Calcineurin Aalpha but not Abeta augments ICl(Ca) in rabbit pulmonary artery smooth muscle cells.
Activation of Ca(2+)-dependent Cl(-) currents (I(Cl(Ca))) increases membrane excitability in vascular smooth muscle cells. Previous studies showed that Ca(2+)-dependent phosphorylation suppresses I(Cl(Ca)) in pulmonary artery myocytes, and the aim of the present study was to determine the role of the Ca(2+)-dependent phosphatase calcineurin on chloride channel activity. Immunocytochemical and Western blot studies with isoform-specific antibodies revealed that the alpha and beta forms of the CaN catalytic subunit are expressed in PA cells but that only the alpha variant translocated to the cell periphery upon a rise in intracellular [Ca(2+)]. I(Cl(Ca)) evoked by pipette solutions containing a [Ca(2+)] set at 500 nm was considerably larger when the pipette solution included constitutively active CaN containing the alpha catalytic isoform. This stimulatory effect was lost by boiling the enzyme or by the inclusion of a specific CaN inhibitory peptide and was not shared by the inclusion of the beta form of the catalytic subunit. In the absence of constitutively active CaN, cyclosporin A, an inhibitor of CaN, suppressed I(Cl(Ca)) evoked by 500 nm Ca(2+) when the current amplitude was relatively large but was ineffective in cells with smaller currents. In perforated patch recordings, cyclosporin A consistently inhibited I(Cl(Ca)) evoked as a consequence of Ca(2+) influx through voltage-dependent calcium channels. These novel data show that in PA myocytes activation of I(Cl(Ca)) is enhanced by Ca(2+)-dependent dephosphorylation and that the regulation of this conductance is highly isoform-specific.
Greenwood
Iain A
IA
Department of Basic Medical Sciences, Pharmacology & Clinical Pharmacology, St. George's Hospital Medical School, London SW17 ORE, United Kingdom.
Ledoux
Jonathan
J
Sanguinetti
Amy
A
Perrino
Brian A
BA
Leblanc
Normand
N
D000595:Amino Acid Sequence; D000818:Animals; D015153:Blotting, Western; D001921:Brain / Q000378:metabolism; D019703:Calcineurin / Q000378:metabolism*; D002118:Calcium / Q000378:metabolism; D015220:Calcium Channels / Q000378:metabolism; D020134:Catalytic Domain; D002460:Cell Line; D002478:Cells, Cultured; D018118:Chloride Channels / Q000737:chemistry; D003062:Codon; D016572:Cyclosporine / Q000494:pharmacology; D018076:DNA, Complementary / Q000378:metabolism; D004594:Electrophysiology; D004730:Endothelium, Vascular / Q000166:cytology; D007150:Immunohistochemistry; D007313:Insecta; D018613:Microscopy, Confocal; D008856:Microscopy, Fluorescence; D008969:Molecular Sequence Data; D032389:Myocytes, Smooth Muscle / Q000166:cytology*; D010455:Peptides / Q000737:chemistry; D011485:Protein Binding; D020033:Protein Isoforms; D017434:Protein Structure, Tertiary; D011651:Pulmonary Artery / Q000378:metabolism*; D011817:Rabbits; D017386:Sequence Homology, Amino Acid
D015220:Calcium Channels; D018118:Chloride Channels; D003062:Codon; D018076:DNA, Complementary; D010455:Peptides; D020033:Protein Isoforms; D016572:Cyclosporine; D019703:Calcineurin; D002118:Calcium