THROMBIN-INDUCED CA2-TYROSINE PHOSPHORYLATION IN ENDOTHELIAL-CELLS ISINHIBITED BY HERBIMYCIN-A( INFLUX AND PROTEIN)

During endothelial cell activation, alpha-thrombin elicits both Ca2+ r elease from internal stores and influx of external Ca2+ across the pla sma membrane. The mechanisms of alpha-thrombin-induced Ca2+ entry into endothelial cells are unclear. Therefore, effects of the specific tyr osine kinase inhibitor herbimycin A on protein tyrosine phosphorylatio n and on intracellular Ca2+ transients were studied in alpha-thrombin- stimulated human umbilical vein endothelial cells. alpha-Thrombin caus ed significant tyrosine phosphorylation of mainly two proteins and evo ked typical biphasic changes of free cytosolic Ca2+ concentration. We show that 24 h pretreatment with herbimycin A inhibited alpha-thrombin -induced endothelial protein tyrosine phosphorylation. Moreover, herbi mycin A significantly attenuated alpha-thrombin-induced Ca2+ influx bu t not release from internal stores. The data suggest that protein tyro sine phosphorylation by alpha-thrombin is involved in the regulation o f alpha-thrombin-induced Ca2+ influx into endothelial cells. (C) 1994 Academic Press, Inc.