Different Requirements for Intrinsic Factor‐Xa Forming Activity and Platelet Factor 3 Activity and their Relationship to Platelet Aggregation and Secretion

S ummary . Platelets provide coagulant activity in part by promoting two essential reactions of intrinsic coagulation: (I) factor‐X activation by a complex of factor IXa, factor VIII, calcium and platelets (intrinsic factor‐Xa forming activity or XaFA); and (2) prothrombin activation by a complex of factor Xa, factor V, calcium and platelets (platelet factor 3 activity or PF 3 A). We found that a chloroform extract of acetone‐dried brain had about half the XaFA of a petroleum ether extract of brain and over twice the PF 3 A of the petroleum ether extract; and that when purified phospholipids other than phosphatidylserine (PS) were added to PS, the resultant activity, compared with PS alone, was reduced in the assay for XaFA and enhanced in the assay for PF 3 A. In experiments with washed platelets stirred with collagen XaFA developed rapidly (100% activity at 30 s), well before maximum [ 14 C]5HT release (2‐4 min) and decayed to 15% at 10 min. In contrast, PF 3 A developed slowly (100% activity after 20 min) well after maximal aggregation and release. Maximal XaFA developed in unstirred platelet suspensions incubated with collagen, whereas PF 3 A did not become available unless platelet suspensions were stirred and underwent secretion. Platelets stirred with ADP released [ 14 C]5HT and developed PF 3 A but not XaFA. Both [ 14 C]5HT release and PF 3 A by collagen were inhibited by p ‐chloromercuriphenylsulphonate, indomethacin and a combination of prostaglandin E 1 and RA 233, but XaFA was only minimally affected by these inhibitors. In contrast concanavalin A inhibited collagen‐induced PF 3 A and XaFA in dose‐dependent fashion but had no effect on [ 14 C]5HT release. Both platelet coagulant activities and [ 14 C]5HT release were inhibited by aspirin, EDTA and a combination of antimycin A and 2‐deoxy‐D‐glucose. These results indicate that the biochemical determinants of XaFA and PF 3 A are different. PF 3 A developed only when aggregation and release occurred whereas XaFA was independent of aggregation and release.