Dieldrin pretreatment does not induce hepatic microsomal and cytosolic epoxide hydrolase activities in rainbow trout (Oncorhyncus mykiss) Public Deposited

http://ir.library.oregonstate.edu/concern/graduate_thesis_or_dissertations/jw827f60s

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  • Previous studies have shown that rainbow trout exposed to dieldrin via diet for 9 to 12 weeks increased biliary excretion of a subsequent dose of [¹⁴C]dieldrin by 500%. This was not explained by induction of the cytochrome P-450 (CYP) system involved in oxidative metabolism of these compounds. We hypothesized that epoxide hydrolase activity increased in dieldrin fed-fish. Epoxide hydrolase is an enzyme that catalyzes the hydrolysis of epoxide compounds to their corresponding diols. For instance, dieldrin is metabolized to 6,7 trans-aldrindihydrodiol. This study investigated the activity of epoxide hydrolase in microsomes and cytosol of rainbow trout fed a diet that contained 0 or 15 ppm dieldrin. Fish were fed control or dieldrin diet (0.324 ug/g body weight/day) for 3, 6, or 9 weeks. There was a small increase in mortality and decrease in body weight among dieldrin-fed fish after 9 weeks. After week 9, dieldrin-fed fish were fed a control diet for an additional 3 weeks because of these signs of toxicity. At week 12, the difference of body weight between control and treated was not significant. Microsomal and cytosolic epoxide hydrolase activities were measured with a radiometric assay which determined differential partitioning of the parent compound (epoxide) in dodecane and the metabolite (diol) in the aqueous phase. Assays were run at optimal pH and temperature using [³H]trans-stilbene oxide (pH 7) as substrate for cytosol and [³H]cis-stilbene oxide (pH 8) as substrate for microsomes. In order to prevent competition for reaction with stilbene oxide, depletion of glutathione was efficiently achieved by dialysis at 4°C for 2 hours at room temperature in buffer [pH 7.5, potassium phosphate 10 mM, KCL 0.15 M, EDTA 1 mM, BHT 0.1 mM, 0.1 mM PMSF]. Protein quantification was determined by using BCA assay and concentrations were always between 5 and 25 ug/ml in the final assay volume. Epoxide hydrolase activities were not significantly different in cytosol or microsomes from control and dieldrin-fed fish. Dieldrin residues in liver were analyzed by gas chromatography with electron capture detection (GC/ECD). The concentration in the liver increased with time of exposure and declined markedly in fish fed dieldrin for 9 weeks and then fed control diet. No dieldrin was detected in livers from control fish.
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