462
Biochimica et Biophysica Acta, 544 (1978) 462--473 © Elsevier/North-Holland Biomedical Press
BBA 28749
PROPERTIES OF EPINEPHRINE-INDUCED ACTIVATION OF CARDIAC ADENOSINE 3',5'-MONOPHOSPHATE-DEPENDENT PROTEIN KINASE
DONNA F. BROWN *, THOMASW. HONEYMANand JAMES G. DOBSONJr. ** Department of Physiology, University of Massachusetts Medical School, Worcester, Mass. 01605 (U.S.A.)
(Received May 30th, 1978)
Summary The effects of epinephrine on cyclic AMP content and protein kinase activity were examined in an in situ rat heart preparation. Bolus injection of epinephrine into the superior vena cava caused an increase in the activity ratio (--cyclic AMP/+cyclic AMP) of 12 000 × g supematant protein kinase. The increase was significant within 5 s and maximal in 10 s. Epinephrine produced a dose-depen'dent increase in both protein kinase activity ratio and cyclic AMP content. The increases in both parameters exhibited a high degree of correlation. The increase in protein kinase activity ratio observed with low doses of epinephrine (less than or equal to 1 #g/kg) resulted from an increase in independent protein kinase activity (--cyclic AMP) without a change in total protein kinase activity (+cyclic AMP). However, the increase in the activity ratio observed with higher doses of epinephrine (greater than 1/~g/kg) was due mainly to a decrease in total protein kinase activity rather than a further increase in independent protein kinase activity. The loss of supematant total protein kinase activity could be accounted for by an increase in activity associated with particulate fractions obtained from the homogenates. A similar redistribution of protein kinase could be demonstrated by the addition of cyclic AMP to homogenates prepared from hearts not stimulated with epinephrine. These results demonstrate that epinephrine over a wide dose range produces a parallel increase in the content of cyclic AMP and the activation of soluble protein kinase. The findings also suggest that protein kinase translocation to particulate material may depend on the degree of epinephrine~induced enzyme activation.
* P r e s e n t a d d r e s s : D e p a r t m e n t o f B i o c h e m i s t r y , Medical College of O h i o , C.S. No. 1 0 0 0 8 , T o l e d o , Ohio 43689, U.S.A. * * T o w h o m c o r r e s p o n d e n c e s h o u l d b e sent.
463
Introduction Adenosine 3',5'-monophosphate (cyclic AMP) and cyclic AMP-dependent protein kinase have been suggested to mediate the activation of cardiac glycogenolysis and the augmentation of myocardial contractility produced by cate~ cholamine stimulation [1--3]. Cyclic AMP-dependent protein kinase activation is thought to be important in the catecholamine-induced activation of phosphorylase presumably by catalyzing the transformation of phosphorylase kinase to its activated form [4--6]. The inotropic response caused by the catecholamines has been suggested to involve cyclic AMP
Biochimica et Biophysica Acta, 544 (1978) 462--473 © Elsevier/North-Holland Biomedical Press
BBA 28749
PROPERTIES OF EPINEPHRINE-INDUCED ACTIVATION OF CARDIAC ADENOSINE 3',5'-MONOPHOSPHATE-DEPENDENT PROTEIN KINASE
DONNA F. BROWN *, THOMASW. HONEYMANand JAMES G. DOBSONJr. ** Department of Physiology, University of Massachusetts Medical School, Worcester, Mass. 01605 (U.S.A.)
(Received May 30th, 1978)
Summary The effects of epinephrine on cyclic AMP content and protein kinase activity were examined in an in situ rat heart preparation. Bolus injection of epinephrine into the superior vena cava caused an increase in the activity ratio (--cyclic AMP/+cyclic AMP) of 12 000 × g supematant protein kinase. The increase was significant within 5 s and maximal in 10 s. Epinephrine produced a dose-depen'dent increase in both protein kinase activity ratio and cyclic AMP content. The increases in both parameters exhibited a high degree of correlation. The increase in protein kinase activity ratio observed with low doses of epinephrine (less than or equal to 1 #g/kg) resulted from an increase in independent protein kinase activity (--cyclic AMP) without a change in total protein kinase activity (+cyclic AMP). However, the increase in the activity ratio observed with higher doses of epinephrine (greater than 1/~g/kg) was due mainly to a decrease in total protein kinase activity rather than a further increase in independent protein kinase activity. The loss of supematant total protein kinase activity could be accounted for by an increase in activity associated with particulate fractions obtained from the homogenates. A similar redistribution of protein kinase could be demonstrated by the addition of cyclic AMP to homogenates prepared from hearts not stimulated with epinephrine. These results demonstrate that epinephrine over a wide dose range produces a parallel increase in the content of cyclic AMP and the activation of soluble protein kinase. The findings also suggest that protein kinase translocation to particulate material may depend on the degree of epinephrine~induced enzyme activation.
* P r e s e n t a d d r e s s : D e p a r t m e n t o f B i o c h e m i s t r y , Medical College of O h i o , C.S. No. 1 0 0 0 8 , T o l e d o , Ohio 43689, U.S.A. * * T o w h o m c o r r e s p o n d e n c e s h o u l d b e sent.
463
Introduction Adenosine 3',5'-monophosphate (cyclic AMP) and cyclic AMP-dependent protein kinase have been suggested to mediate the activation of cardiac glycogenolysis and the augmentation of myocardial contractility produced by cate~ cholamine stimulation [1--3]. Cyclic AMP-dependent protein kinase activation is thought to be important in the catecholamine-induced activation of phosphorylase presumably by catalyzing the transformation of phosphorylase kinase to its activated form [4--6]. The inotropic response caused by the catecholamines has been suggested to involve cyclic AMP
