Thromboxane A2/prostaglandin H2 mobilizes calcium in human blood platelets

The present study investigated the mechanism by which thromboxane A2/prostaglandin H2 (TXA2/PGH2) stimulates platelet activation. Previous studies in isolated platelet vesicles have suggested that TXA2/PGH2 functions to release calcium from intraplatelet stores. On this basis, we investigated whether TXA2/PGH2 causes mobilization of calcium in intact platelets. Calcium redistribution was measured using the fluorescent probe, chlortetracycline (CTC), and a photon-counting microspectrofluorometer. Human platelet-rich plasma was incubated with CTC (50 microM) for 40 min at 25 degrees C. Shape change was induced with arachidonic acid (AA, 100 microM) or ADP (0.75-1.0 microM). It was found that AA addition resulted in a significant release of intraplatelet calcium. This release was blocked by inhibition of the cyclooxygenase with indomethacin (20 microM) or the specific TXA2/PGH2 antagonist, 13-azaprostanoic acid (13-APA, 100 microM). On the other hand, neither indomethacin nor 13-APA had any effect on calcium release stimulated by ADP. However, prostacyclin (13 nM) inhibited both AA- and ADP-induced calcium release. These findings provide evidence that cyclooxygenase products of AA, i.e., TXA2 and/or PGH2 directly caused the mobilization of intraplatelet calcium. Furthermore, this calcium mobilization appears to be mediated through a specific TXA2/PGH2 receptor interaction.