title: Vesicular exocytosis contributes to volume-sensitive ATP release in biliary cells.
abstract: Extracellular ATP is a potent autocrine/paracrine signal that regulates a broad range of liver functions through activation of purinergic receptors. In biliary epithelium, increases in cell volume stimulate ATP release through a phosphoinositide 3-kinase (PI3-kinase)-dependent mechanism. Because PI3-kinase also regulates vesicular exocytosis, the purpose of these studies was to determine whether volume-stimulated vesicular exocytosis contributes to cellular ATP release. In a human cholangiocarcinoma cell line, exocytosis was measured by using the plasma membrane marker FM1-43, whereas ATP release was assessed by using a luciferase-luciferin assay. Under basal conditions, cholangiocytes exhibited constitutive exocytosis at a rate of 1.6%/min, and low levels of extracellular ATP were detected at 48.2 arbitrary light units. Increases in cholangiocyte cell volume induced by hypotonic exposure resulted in a 10-fold increase in the rate of exocytosis and a robust 35-fold increase in ATP release. Both vesicular exocytosis and ATP release were proportional to cell volume, and both exhibited similar regulatory properties including: 1) dependence on intact PI3-kinase, 2) attenuation by inhibition of PKC, and 3) potentiation by activation of PKC before hypotonic exposure. These findings demonstrate that increases in cholangiocyte cell volume stimulate ATP release and vesicular exocytosis through similar regulatory paradigms. Functional interactions among cell volume, PKC, and PI3-kinase modulate exocytosis, thereby regulating ATP release and purinergic signaling in cholangiocytes. It is hypothesized that PKC is involved in the recruitment of a volume-sensitive vesicular pool to a readily releasable state.
authors: Department of Medicine, University of Colorado Health Sciences Center, Denver, CO 80262, USA. david.gatof@uchsc.edu
authors: David
authors: D
authors: Gatof
authors: Gordan
authors: G
authors: Kilic
authors: J Gregory
authors: JG
authors: Fitz
mesh_terms: D000230:Adenocarcinoma / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000378:metabolism*; D045744:Cell Line, Tumor; D002462:Cell Membrane / Q000378:metabolism; D048429:Cell Size; D000242:Cyclic AMP / Q000502:physiology; D020536:Enzyme Activators / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004847:Epithelial Cells / Q000187:drug effects / Q000378:metabolism; D005089:Exocytosis / Q000187:drug effects / Q000502:physiology*; D005453:Fluorescence; D005704:Gallbladder / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism*; D005706:Gallbladder Neoplasms / Q000378:metabolism; D006801:Humans; D009994:Osmolar Concentration; D019869:Phosphatidylinositol 3-Kinases / Q000502:physiology; D000081082:Phosphoinositide-3 Kinase Inhibitors; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000502:physiology
chemical_list: D000255:Adenosine Triphosphate; D000242:Cyclic AMP; D020536:Enzyme Activators; D004791:Enzyme Inhibitors; D019869:Phosphatidylinositol 3-Kinases; D000081082:Phosphoinositide-3 Kinase Inhibitors; D011493:Protein Kinase C