Multifunctional Ca2+/calmodulin-dependent protein kinase mediates Ca(2+)-induced enhancement of the L-type Ca2+ current in rabbit ventricular myocytes.
The intracellular mechanism underlying the Ca(2+)-induced enhancement of the L-type Ca2+ current (ICa) was examined in adult rabbit cardiac ventricular myocytes by using patch-clamp methodology. Internal Ca2+ was elevated by flash photolysis of the Ca2+ chelator Nitr 5, and intracellular Ca2+ levels were simultaneously monitored by Fluo 3 fluorescence. Flash photolysis of Nitr 5 produced a rapid (< 1-second) elevation of internal Ca2+, which led to enhancement (39% to 51% above control) of the peak inward Ca2+ current after a delay of 20 to 120 seconds. Internal dialysis of myocytes with synthetic inhibitory peptides derived from the pseudosubstrate (peptide 273-302) and calmodulin binding (peptide 291-317) regions within the regulatory domain of multifunctional Ca2+/calmodulin-dependent protein kinase (CaM kinase) blocked enhancement of ICa produced by elevation of internal Ca2+ but not that produced by beta-adrenergic stimulation. These inhibitory peptides also had no effect on the elevation of internal Ca2+ produced by flash photolysis of Nitr 5. A pseudosubstrate inhibitory peptide derived from protein kinase C had no significant effect on Ca(2+)-dependent enhancement of ICa. We conclude that CaM kinase mediates the Ca(2+)-induced enhancement of ICa in mammalian cardiac myocytes by a mechanism likely involving direct phosphorylation of the L-type Ca2+ channel complex or an associated regulatory protein.
Anderson
M E
ME
Falk Cardiovascular Research Center, Department of Medicine, Stanford University School of Medicine 94305-5401.
Braun
A P
AP
Schulman
H
H
Premack
B A
BA
D000814:Aniline Compounds; D000818:Animals; D002118:Calcium / Q000378:metabolism / Q000502:physiology*; D015220:Calcium Channels / Q000187:drug effects; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000502:physiology*; D002470:Cell Survival; D002614:Chelating Agents / Q000494:pharmacology; D004533:Egtazic Acid / Q000031:analogs & derivatives / Q000494:pharmacology; D005453:Fluorescence; D005456:Fluorescent Dyes; D006352:Heart Ventricles / Q000166:cytology; D066298:In Vitro Techniques; D010782:Photolysis; D011817:Rabbits; D016276:Ventricular Function; D014966:Xanthenes
D000814:Aniline Compounds; D015220:Calcium Channels; D002614:Chelating Agents; D005456:Fluorescent Dyes; D014966:Xanthenes; C052455:nitr 5; C059715:Fluo-3; D004533:Egtazic Acid; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D002118:Calcium