CAMP CAN RAISE OR LOWER CARDIAC ACTOMYOSIN ATPASE ACTIVITY DEPENDING ON ALPHA-ADRENERGIC ACTIVITY

Adenosine 3',5'-cyclic monophosphate (cAMP) or beta-adrenergic stimulation has been shown to increase actomyosin adenosinetriphosphatase (ATPase) activity in cardiac muscle. Because the major catecholamine transmitters have both alpha- and beta-adrenergic activity, the possibility of a role for alpha-adrenergic stimulation in the regulation of ATPase activity has been investigated. Histochemical measurement of actomyosin ATPase activity in quickly frozen rat hearts has been used as the assay of enzymatic function of the contractile proteins. The dose-response curve of ATPase activity to cAMP shows an increase in ATPase activity at a threshold concentration of 0.01 mu M, a peak effect at 0.5-1.0 mu M, and a decline beyond 1.5 mu M to a level below control at 10 mu M cAMP. Kinetic studies varying ATP concentration from 0.5 to 10 mM indicated the existence of multiple forms of actomyosin ATPase activity in the absence of cAMP and only one form with a higher maximum velocity in the presence of 1 mu M cAMP. Apparently cAMP raises the enzymatic activity of the individual actomyosin molecule rather than increasing the number of active molecules. The addition of an a-adrenergic blocker had no significant effect in the absence of added cAMP, but in the presence of the cyclic nucleotide, 1 mu M prazosin always produced a negative effect on ATPase activity. Over the entire range of 0.01-10 mu M, cAMP lowered ATPase activity when the alpha-adrenergic antagonist was present. The integrity of the cAMP regulatory system was sensitive to the tissue oxygen tension at the time the heart was quickly frozen. At certain oxygen tension, the stimulatory component of the cAMP regulation was observed without any inhibitory component, suggesting that there are two relatively independent parts of the regulatory mechanism, an inhibitory and a stimulatory. In the presence of gamma-labeled [P-32]ATP, P-32 was incorporated into several proteins, including the inhibitory subunit of troponin (TNI), C protein, and the regulatory light chain of myosin. cAMP (1 mu M) caused an increase in P-32 labeling of TNI and C protein. The addition of prazosin with cAMP caused a decrease in the overall level of phosphorylation with specific dephosphorylation of C protein and TNI, the former to a degree similar to the decrease in actomyosin ATPase activity, the latter to a greater degree. These results indicate that a-adrenergic activity modulates the balance between kinase and phosphatase activity in the presence of cAMP, probably by inhibiting phosphatase activity. A model to integrate these data involving protein kinase A phosphorylation of C protein, a guanosine 3',5'-cyclic monophosphate-activated phosphatase that is also activated by higher concentrations of cAMP, and a protein kinase C-dependent inhibition of phosphate activity controlling the alpha-adrenergic system is proposed.