Cyclooxygenase products are thought to mediate adenosine-stimulated contraction of nonpregnant myometrium. We have examined the effects of the cyclooxygenase inhibitors, indomethacin and meclofenamate, upon adenosine-stimulated contractions in isolated, endometrium-free strips of uterine smooth muscle from virgin guinea pigs. Indomethacin (30 microM) had no effect on adenosine-induced contractions; addition of meclofenamate at 30 microM, however, rapidly and reversibly blocked contractions in response to adenosine, yet had no effect upon acetylcholine-stimulated contractions. Application of prostaglandin F2 alpha at a final concentration of 1.1 microM elicited contractions in indomethacin-treated (but not meclofenamate-treated) tissues. The adenosine receptor antagonist, 8-(p-sulfophenyl)theophylline, interfered with adenosine-mediated contractions when present at a final concentration of 100 microM, confirming the presence of extracellular binding sites for the purine on the smooth muscle. Treatment of muscle strips with either of the adenosine uptake blockers, dipyridamole or S-(p-nitrobenzyl)-6-thio-guanosine, resulted in dramatic reductions in adenosine-stimulated contractions, findings consistent with an additional, intracellular site of action of adenosine in uterine smooth muscle. These results suggest that, in contrast to findings in intact strips of whole uterine tissue, cyclooxygenase activity is not required for adenosine-stimulated contraction in isolated myometrium from nonpregnant guinea pig. It also is suggested that the nonsteroidal anti-inflammatory compound, meclofenamate, may interfere with the binding of another (non-cyclooxygenase) product of arachidonic acid metabolism.