There are at least 3 mechanisms involved in thrombin-induced aggregation and release: (1) released ADP, (2) formation of thromboxane A2 and (3) a third mechanism(s). We have examined whether the third pathway is due to formation or release of a substance from platelets which affects other platelets. Washed human platelets were exposed to thrombin (2.5 u/ml) for 15 min at 37°C in the presence of indomethacin to block thromboxane A2 formation. Platelets were removed by centrifugation and the thrombin neutralized with hirudin or DFP. Addition of the superna te to washed human platelets prelabeled with 14C-serotonin caused platelet aggregation but release did not occur. Treatment of the supernate with apyrase, CP/CPK or dialysis abolished aggregation, indicating that the material was ADP. Thus, the mechanism by which thrombin induces aggregation and release with human platelets in the presence of agents which destroy ADP and block the formation of thromboxane A2 is a direct effect of thrombin on platelets and does not involve a substance freed from platelets. In contrast, when washed rabbit platelets were treated with thrombin in the presence of indomethacin and the released ADP was removed, material remained in the supernate which caused aggregation and release from washed rabbit platelets but was without effect on washed human platelets. The activity of this material (MW > 10,000) was not abolished by dialysis or boiling. Therefore rabbit platelets differ from human platelets because they have a mechanism in addition to released ADP, thromboxane A2 and the direct effect of thrombin on platelets that can cause aggregation and release.
The G protein coupled to the thromboxane A2 receptor in human platelets is a member of the novel Gq family
