Concurrent studies of oxygen consumption and aggregation in stimulated human platelets

Abstract

A new approach to the study of the “burst” of oxygen consumption in stimulated human platelets is described. Following addition of collagen, thrombin, or arachidonic acid to washed platelets, O2 consumption and aggregation were measured concurrently. In contrast to thrombin and arachidonic acid, collagen induced a calcium-dependent O2 burst after a lag phase of 25-35 sec. Elevation of platelet cAMP levels completely inhibited the oxygen burst and aggregation response. Blockade of energy metabolism also prevented the oxygen burst; however, the aggregation response was not completely suppressed. Eicosatetraynoic acid and indomethacin—inhibitors of arachidonic acid oxygenation—also abolished the collagen-induced O2 burst but not aggregation. These findings indicate that the platelet O2 burst is utilized for formation of prostaglandin endoperoxides, which are apparently not essential for collagen-induced aggregation. Comparison of the oxygen consumption and aggregation curves elicited by arachidonic acid in the presence of increasing amounts of ADP scavengers demonstrated the obligatory participation of released ADP in aggregation induced by this fatty acid. Stored platelets did not exhibit an oxygen burst when challenged by collagen or thrombin, although the aggregation response was relatively intact. In contrast, addition of arachidonic acid to stored platelets elicited both an O2 burst and aggregation, suggesting that phospholipase A2 activity, or steps leading to its activation, are more labile to storage than the platelet cyclooxygenase. It is concluded that concurrent comparison of oxygen consumption and aggregation responses permits a direct evaluation of the role of endoperoxide formation in human platelet aggregation.