To study the role of Kupffer cells in the aggravation of liver injury, effects of zymosan on acute liver damage were explored using perfused livers of rats 24 h after intraperitoneal injection of D-galactosamine (800 mg/kg). The leakage of lactate dehydrogenase and aspartate aminotransferase into the effluent was used to indicate acute liver damage. Infusion of zymosan (30 µg/ml) into the portal vein rapidly increased the leakage of lactate dehydrogenase and aspartate aminotransferase from galactosamine-treated liver with decreased perfusion flow. Pretreatment of animals with gadolinium, which diminished an immunostaining of resident macrophages in the injured liver, significantly attenuated the flow reduction induced by zymosan, whereas it did not affect the increases in enzyme leakage. Infusions of PGF₂, PGE₂, and leukotriene D₄, the eicosanoids mainly produced by Kupffer cells, decreased perfusion flow without rapid augmentation of enzyme leakage from galactosamine-treated liver. These results indicate that zymosan potentiates acute liver damage after galactosamine injection and suggest that certain types of nonparenchymal cells other than Kupffer cells are mainly involved in the action of zymosan.