Species-specific antagonism of Ah receptor action by 2,2',5,5'-tetrachloro- and 2,2',3,3'4,4'-hexachlorobiphenyl.

Using recombinant cell lines showing Ah receptor-controlled expression of a luciferase reporter gene, the interaction of di-ortho-substitute polychlorinated biphenyls (PCBs) with Ah receptor agonists was studied. In the recombinant Hepa1c1c7 mouse hepatoma (H1L1.1c7) cells strong antagonistic interaction of 2,2',5,5'-tetrachlorobiphenyl (PCB52) with luciferase expression induced by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) or 3,3',4,4'-tetrachlorobiphenyl (PCB77) was observed, and similarly, between 2,2',3,3',4,4'-hexachlorobiphenyl (PCB128) and PCB77. Accordingly, PCB52 was found to inhibit ethoxyresorufin-O-deethylase (EROD) induction by PCB77 in wild-type Hepa1c1c7 cells. In contrast, the antagonistic effect of PCB52 on TCDD-induced luciferase expression was only minor in recombinant guinea pig GPC16 colon adenocarcinoma (G16L1.1c8) and human HepG2 hepatoma (HG2L1.1c3) cells, and intermediate in recombinant H4IIE rat hepatoma (H4L1.1c4) cells. Gel retardation studies using a 32 P-labelled dioxin responsive element (DRE)-containing oligonucleotide, and ligand binding studies using [3H]TCDD, demonstrated that the species-specific antagonistic activity of PCB52 on Ah receptor-controlled luciferase expression is due to inhibition of Ah receptor ligand and DNA binding. We conclude, that Ah-mediated luciferase expression provides a useful tool to study the species specificity of Ah receptor (ant)agonists.

Stress factors that can affect studies of drug metabolism in fish.

The effects of environmental stress conditions on the defense response of rainbow trout following a four week exposure to subacute levels of un-ionized ammonia or temperatures 5 C above and 5 C below the temperature optimum (15 C) were investigated. These experimental studies can serve as a model to evaluate the metabolic response of fish to external agents (e.g., drugs, vaccines) under environmental conditions seen in the culture of fish. Blood and tissue immune parameters measured include hematocrits, antibody levels and differentiation of white blood cell populations in tissue imprints of the anterior kidney. These analyses were compared to the growth parameter, average percent weight gain. Fish given primary and secondary immunization with a bacterial vaccine (Aeromonas hydrophila) were exposed to sublethal concentrations of un-ionized ammonia of 0.2, 0.3 or 0.4 mg/ml. Fish exposed to the higher concentrations of ammonia showed a decrease in growth compared to control fish. Several significant changes were observed in the leukocytes of the anterior kidney at the various concentrations of ammonia tested. A decrease in antibody titers to A hydrophila was seen at the two higher concentrations of ammonia. In a second study, the effects of non-optimum temperature conditions (10 C and 20 C) were compared with an optimum temperature (15 C). Fish held at sub-optimum temperatures had significantly lower hematocrits than the control fish maintained at 15 C. Several significant changes were also seen in the anterior kidney leukocytes. Antibody titers to A hydrophila were significantly lower at the end of the stressing period in the trout maintained at 10 C compared to the immunized controls at 15 C. In contrast, fish held at 20 C had significantly higher antibody titers than did the immunized controls. Compared to controls, fish growth was increased at 10 C and decreased at 20 C. These studies confirm that environmental factors can induce stress and affect the metabolism and health of the fish.