To examine whether activation of polymorphonuclear leukocytes attenuates release of adenosine through attenuation of their own ecto-5'-nucleotidase activity, human polymorphonuclear leukocytes were incubated with and without exposure to either N-formyl-methionyl-leucyl-phenylalanine (FMLP) or complement C5a. Ecto-5'-nucleotidase activity of polymorphonuclear leukocytes was attenuated by both FMLP and complement C5a (22.7 +/- 3.6 vs 9.7 +/- 2.6 nmol/min per 10(7) cells at 10(-6) M FMLP, P < .05; 21.5 +/- 2.2 vs 10.2 +/- 1.2 nmol/min per 10(7) cells at 5 x 10(-7) g/mL complement C5a, P < .001), whereas cytosolic 5'-nucleotidase activity was not affected by either FMLP or complement C5a. These reductions of ecto-5'-nucleotidase activity that were caused by both FMLP and complement C5a were dose and time dependent and were inhibited by superoxide dismutase. Desferrioxamine did not inhibit the decreases in ecto-5'-nucleotidase. In accordance with the decreases in ecto-5'-nucleotidase activity, release of adenosine was attenuated in the FMLP-pretreated and complement C5a-pretreated polymorphonuclear leukocytes, which were restored by concomitant administration of superoxide dismutase. The viability of FMLP-pretreated and complement C5a-pretreated polymorphonuclear leukocytes was markedly decreased compared with the untreated group after 60 minutes of hypoxia followed by 60 minutes of reoxygenation. Thus, we conclude that: (1) activation of polymorphonuclear leukocytes attenuates their own ecto-5'-nucleotidase activity and thereby reduces adenosine release, (2) reduction of ecto-5'-nucleotidase activity is attributable to generated superoxide anion in polymorphonuclear leukocytes, and (3) viability of polymorphonuclear leukocytes after hypoxia and reoxygenation largely depends on the extents of decreases in ecto-5'-nucleotidase activity.