Nine C2-substituted adenosine analogues that are potent and selective for the A2-adenosine receptor were tested for their ability to induce relaxations of the guinea pig aorta. Compounds tested were 2-phenylethoxyadenosine (PEA), 2-phenylethoxy-5'-N-ethylcarboxamidoadenosine (PENECA), 2-cyclohexylethoxyadenosine (CEA), 2-fluorophenylethoxyadenosine (FPEA), 2-methoxyphenylethoxyadenosine (MPEA), 2-naphthylethoxyadenosine (NEA), 2-phenylaminoadenosine (CV-1808), 2-phenylethylaminoadenosine (PEAA) and 2-carboxyethylphenethylamino-5'-N-ethylcarboxamidoadenosine (CGS21680). The responses to these agents were compared to those of three standard adenosine receptor agonists, 5'-N-ethylcarboxamidoadenosine (NECA), N6-cyclohexyladenosine (CHA) and R-N6-phenylisopropyladenosine (R-PIA). The C2-ethoxyadenosine analogues were 30- to 140-fold less potent than NECA and the C2-amino-substituted analogues were 250 to 1000-fold less potent than NECA at inducing relaxations of the guinea pig aorta. All of the analogues were also less potent than the A1-selective agonist R-PIA. However, only responses to NECA were competitively antagonized by the non-selective adenosine receptor antagonist 8-phenyltheophylline (8-PT), pKB = 6.83 +/- 0.05. The results suggest that the C2-substituted analogues produce relaxations of the guinea pig aorta through a combination of actions at A2-adenosine receptors and at xanthine resistant sites. The lack of potency of these analogues at activating the xanthine sensitive A2-receptors in the guinea pig aorta suggests that these adenosine receptors may be of the A2b-subtype.