The effects of adenosine analogues on pain have been shown to depend on the subtype receptor involved as well as on the nociceptive stimuli and on the route of administration. In the first experiment of the present study intraperitoneal administration of the A(1) receptor agonist N(6)-cyclopentyladenosine (CPA) (0.015, 0.03, 0.09, 0.15, 0.21, 0.3 mg/kg) induced dose-dependent analgesia to formalin pain in both phases characterizing the test. The A(2a) receptor agonist 2-[p-2-(carbonyl-ethyl)-phenyethylamino]-5'-N-ethylcarboxaminoadenosine (CGS21680) (0.025, 0.05, 0.1, 0.15 mg/kg) significantly affected behavioral responses to formalin only during the early phase. In the second experiment the interaction between adenosine and the opioid system was investigated through both behavioral and neurochemical studies. The opioid antagonist naltrexone (0.1 mg/kg) did not affect the antinociception induced by CPA (0.21 mg/kg) and CGS21680 (0.05 mg/kg). Autoradiographic studies showed that formalin administration significantly modified mu-opioid receptor density in the superficial laminae of the spinal cord and in the paracentral thalamic nucleus, contralateral to the side of formalin injection. CPA and CGS21680 counteracted these effects induced by formalin. In conclusion the present study confirms and extends the role of A(1) and A(2a) adenosine receptors in the modulation of inflammatory pain and their interaction with the mu-opioid system, and suggests further investigation of these purinergic receptors from a therapeutic perspective.