Trypsin-induced acute inflammation was studied in hamster cheek pouch using intravital microscopy, correlative histology, and electron microscopy. Vascular permeability changes were monitored with intravital fluoroscopy, after intravenous injection of FITC-dextran (Mw 150,000), by counting the number of FITC-dextran leakages around the vessels. The number of extravasated polymorphonuclear leukocytes (PMNLs) was calculated by a histological technique. A dose-dependent increase in the number of FITC-dextran leakages, as well as the number of accumulated PMNLs, was found when trypsin was locally deposited in concentrations of 0.25-2.5 microM (15 microliters during 5 min). Local deposition of autologous serum treated with trypsin at final concentrations of 0.25-2.5 microM caused an increase in vascular permeability as equally pronounced as that of pure trypsin, but only a moderate PMNL accumulation which was not dose dependent. Trypsin at a 25 microM concentration resulted in numerous microbleedings and cessation of flow. The electron microscopy demonstrated inflammatory events (PMNL adhesion, diapedesis, and interstitial infiltration) in all treatment groups but they were more pronounced after trypsin exposure. Trypsin did not cause disintegration, cellular lysis, or increased mast cell degranulation. The permeability changes induced by trypsin (2.5 microM) and trypsinated serum (2.5 microM) were significantly suppressed by the addition of the chelating agent potassium-EDTA to the reaction mixture, indicating a calcium- or magnesium-dependent mechanism. Pretreatment of the animals with cobra venom factor (CVF), by which the plasma C3 concentration was reduced to less than 10%, inhibited the vascular leakages almost completely. The trypsin-induced accumulation of PMNLs was significantly reduced by potassium-EDTA as well as by pretreatment with CVF (P less than 0.01). These findings indicate a central role of complement activation in trypsin-induced acute inflammation in the hamster cheek pouch.