In VivoSteroid Metabolism in Embryonic and Newly Hatched Steelhead Trout (Oncorhynchus mykiss)

Abstract

Radioactive pregnenolone (P5), testosterone (T), or 17β-estradiol (E2) was microinjected into steelhead trout,Oncorhynchus mykiss,embryos and newly hatched yolksac fry (alevins) to detectin vivometabolism. We also assayed the water used to incubate animals for 10 hr after microinjection to detect possible metabolite excretion. High pressure liquid chromatography and thin layer chromatography were used to separate and tentatively identify steroid metabolites. Metabolites of P5 were androstenedione (AN), E2, T, and glucuronides of E2 and T in embryos and AN, E2, progesterone, 17α, 20β-dihydroxyprogesterone, and P5 glucuronide in alevins. E2 and its glucuronide were synthesized from precursor T in the embryos and alevins; however, the amounts of E2 and E2 glucuronide synthesized in the embryos were 10 and 3 magnitudes greater than those detected in alevins. Testosterone glucuronide was synthesized in similar amounts in both stages of development. Embryos did not synthesize free metabolites from E2 precursor, but E2 glucuronide was detected from E2 precursor. Estradiol in alevins was metabolized into unidentified free and glucuronide-conjugated steroids. Three unknown metabolites synthesized from P5 precursors and seven unknown substances produced in animals injected with testosterone or estradiol precursors were detected. Free metabolites were detected in the incubation water that held the animals (embryos and alevins) for 10 hr after microinjection with T or E2. Glucuronide metabolites were not excreted by embryos into the incubation water 10 hr after microinjections with any of the steroid precursors; however, alevins excreted glucuronides into the incubation water when supplied with precursor T. These results imply that endogenous steroid metabolism of maternally contributed steroids is active during early development.