Ca /Calmodulin Kinase II–Dependent Phosphorylation of Ryanodine Receptors Suppresses Ca Sparks and Ca Waves in Cardiac Myocytes

The multifunctional Ca /calmodulin-dependent protein kinase II C (CaMKII C) is found in the macromolecular complex of type 2 ryanodine receptor (RyR2) Ca release channels in the heart. However, the functional role of CaMKII-dependent phosphorylation of RyR2 is highly controversial. To address this issue, we expressed wild-type, constitutively active, or dominant-negative CaMKII C via adenoviral gene transfer in cultured adult rat ventricular myocytes. CaMKII-mediated phosphorylation of RyR2 was reduced, enhanced, or unaltered by dominant-negative, constitutively active, or wild-type CaMKII C expression, whereas phosphorylation of phospholamban at Thr17, an endogenous indicator of CaMKII activity, was at 73%, 161%, or 115% of the control group expressing -galactosidase ( -gal), respectively. In parallel with the phospholamban phosphorylation, the decay kinetics of global Ca transients was slowed, accelerated, or unchanged, whereas spontaneous Ca spark activity was hyperactive, depressed, or unchanged in dominant-negative, constitutively active, or wild-type CaMKII C groups, respectively. When challenged by high extracellular Ca , both wild-type and constitutively active CaMKII C protected the cells from store overload–induced Ca release, manifested by a 60% suppression of Ca waves (at 2 to 20 mmol/L extracellular Ca ) in spite of an elevated sarcoplasmic reticulum Ca content, whereas dominant-negative CaMKII C promoted Ca wave production (at 20 mmol/L Ca ) with significantly depleted sarcoplasmic reticulum Ca . Taken together, our data support the notion that CaMKII C negatively regulates RyR2 activity and spontaneous sarcoplasmic reticulum Ca 2 release, thereby affording a negative feedback that stabilizes local and global Ca -induced Ca release in the heart. (Circ Res. 2007;100:399-407.)