Exposure of killifish to benzo[a]pyrene: comparative metabolism, DNA adduct formation and aryl hydrocarbon (Ah) receptor agonist activities

Abstract

Benzo[ a]pyrene (BaP) elicited a dose-response induction of hepatic CYP1A1 gene expression in killifish characterized by increased CYP1A1 mRNA levels, ethoxyresorufin O-deethylase (EROD) and aryl hydrocarbon hydroxylase (AHH) activities. There were marked differences in the maximally-induced EROD (1529 pmol mg −1min −1) and AHH (377 pmol mg −1 min −1) activities and these differences were observed at all üme points and at concentrations of BaP from 1 to 50 mg/kg. Treatment of killifish with BaP did not significantly affect binding of [ 125I]epidermal growth factor (EGF) to hepatic plasma membranes containing the EGF receptor. There was a dose-dependent increase in formation of DNA adducts in killifish treated with 1-50 mg/kg of BaP. Two-dimensional thin-layer chromatographic analysis of the [ 32P] labeled adducted nucleoüdes revealed one major spot. In a time course study over a period of 2–14 days, the relative DNA adduct levels in fish treated with 5 mg/kg BaP were constant. The most sensitive indicator of exposure to BaP was the dose-dependent formation of biliary metabolites in which there was a > 40-fold increase (compared to untreated animals) in fluorescent metabolites formed at the lowest dose of BaP (1 mg/kg) used in this study. Killifish expressed relatively high levels (203 fmol/mg) of the nuclear aryl hydrocarbon (Ah) receptor which was isolated from nuclear extracts of fish treated with [ 3H]-2,3,7,8-tetrachlorodibenzo p-dioxin. The nuclear Ah receptor complex sedimented at 6.0 S which was similar to that observed in other animal species. Based on their CYPIA1 induction response to BaP and Ah receptor levels, the results of this study indicate that killifish are highly Ah-responsive.