title: Regulation of Cl- transport by IBMX in renal A6 epithelium.
abstract: We studied regulation of Cl- transport by cAMP and Ca2+ in renal epithelial A6 cells. Stimulation of A6 cells by 1 mM 3-isobutyl-1-methylxanthine (IBMX, an inhibitor of phosphodiesterase), which increased cytosolic cAMP, elicited biphasic increases in short-circuit current (Isc), i.e., a transient phase followed by a sustained one. Apical application of 5-nitro-2-(3-phenylpropylamino)-benzoate (NPPB, a Cl- channel blocker) markedly and dose-dependently inhibited the IBMX-induced Isc. Pretreatment with nifedipine (100 microM, a Ca2+ channel blocker) or 1,2-bis(o-aminophenoxy)-ethane-N,N,N',N'-tetraacetic acid tetra-(acetoxymethyl)-ester (BAPTA/AM, 10 microM, a Ca2+ chelator) partially but markedly inhibited the Isc. On the other hand, a cAMP-dependent protein kinase inhibitor, H89 (0.5 microM for 1 h), also reduced the IBMX-induced Isc to a level similar to that following nifedipine or BAPTA pretreatment. Nifedipine had no synergistic effects on the IBMX-induced Isc in cells treated with H89. Ionomycin (a Ca2+ ionophore) could mimic the transient increase dose dependently, and H89 did not block the ionomycin-induced Isc. Taken together, our observations suggest that: (1) part of the IBMX-stimulated Cl- release is regulated by an increased cytosolic Ca2+ through nifedipine-sensitive Ca2+ influx; (2) cAMP-dependent phosphorylation may be required for elevation of the cytosolic Ca2+ concentration but not for activation of Cl- channels, which are directly activated by cytosolic Ca2+; and (3) the IBMX-induced sustained Cl- release requires cAMP elevation in addition to an increase in the cytosolic Ca2+ concentration.
authors.affiliation: Laboratory for Cellular and Molecular Physiology, Division of Respiratory Research, Hospital for Sick Children Research Institute, University of Toronto Faculty of Medicine, Toronto, Ontario M5G 1X8, Canada.
authors.forename: N
authors.initials: N
authors.lastname: Niisato
authors.forename: Y
authors.initials: Y
authors.lastname: Marunaka
mesh_terms: D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine / Q000494:pharmacology; D015056:1-Methyl-3-isobutylxanthine / Q000494:pharmacology*; D000818:Animals; D002118:Calcium / Q000378:metabolism; D002121:Calcium Channel Blockers / Q000494:pharmacology; D002460:Cell Line; D018118:Chloride Channels / Q000037:antagonists & inhibitors; D002712:Chlorides / Q000378:metabolism*; D000242:Cyclic AMP / Q000378:metabolism; D003600:Cytosol / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004848:Epithelium / Q000187:drug effects / Q000378:metabolism; D017136:Ion Transport; D007546:Isoquinolines / Q000494:pharmacology; D007668:Kidney / Q000187:drug effects* / Q000378:metabolism*; D007700:Kinetics; D009579:Nitrobenzoates / Q000494:pharmacology; D010726:Phosphodiesterase Inhibitors / Q000494:pharmacology; D047428:Protein Kinase Inhibitors; D013449:Sulfonamides; D014982:Xenopus laevis
chemical_list: D002121:Calcium Channel Blockers; D018118:Chloride Channels; D002712:Chlorides; D004791:Enzyme Inhibitors; D007546:Isoquinolines; D009579:Nitrobenzoates; D010726:Phosphodiesterase Inhibitors; D047428:Protein Kinase Inhibitors; D013449:Sulfonamides; C058176:5-nitro-2-(3-phenylpropylamino)benzoic acid; D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine; D000242:Cyclic AMP; C063509:N-(2-(4-bromocinnamylamino)ethyl)-5-isoquinolinesulfonamide; D002118:Calcium; D015056:1-Methyl-3-isobutylxanthine