Subcellular distribution of high-affinity type IV cyclic AMP phosphodiesterase activity in rabbit ventricular myocardium: relations to the effects of cardiotonic drugs.
Rabbit ventricular myocardium contains distinct cytosolic and particulate cyclic AMP (cAMP) phosphodiesterase activities that exhibit characteristics ascribed to a high-affinity type IV cAMP phosphodiesterase activity found in several tissues. The particulate activity associated with sarcoplasmic reticulum vesicles has an apparent Km for cAMP of about 0.3 microM and a Vmax of 2.45 +/- 0.55 nmol/min/mg. Cyclic GMP (cGMP) inhibits hydrolysis measured at 0.25 microM cAMP with an IC50 value of 0.28 microM. In comparison, a ventricular cytosolic high-affinity cAMP phosphodiesterase activity obtained by anion exchange chromatography (Peak III) has an apparent Km of 0.93 microM and a Vmax of 17 +/- 1 nmol/min/mg. Hydrolysis of 0.25 microM cAMP by this cytosolic activity is weakly inhibited by cGMP with an IC50 value of 142 microM. Particulate enzyme activity is 60-fold more sensitive to inhibition by milrinone than is the cytosolic form (Ki = 0.18 versus 11 microM, respectively); the pyridazinone imazodan is a 12-fold more potent inhibitor of the particulate activity than of the cytosolic form (Ki = 1.5 versus 18 microM, respectively). Inhibition of both cytosolic and particulate enzyme activities appears competitive in nature. Solubilization of particulate activity did not significantly alter its affinity for substrate or sensitivity to inhibition by cGMP. In the presence of a submaximally activating concentration of forskolin (0.4 microM), selective phosphodiesterase inhibitors potentiated the activation of protein kinase in isolated ventricular septal slices. Under these conditions, changes in cAMP-dependent protein kinase activity ratios correlated more closely with contractile responses than did changes in intracellular content of cAMP.(ABSTRACT TRUNCATED AT 250 WORDS)
Kithas
P A
PA
Department of Pharmacology, College of Medicine, University of South Alabama, Mobile.
Artman
M
M
Thompson
W J
WJ
Strada
S J
SJ
D015105:3',5'-Cyclic-AMP Phosphodiesterases / Q000145:classification / Q000378:metabolism*; D000818:Animals; D002147:Calmodulin / Q000378:metabolism; D002316:Cardiotonic Agents / Q000494:pharmacology*; D003600:Cytosol / Q000201:enzymology; D004789:Enzyme Activation / Q000187:drug effects; D006352:Heart Ventricles; D009200:Myocardial Contraction / Q000187:drug effects; D009206:Myocardium / Q000201:enzymology*; D011494:Protein Kinases / Q000378:metabolism; D011817:Rabbits; D012519:Sarcoplasmic Reticulum / Q000201:enzymology; D013347:Subcellular Fractions / Q000201:enzymology*
D002147:Calmodulin; D002316:Cardiotonic Agents; D011494:Protein Kinases; D015105:3',5'-Cyclic-AMP Phosphodiesterases