Effect of zooxanthellatoxin-A, an unique marine product, on arachidonic acid cascade in rabbit platelets

Zooxanthellatoxin-A (ZT-A), a bioactive substance isolated from a symbiotic marine alga Symbiodinium sp., caused rabbit platelet aggregation. ZT-A-induced aggregation was dependent on the presence of external Ca2+, and was inhibited by several Ca2+ channel antagonists except L-type one. Furthermore, ZT-A-induced aggregation was attenuated by genistein, indomethacin and SQ29548, indicating that tyrosine phosphorylation and thromboxane A2 (TXA2) are involved in the aggregation. In fact, ZT-A released arachidonic acid and accumulated TXB2, a stable metabolite of TXA2, which was inhibited by genistein. ZT-A caused phosphorylation and activation of mitogenactivated protein kinase (MAPK), which was known to activate cytosolic phospholipase A2 (cPLA2). ZT-A caused the activation of phospholipase C (PLC)-gamma 2, resulting in an accumulation of diacylglycerol that activates protein kinase C (PKC). The MAPK activation was inhibited by genistein and staurousporine. ZT-A is not a Ca(2+)-lonophore, since its different responsibility from ionomycin to external Ca2+, indomethacin and 12-HETE, a platelet lipoxygenase product. These results suggest that ZT-A stimulates PKC a tyrosine kinase with influxed Ca2+, resulting in the activation PLC-gamma 2 that stimulates via diacylglycerol. Then, MAPK is activated by a PKC pathway, then cPLA2 is activated by MAPK. The released arachidonic acid is rapidly converted to TXA2 which causes platelet aggregation.