Thrombopoietin (TPO) regulates stem cell proliferation and maturation of megakaryocytes by activating the c-Mpl-receptor, a member of the hematopoietic cytokine family. As human platelets possess c-Mplreceptors and supraphysiological concentrations of TPO trigger platelet aggregation and secretion, we searched for the signalling pathways through which the c-Mpl-receptor might activate platelets. A physiological concentration of TPO (20 ng/mL) did not trigger platelet functions, but increased their sensitivity to α-thrombin resulting in a 4-fold faster dense granule secretion. The effect of TPO was abolished by indomethacin and caused by synergism with signal generation by α-thrombin at the level of the cytosolic phospholipase A₂ (cPLA₂) pathway resulting in more arachidonate release, cPLA₂ phosphorylation and thromboxane A₂ formation. A similar synergism was seen at the level of extracellular signal-regulated kinase 2 (ERK2 or p42-MAPK). These data suggest, that TPO increases the sensitivity of platelets to α-thrombin by enhancing cPLA₂ activation via the ERK2-cPLA₂ pathway.