Abstract
It is well-established that in heart, both the L-type Ca2+channel and the cystic fibrosis transmembrane conductance regulator Cl− channel are regulated by cAMP-dependent phosphorylation. However, it is not clear whether both of these channels are regulated in concert by protein kinase A (PKA) or whether there are mechanisms that independently control the phosphorylation of these two PKA targets. The purpose of this study was to compare the effects of various protein phosphatase and protein kinase inhibitors on these two ionic currents (ICa and ICl) in guinea pig ventricular myocytes to gain insight into these questions. We found that both the stimulation and washout of the effects of isoproterenol on ICl are about twice as fast as the effects on ICa, probably because the dephosphorylation reaction for ICl is faster than that for ICa. In contrast, inhibition of protein phosphatases with 10 μm microcystin stimulated both ICa and ICl, but the stimulation of ICl was much slower and smaller than the stimulation of ICa. The effect of microcystin was inhibited by staurosporine (Ki = 171.5 and 161 nm for ICa and ICl, respectively), suggesting that the stimulation was due to a kinase. The kinase was not protein kinase C (PKC) because it was not inhibited by the specific pseudosubstrate inhibitor of PKC, PKC(19–31), and it was not PKA because it was not inhibited by adenosine 3′,5′-cyclic phosphorothioate. These results suggest that although both the Ca2+ and Cl− channels are regulated by cAMP-dependent phosphorylation, another protein kinase may also regulate these channels, and the kinetics of the response of the channels to phosphorylation can be modulated independently by protein phosphatases.
Footnotes
- Received April 8, 1997.
- Accepted June 18, 1997.
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Send reprint requests to: Dr. H. Criss Hartzell, Department of Cell Biology, Emory University School of Medicine, 1648 Pierce Drive, Atlanta, GA 30322-3030. E-mail: criss{at}anatomy.emory.edu
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This work was supported by National Institutes of Health Grant HL21195.
- The American Society for Pharmacology and Experimental Therapeutics
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