title: Modulation of Ca2+-dependent Cl- channels by calcineurin in rabbit coronary arterial myocytes.
abstract: The role of the Ca2+-dependent phosphatase calcineurin (CaN) in the modulation of Ca2+-dependent Cl- channels (ClCa) was studied in freshly isolated rabbit coronary arterial myocytes. Immunocytochemical experiments showed that calmodulin-dependent protein kinase II (CaMKII) and CaN were distributed evenly throughout the cytoplasm of coronary myocytes at rest and translocated to the plasmalemma when intracellular Ca2+ was increased. ClCa currents (ICl(Ca)) elicited by cell dialysis with fixed intracellular Ca2+ levels up to 500 nM were inhibited by 10 microM cyclosporin A (CsA), a specific inhibitor of CaN, in a voltage-dependent manner, whereas currents evoked by 1 microM Ca2+ were not affected. Inhibition of CaN with CsA also led to a significant reduction in Ca2+ sensitivity of the channel at +50 mV; half-maximal activation increased from 363 +/- 16 nM Ca2+ in control to 515 +/- 40 nM Ca2+ in the presence of CsA. Similar effects were observed on ICl(Ca) when a specific peptide fragment inhibitor of CaN (CaN-AF, 5 microM) was dialysed into the cell via the pipette (500 nM Ca2+). Application of KN-93 (10 microM), a specific inhibitor of CaMKII, enhanced ICl(Ca) in myocytes dialysed with 1 microM Ca2+ but produced no significant effect on this current when the cells were dialysed with 350 or 500 nM Ca2+. These results are consistent with the notion that in coronary arterial cells, the activity of ClCa is enhanced by dephosphorylation of the channel or a closely associated regulatory protein. Moreover the balance of CaN and CaMKII regulating ICl(Ca) is dependent on the level of Ca2+ used to activate ICl(Ca).
authors.affiliation: Department of Physiology, University of Montréal and Research Centre, Montréal Heart Institute, Montréal, Québec, Canada.
authors.forename: Jonathan
authors.initials: J
authors.lastname: Ledoux
authors.forename: Iain
authors.initials: I
authors.lastname: Greenwood
authors.forename: Louis R
authors.initials: LR
authors.lastname: Villeneuve
authors.forename: Normand
authors.initials: N
authors.lastname: Leblanc
mesh_terms: D000818:Animals; D001158:Arteries; D001596:Benzylamines / Q000494:pharmacology; D019703:Calcineurin / Q000378:metabolism / Q000502:physiology*; D065095:Calcineurin Inhibitors; D002118:Calcium / Q000378:metabolism*; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D018118:Chloride Channels / Q000187:drug effects / Q000378:metabolism* / Q000502:physiology; D003331:Coronary Vessels / Q000166:cytology / Q000378:metabolism*; D016572:Cyclosporine / Q000494:pharmacology; D004553:Electric Conductivity; D004791:Enzyme Inhibitors / Q000494:pharmacology; D007150:Immunohistochemistry; D009131:Muscle, Smooth, Vascular / Q000378:metabolism*; D032389:Myocytes, Smooth Muscle / Q000378:metabolism*; D010446:Peptide Fragments / Q000494:pharmacology; D011817:Rabbits; D013449:Sulfonamides / Q000494:pharmacology; D000096926:Benzenesulfonamides
chemical_list: D001596:Benzylamines; D019703:Calcineurin; D065095:Calcineurin Inhibitors; D002118:Calcium; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D018118:Chloride Channels; D016572:Cyclosporine; D004791:Enzyme Inhibitors; D010446:Peptide Fragments; D013449:Sulfonamides; C487831:calcineurin-AF; C072105:KN 93; D000096926:Benzenesulfonamides