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1 Cyclic GMP (cGMP) has been shown to be an important modulator of cardiac contractile function. A major component of cGMP regulation of contractility is cGMP-mediated inhibition of the cardiac calcium current (I Ca ). An under-appreciated aspect of cyclic nucleotide signalling is hydrolysis of the cyclic nucleotide (i.e., breakdown by phosphodiesterases (PDEs)). The role of cGMP hydrolysis in regulating I Ca has not been studied. Thus the purpose of this study was to investigate if inhibition of cGMP hydrolysis can modulate I Ca in isolated guinea-pig ventricular myocytes. 2 Zaprinast, a selective inhibitor of cGMP-speci®c PDE (PDE5), caused a signi®cant increase in cGMP levels in myocytes, but was without a ect on basal or b-adrenergic stimulated cAMP levels (consistent with its actions as a speci®c inhibitor of PDE5). 3 Zaprinast inhibited I Ca that was pre-stimulated with cAMP elevating agents (isoproterenol, a badrenergic agonist; or forskolin, a direct activator of adenylate cyclase). The e ect of zaprinast was greatly reduced by KT5823, an inhibitor of cGMP-dependent protein kinase (PKG). 4 Zaprinast also signi®cantly inhibited basal I Ca when perforated-patch or whole-cell recording with physiological pipette calcium concentration (10 77 M) was used. However, this e ect was not observed when using standard calcium-free whole-cell recording conditions. 5 These results indicate that inhibition of cGMP hydrolysis can decrease both basal and cAMPstimulated I Ca . Thus, cGMP hydrolysis may likely be an important step for physiological modulation of I Ca . This regulation may also be important in disease states in which cGMP production is increased and PDE5 expression is altered, such as heart failure.

The effects of adenosine analogues on beta-adrenergic receptor and receptor-independent elicited increases in adenylate cyclase activity were investigated using membranes obtained from primary cultures of adult rat ventricular myocytes. Phenylisopropyladenosine, an A1-receptor agonist, at concentrations of 0.1, 1.0, and 10 microM, maximally inhibited isoproterenol-stimulated adenylate cyclase activity by 35, 55, and 41%, respectively. The inhibition by phenylisopropyladenosine was antagonized by 10 microM theophylline. One micromolar phenylisopropyladenosine was much less effective at attenuating forskolin-stimulated activity, such that the maximum inhibition was 26%. Phenylisopropyladenosine had no effect on adenylate cyclase stimulation by 5'-guanylylimidodiphosphate. Phenylaminoadenosine, an A2 agonist, at 10 microM or greater stimulated adenylate cyclase activity. This effect was not significantly inhibited by theophylline or 0.1 microM 1,3-dipropyl-8-cyclopentylxanthine (DPCPX), which antagonized phenylisopropyladenosine inhibition of isoproterenol-stimulated adenylate cyclase activity. Additionally, N-ethylcarboxamidoadenosine, a nonselective adenosine-receptor agonist, had no effect on adenylate cyclase activity in the absence of DPCPX but stimulated adenylate cyclase activity in the presence of DPCPX. These results indicate that both A1 and A2 receptors exist on the ventricular myocyte sarcolemma. More importantly, the findings suggest that adenosine inhibition of catecholamine-stimulated adenylate cyclase activity is primarily due to an alteration in beta-adrenergic receptor-mediated transduction and perhaps in part by a direct inhibition of the catalytic component.

The contractile response of the aged adult heart to j3-adrenergic stimulation is known to be reduced compared with the young adult heart. Since endogenous adenosine exerts an antiadrenergic action in the heart, this study was undertaken to determine if the basal endogenous level of myocardial adenosine increases with age and whether this increase mediates the reduced responsiveness of aged heart to 3-adrenergic stimulation. Young (3-5 months) and aged (12-22 months) Sprague-Dawley adult rat hearts of CD and SD stock were perfused at constant pressure and paced at 270 contractions/min. The two age groups had a similar level of +dP/dtmax (index of contractility) under control conditions. Adenosine release into the coronary effluent was 30±3 nmol/min/g dry wt from young and 54±9 nmol/min/g dry wt from aged hearts. Inosine release was also greater from the aged hearts. Isoproterenol (10-8 M) stimulation increased contractile state by 113% in young hearts and only 69o in aged hearts. Isoproterenol further increased the adenosine and inosine release from both age groups. Theophylline (5 X iO-M), an adenosine antagonist, prevented the difference in the contractile response to isoproterenol stimulation between the young and aged hearts. Elevation of external calcium from 2 to 4 mM increased contractility equally in both age groups without influencing adenosine release. Myocardial oxygen consumption, coronary effluent Po2, oxygen supplydemand ratio, and lactate release were similar for both age groups, indicating that under the conditions studied the elevated release of adenosine by the aged hearts was not due to hypoxia. Aged (10-14 months) adult guinea pig hearts also displayed a reduced responsiveness to the isoproterenol stimulation and released more adenosine compared with young (3-4 months) adult guinea pig hearts. These findings suggest that enhanced adenosine levels that are present in the aged myocardium are responsible, in part, for the reduced contractile responsiveness of the older adult heart to j3-adrenergic stimulation. (Circulation Research 1990;66:1381-1390 As adult mammals age, profound changes occur in cardiovascular function. For example, augmentation of cardiac contractile performance during stress is diminished in rats1'2 and humans.3'4 Normally, during stress the contractile performance of the heart is increased as a result of sympathetic nervous system-mediated release of the neurotransmitter norepinephrine and enhanced levels of circulating epinephrine. These catecholamines interact with myocardial ,B-adrenoceptors, thereby actiFrom the Department of Physiology, University of Massachusetts Medical School, Worcester, Massachusetts.Presented as a preliminary report in abstract form (Fed Proc 1986;45:750).Supported primarily by HL-36964 and in part by HL-22828 Public Health Service grants from the National Institutes of Health.Address for reprints: James G. Dobson Jr., PhD, Department of Physiology, University of Massachusetts Medical School, 55 Lake Avenue North, Worcester, MA 01655.Received No...

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