Serine proteinases are involved in several physiological processes and elicit profound cellular effects in a variety of tissues. Besides the thrombin receptor a second receptor, activated by trypsin, the proteinase-activated receptor 2 (PAR-2), was cloned and characterized. Both enzymes generate a new extracellular N-terminus by limited proteolytic cleavage which functions as tethered ligand to activate the receptor. Synthetic peptides corresponding to the sequences of the newly generated N-terminus are able to mimic the effects of the enzymes. In porcine pulmonary arteries trypsin and the receptor-derived peptide SLIGRL elicited an endothelium-dependent transient relaxation of PGF2alpha-precontracted vessels. The EC50 values for trypsin and SLIGRL amounted to 1.1 +/- 0.2 nM and 5.4 +/- 0.6 microM, respectively. Trypsin and SLIGRL caused a homologous desensitization but thrombin and the thrombin receptor-activating peptide SFLLRN were still able to elicit pronounced relaxant effects. The trypsin- and SLIGRL-induced relaxant responses were markedly diminished after blockade of the nitric oxide synthesis by N(G)-nitro-L-arginine methyl ester (200 microM) and were absent in endothelium-denuded vessels. Indomethacin and hirudin did not influence the relaxant effects. The effect of trypsin but not that of SLIGRL was blocked by the proteinase inhibitor aprotinin suggesting that only proteolytically active trypsin activates the receptor. Benzamidine derivatives of the 3-amidinophenylalanine type with different affinity for trypsin and thrombin inhibited the vascular effects of trypsin (IC50 0.007-0.7 microM) correlating with its antitrypsin activity. The data suggest that the vascular effects of trypsin and SLIGRL are mediated through activation of PAR-2 which differs from the thrombin receptor.