Effects of felodipine (a dihydropyridine calcium channel blocker) and analogues on calmodulin-dependent enzymes.
We have examined the effects on the activities of three calmodulin-dependent enzymes (cAMP phosphodiesterase, caldesmon kinase and myosin light chain kinase) of the dihydropyridine Ca2+ channel blocker felodipine and three analogues (p-chloro, oxidized and t-butyl) exhibiting different pharmacological potencies. The cAMP phosphodiesterase was inhibited completely by felodipine and the p-chloro analogue with IC50 values of 3.7 and 1.5 microM respectively. The oxidized and t-butyl analogues were relatively ineffective in inhibiting cAMP phosphodiesterase. Felodipine and the p-chloro analogue inhibited the basal (Ca2+/calmodulin-independent) activity of cAMP phosphodiesterase as well as the calmodulin-stimulated activity. Calmodulin was relatively ineffective in preventing inhibition of cAMP phosphodiesterase by felodipine and the p-chloro analogue. These observations suggest that felodipine may act directly on the phosphodiesterase as well as through calmodulin. Felodipine and the p-chloro analogue inhibited Ca2+/calmodulin-dependent caldesmon kinase with similar potencies (IC50 = 17.4 microM), whereas the oxidized and t-butyl analogues caused no inhibition. Similarly, felodipine and the p-chloro analogue inhibited myosin light chain kinase activity whether the isolated 20 kD light chain (IC50 = 12.6 microM) or intact myosin (IC50 = 11.0 microM) was used as substrate. Inhibition in each case was prevented by excess calmodulin. The oxidized and t-butyl derivatives caused little or no inhibition. Finally, the effects of felodipine and the three analogues on two processes which are dependent on myosin phosphorylation were examined, namely the actin-activated Mg2+-ATPase activity of myosin and the assembly of myosin filaments. Felodipine and the p-chloro analogue inhibited the actin-activated Mg2+-ATPase activity of smooth muscle myosin (IC50 = 25.1 microM). The oxidized and t-butyl analogues exhibited no inhibition. Similarly, felodipine and the p-chloro analogue blocked myosin filament assembly induced by low concentrations of calmodulin, whereas the oxidized and t-butyl analogues did not. Again, inhibition of the actin-activated myosin Mg2+-ATPase and myosin filament assembly by felodipine and the p-chloro analogue could be reversed by raising the calmodulin concentration. These observations suggest that some of the pharmacological actions of felodipine on smooth muscle may involve inhibition of calmodulin-dependent enzymes which are functionally involved in the regulation of smooth muscle contraction.
Walsh
M P
MP
Department of Medical Biochemistry, Faculty of Medicine, University of Calgary, Alberta, Canada.
Sutherland
C
C
Scott-Woo
G C
GC
D015105:3',5'-Cyclic-AMP Phosphodiesterases / Q000037:antagonists & inhibitors*; D000199:Actins / Q000502:physiology; D000818:Animals; D017301:Ca(2+) Mg(2+)-ATPase / Q000037:antagonists & inhibitors; D002121:Calcium Channel Blockers / Q000494:pharmacology*; D002147:Calmodulin / Q000502:physiology*; D002148:Calmodulin-Binding Proteins / Q000378:metabolism; D002645:Chickens; D054677:Cyclic Nucleotide Phosphodiesterases, Type 1; D004591:Electrophoresis, Polyacrylamide Gel; D015736:Felodipine; D009130:Muscle, Smooth / Q000201:enzymology; D009219:Myosin-Light-Chain Kinase / Q000037:antagonists & inhibitors*; D009218:Myosins / Q000378:metabolism; D009568:Nitrendipine / Q000031:analogs & derivatives* / Q000494:pharmacology; D010766:Phosphorylation; D047428:Protein Kinase Inhibitors
D000199:Actins; D002121:Calcium Channel Blockers; D002147:Calmodulin; D002148:Calmodulin-Binding Proteins; D047428:Protein Kinase Inhibitors; D009568:Nitrendipine; D009219:Myosin-Light-Chain Kinase; D015105:3',5'-Cyclic-AMP Phosphodiesterases; D054677:Cyclic Nucleotide Phosphodiesterases, Type 1; D017301:Ca(2+) Mg(2+)-ATPase; D009218:Myosins; D015736:Felodipine