It is now generally well accepted that heparin and related substances increase the fibrinolytic activity in vivo. The stimulation of the amidolytic, plasminogenolytic and fibrinogenolytic activity of tissue plasminogen activator and urinary plasminogen activator through heparin was investigated in vitro. A concentration-dependent stimulation of the plasminogenolytic and fibrinogenolytic activity of both urinary and tissue-type plasminogen activators was observed in the presence of heparin. No heparin dependence was observed in the amidolytic assay. Heparin stimulates the plasminogenolytic activity of tissue plasminogen activators in the same manner as fibrin. Both activators form complexes with heparin; the heparin-binding-site seems to be identical or related with the fibrin-binding-site of tissue plasminogen activator. The physiological role of these interactions is discussed.