Regulation of arachidonic acid-dependent Ca++ influx in human platelets

Quin2 was used to study the rise in cytoplasmic free calcium ([Ca++]i) and the role of prostaglandin (PG) endoperoxides/thromboxane A2 (TxA2), reduced glutathione (GSH), ADP and the glycoprotein (GP) IIb-IIIa complex in mediating [Ca++]i rise during arachidonic acid (AA)-induced platelet aggregation. Ca++ mobilization, mostly due to an influx across the plasma membrane, is completely inhibited by aspirin and persists after selective blockade of TxA2 synthase by dazoxiben. GSH total depletion causes a complete aggregation block and 90% inhibition of the transient: U-46619, a stable analog of cyclic endoperoxide PGH2, stimulates [Ca++]i transient in aspirin-treated or in GSH-depleted platelets. ADP-scavengers, ATP (which competes for the ADP receptor), and monoclonal antibodies against the GP IIb-IIIa complex reduce AA-induced Ca++ influx. Therefore, PG endoperoxides alone or a PGH2/TxA2 mimetic stimulate Ca++ influx. Synthesis of PGH2 and TxA2 depends on the availability of GSH, which acts as the reducing cofactor for the PG-peroxidase activity. ADP and GP IIb-IIIa are regulating factors of AA-mediated Ca++ influx during platelet activation.