Further Evidence of the Involvement of Thrombin in Platelet Release by ADP and Other Agents
There is evidence of the involvement of coagulation factors in platelet aggregation and the release reaction caused by ADP and collagen but this has been challenged. This report concerns further experiments which can explain the apparently conflicting observations of various laboratories and which provide additional evidence of the involvement of coagulation factors and thrombin in the platelet release reaction caused by ADP and other agents.Washed platelets suspended in a buffer solution responded poorly to ADP with no second phase aggregation or release of [3H]5HT. In contrast, washed platelets suspended in dialysed plasma underwent second phase aggregation and released radioactivity. This response depended on calcium. Dialysed plasma deficient in factors XI or X did not restore second phase aggregation or the release reaction. Hirudin and heparin inhibited second phase aggregation and release by ADP and epinephrine. However, the inhibitory effects of heparin and hirudin on ADP,epinephrine and collagen were not observed when citrate was present. To explore the possibility that a reduction of the free calcium concentration accounts for the inhibition of action of these antithrombin agents by citrate experiments with EGTA were carried out. This alternative chelating agent also prevented the inhibitory actions of heparin and hirudin.These observations support the view that platelets and clotting function cooperatively in platelet reactions involved in haemostasis. We suggest that small amounts of thrombin formed prior to fibrin clotting mediate platelet reactions in haemostasis initiated by collagen and ADP and that secondary aggregation does not represent a citrate artifact.