Adenosine via an adenosine A1 receptor (A1R) is a negative feedback inhibitor of adrenergic stimulation in the heart, protecting it from toxic effects of overstimulation. Stimulation of the A 1R results in the activation of Gi protein, release of free G␤␥-subunits, and activation/translocation of PKC-ε to the receptor for activated C kinase 2 protein at the Z-line of the cardiomyocyte sarcomere. Using an anti-G␤␥ peptide, we investigated the role of these subunits in the A1R stimulation of phospholipase C (PLC), with the premise that the resulting diacylglycerol provides for the activation of PKC-ε. Inositol 1,4,5-triphosphate release was an index of PLC activity. Chlorocyclopentyl adenosine (CCPA), an A1R agonist, increased inositol 1,4,5-triphosphate production by 273% in mouse heart homogenates, an effect absent in A1R knockout hearts and inhibited by anti-G␤␥ peptide. In a second study, p38 MAPK and heat shock protein 27 (HSP27), found by others to be associated with the loss of myocardial contractile function, were postulated to play a role in the actions of A1R. Isoproterenol, a ␤-adrenergic receptor agonist, increased the Ca 2ϩ transient and sarcomere shortening magnitudes by 36 and 49%, respectively. In the rat cardiomyocyte, CCPA significantly reduced these increases, an action blocked by the p38 MAPK inhibitor SB-203580. While CCPA significantly increased the phosphorylation of HSP27, this action was inhibited by isoproterenol. These data indicate that the activation of PKC-ε by A1R results from the activation of PLC via free G␤␥-subunits released upon A1R-induced dissociation of Gi␣␤␥. Attenuation of ␤-adrenergic-induced contractile function by A1R may involve the activation of p38 MAPK, but not HSP27. G␤␥ subunits; antiadrenergic; rodent; contractility ADENOSINE IS RELEASED ENDOGENOUSLY in the heart that is stressed by hypoxia, ischemia, or adrenergic stimulation (13,15). In the interstitium, adenosine activates cell surface receptors that initiate signal cascades within the cardiomyocyte, ultimately reducing the cardiotoxic manifestations of the initial insult (16,43). For many years, study has focused on the antiadrenergic action of adenosine, or the ability of this nucleoside to protect the heart against excessive stimulation (34) by norepinephrine released during ischemia (36). In manifesting this action, adenosine A 1 receptors (A 1 R) attenuate ␤-adrenergic catecholamine-elicited increases in G s protein cycling (17), adenylyl cyclase activity (33), cAMP formation (5), activation of protein kinase A (PKA) (6), protein phosphorylation (12), myocardial Ca 2ϩ transient magnitude (14), and ventricular contractility (6, 16).Adenosine also reduces the responsiveness of the myocardium to catecholamine stimulation by a mechanism independent of changes in adrenergic-stimulated cyclase activity. This adenoprotection by the A 1 R agonist chlorocyclopentyl adenosine (CCPA) has been demonstrated by us to involve the translocation of PKC-ε to receptor for activated C kinase 2 (RACK2) in rat and mouse myocard...

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