5'-N-ethylcarboxamideadenosine (NECA) greater than 2-chloroadenosine greater than adenosine greater than (-)-N6-(R-phenyl-isopropyl)-adenosine [(-)-R-PIA] greater than (+)-N6-(S-phenyl-isopropyl)-adenosine [(+)-S-PIA] inhibited in vitro human platelet aggregation in a dose-dependent fashion. 6-nitrobenzylthioinosine and dipyridamole, which inhibit adenosine uptake, and erythro-9-(2-hydroxy-3-nonyl)-adenine, which blocks adenosine metabolism, did not impair the inhibition induced by NECA and adenosine. 8-phenyltheophylline and theophylline, two competitive antagonists of adenosine receptors, blocked the inhibition of platelet aggregation caused by NECA and adenosine. NECA greater than 2-chloroadenosine greater than adenosine greater than (-)-R-PIA greater than (+)-S-PIA increased platelet cyclic adenosine monophosphate (cAMP) levels in a dose-dependent fashion. A significant linear correlation (r = 0.70, p less than 0.001) was found between the increase of platelet cAMP and the inhibition of platelet aggregation induced by adenosine and its analogs. 8-phenyltheophylline, which is a competitive antagonist of adenosine in platelets, also blocked the cAMP accumulation caused by NECA. These data suggest that NECA and other adenosine analogs activate a specific cell surface adenylate cyclase-linked adenosine receptor whose properties are similar to those of an adenosine A2/Ra receptor.