Calcineurin Aα but not Aβ augments ICl(Ca) in rabbit pulmonary artery smooth muscle cells

Activation of Ca2+-dependent Cl- currents (I-Cl(Ca)) increases membrane excitability in vascular smooth muscle cells. Previous studies showed that Ca2+-dependent phosphorylation suppresses I-Cl(Ca) in pulmonary artery myocytes, and the aim of the present study was to determine the role of the Ca2+-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 [Ca2+]. I-Cl(Ca) evoked by pipette solutions containing a [Ca2+] 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 Ca2+ 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 Ca2+ influx through voltage-dependent calcium channels. These novel data show that in PA myocytes activation of I-Cl(Ca) is enhanced by Ca2+-dependent dephosphorylation and that the regulation of this conductance is highly isoform-specific.