Microcystin-LR toxicodynamics, induced pathology, and immunohistochemical localization in livers of blue-green algae exposed rainbow trout (Oncorhynchus mykiss)

With this retrospective study, we investigated the temporal pattern of toxi n exposure and pathology, as well as the topical relationship between hepat otoxic injury and localization of microcystin-LR, a potent hepatotoxin, tum or promoter, and inhibitor of protein phosphatases-1 and -2A (PP), in liver s of MC-gavaged rainbow trout (Oncorhynchus mykiss) yearlings, using an imm unohistochemical detection method and MC-specific antibodies. H&E stains of liver sections were used to determine pathological changes. Nuclear morpho logy of hepatocytes and ISEL analysis were employed as endpoints to detect the advent of apoptotic cell death in hepatocytes. Trout had been gavaged w ith lyophilized cyanobacteria (Microcystis aeruginosa, strain PCC 7806) at acutely toxic doses of 5700 mu g microcystin (MC) per kg of body weight (bw ), as described previously (Tencalla and Dietrich, 1997). Briefly, 3 contro l and 3 test animal were killed 1, 3, 12, 24, 48, and 72 h after bolus dosi ng, and livers were fixed and paraffin embedded for histological analysis a nd later retrospective histochemical analyses. The results of the immunohis tochemistry reported here revealed a time dependent, discernible increase i n MC-positive staining intensity throughout the liver, clearly not concurri ng with the kinetics of hepatic PP inhibition observed in the same fish and reported in an earlier publication by Tencalla and Dietrich (1997). After 3 h, marked and increasing MC-immunopositivity was observed in the cytoplas m, as well as the nuclei of hepatocytes. Apoptotic cell death could be dete cted after 48 h, at the very earliest. These data suggest that accumulation of MC and subsequent changes in cellular morphology, PP inhibition, and he patocyte necrosis represent the primary events in microcystin induced hepat otoxicity and appear to be associated with the reversible interaction of MC with the PP. In contrast, apoptotic cell death, as demonstrated here, seem s to be of only secondary nature and presumably results from the covalent i nteraction of MC with cellular and nuclear PP as well as other thiol contai ning cellular proteins.