In the heart, endogenous adenosine attenuates the -adrenergic-elicited increase in contractile performance via activation of adenosine A₁ receptors. It has been recently reported that this function of adenosine becomes more pronounced with myocardial maturation. The purpose of the present study was to determine whether mature hearts possess a greater sensitivity than immature hearts to this antiadrenergic effect of adenosine. Isolated perfused hearts or atria from immature (ca. 23 days) and mature (ca. 80 days) rats were stimulated with isoproterenol (Iso), a -adrenergic agonist, at 10⁸ M and concomitantly exposed to increasing concentrations of 2-chloroN⁶-cyclopentyladenosine (CCPA), a highly selective and potent adenosine A₁-receptor agonist, from 10¹² to 10⁶ M. CCPA at 10¹⁰-10⁶ M dose dependently reduced the Iso-elicited contractile response more in immature than in mature hearts or atria. At 10⁶ M, CCPA reduced the Iso-elicited contractile response by 103% in immature hearts and by 55% in mature hearts. These effects of CCPA were attenuated by the adenosine A₁-receptor antagonist 8-cyclopentyl-1,3-dipropylxanthine at 10⁷ M. In additional experiments, CCPA exhibited similar effectiveness in reducing the spontaneous heart rate of immature and mature hearts, an effect also mediated by activation of adenosine A₁ receptors. Similar to CCPA, the adenosine A₁-receptor agonist R-N⁶-(2-phenylisopropyl)adenosine reduced the Iso-elicited contractile response more in immature than in mature hearts, albeit with less effectiveness than CCPA. In agreement with these results, CCPA reduced Iso-elicited adenylyl cyclase activity more in immature than in mature hearts. Overall, in contrast with our original hypothesis, these results indicate that immature hearts display greater sensitivity than mature hearts to the antiadrenergic effect of adenosine A₁-receptor activation.