title: Cytosolic Ca2+ and protein kinase Calpha couple cellular metabolism to membrane K+ permeability in a human biliary cell line.
abstract: Cholangiocytes represent an important target of injury during the ischemia and metabolic stress that accompanies liver preservation. Since K+ efflux serves to minimize injury during ATP depletion in certain other cell types, the purpose of these studies was to evaluate the effects of ATP depletion on plasma membrane K+ permeability of Mz-ChA-1 cells, a model human biliary cell line. Cells were exposed to dinitrophenol (50 microM) and 2-deoxyglucose (10 mM) as the standard model of metabolic injury. Whole-cell and single K+ channel currents were measured using patch clamp techniques; and intracellular [Ca2+] ([Ca2+]i) was estimated by calcium green-1 fluorescence. Metabolic stress increased [Ca2+]i, and stimulated translocation of the alpha isoform of protein kinase C (PKCalpha) from cytosolic to particulate cell fractions. The same maneuver increased membrane K+ permeability 40-70-fold as detected by (a) activation of K+selective whole cell currents of 2,176+/-218 pA (n = 34), and (b) opening of apamin-sensitive K+ channels with a unitary conductance of 17.0+/-0.2 pS. PKCalpha translocation and channel opening appear to be related since stress-induced K+ efflux is inhibited by chelation of cytosolic Ca2+, exposure to the PKC inhibitor chelerythrine (25 microM) and downregulation of PKC by phorbol esters. Moreover, K+ currents were activated by intracellular perfusion with recombinant PKCalpha in the absence of metabolic inhibitors. These findings indicate that in biliary cells apamin-sensitive K+ channels are functionally coupled to cell metabolism and suggest that cytosolic Ca2+ and PKCalpha are selectively involved in the response.
authors.affiliation: Department of Medicine, Duke University Medical Center, Durham, North Carolina 27710, USA.
authors.forename: Y
authors.initials: Y
authors.lastname: Wang
authors.forename: R
authors.initials: R
authors.lastname: Roman
authors.forename: T
authors.initials: T
authors.lastname: Schlenker
authors.forename: Y A
authors.initials: YA
authors.lastname: Hannun
authors.forename: J
authors.initials: J
authors.lastname: Raymond
authors.forename: J G
authors.initials: JG
authors.lastname: Fitz
mesh_terms: D019297:2,4-Dinitrophenol / Q000494:pharmacology; D000255:Adenosine Triphosphate / Q000008:administration & dosage / Q000378:metabolism; D001652:Bile Ducts / Q000378:metabolism* / Q000648:ultrastructure; D002118:Calcium / Q000378:metabolism*; D002463:Cell Membrane Permeability; D002614:Chelating Agents / Q000494:pharmacology; D018281:Cholangiocarcinoma; D003600:Cytosol / Q000378:metabolism*; D003847:Deoxyglucose / Q000494:pharmacology; D004553:Electric Conductivity; D006801:Humans; D008856:Microscopy, Fluorescence; D018408:Patch-Clamp Techniques; D011188:Potassium / Q000378:metabolism*; D015221:Potassium Channels / Q000502:physiology; D011493:Protein Kinase C / Q000378:metabolism*; D014407:Tumor Cells, Cultured
chemical_list: D002614:Chelating Agents; D015221:Potassium Channels; D000255:Adenosine Triphosphate; D003847:Deoxyglucose; D011493:Protein Kinase C; D019297:2,4-Dinitrophenol; D011188:Potassium; D002118:Calcium