Identification of Thyroid Hormone Conjugates Produced by Isolated Hepatocytes and Excreted in Bile of Rainbow Trout,Oncorhynchus mykiss

Abstract

We studied thyroid hormone (TH) conjugation in fasted trout by incubating isolated hepatocytes with either [125I]T4or [125I]T3, and by analyzing bile from trout injected with either [125I]T4or [125I]T3. Glucuronide conjugates were identified by hydrolysis with β-glucuronidase and sulfate conjugates by acid solvolysis with ethyl acetate/trifluoroacetic acid (1%). We used Sephadex LH-20 chromatography to concentrate the conjugate fractions from hepatocyte incubates prior to HPLC analysis. Glucuronide conjugates of T4and T3were producedin vitroand glucuronides of T4, T3, and 3,3′-T2were foundin vivo.Sulfation of T4occurredin vitroandin vivo.T3sulfation was not establishedin vitro,but sulfate conjugates of T3and T2were found in bile. Significant proportions of unconjugated T4and T3also occurred in the bile. We conclude that (i) as in other vertebrates, iodothyronines undergo hepatic glucuronidation and are excreted as such in the bile, (ii) T4and T3undergo sulfation, and in contrast to mammals, are excreted in significant amounts in the bile, (iii) 3,3′-T2, a prominent deiodination product of T3, is excreted as both glucuronide and sulfate conjugates, and (iv) the isolated hepatocyte system is appropriate for studying aspects of TH metabolism in trout.