRIPTION (taken from application) Preliminary investigations have identified new insights regarding the roles of Cl- channels and cellular ATP release during liver cell swelling, and molecular correlates of the Cl- channels involved in these pathways. The goals of these studies are to better elucidate the cellular and molecular mechanisms involved in liver cell volume regulation, and implications for liver function. The specific aims are to i) characterize channel-mediated ATP efflux and auto-/ paracrine modulation of liver cell volume and bile formation, ii) investigate the role of liver ATP-binding cassette (ABC) proteins as ATP channels or channel regulators, iii) assess the role of protein kinase C (PKC) as a signaling factor that regulates volume recovery, and iv) identify the molecular basis for increased Cl-permeability during swelling and purinergic stimulation. Using both hepatocyte and biliary cells and cell lines, techniques will include a) study of swelling-activated Cl- and ATP channels using patch clamp analysis, b) measurement of cell volume changes with a Coulter Multisizer, c) characterization of epithelial transport with Ussing chamber voltage-clamp experiments in polarized primary rat cholangioctye monolayers and d) cloning and functional characterization of hepatobiliary CLC Cl- channel homologues. Studies will be performed by the applicant at the University of Colorado Health Sciences Center under the preceptorship of Dr. J. Gregory Fitz. During this mentored research period, investigative tools in cell biology, physiology, and molecular biology will be developed to compliment electrophysiologic study of liver ion channels, and serve as a basis for future contributions in the unexplored area of hepatobiliary ion channel genetics. The long term objective of these investigations is to provide a physiologic basis for the development of pharmacological approaches to modulate biliary secretion in cholestatic liver diseases such as cystic fibrosis, and to ameliorate cellular injury during metabolic stress.