Identification of thyroid hormone conjugates produced by isolated hepatocytes and excreted in bile of rainbow trout, Oncorhynchus mykiss

We studied thyroid hormone (TH) conjugation in fasted trout by incubating isolated hepatocytes with either [I-125]T-4 or [I-125]T-3, and by analyzing bile from trout injected with either [I-125]T-4 or [I-125]T-3. Glucuronide conjugates were identified by hydrolysis with beta-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 T-4 and T-3 were produced in vitro and glucuronides of T-4, T-3, and 3,3'-T-2 were found in vivo. Sulfation of T-4 occurred in vitro and in vivo. T-3 sulfation was not established in vitro, but sulfate conjugates of T-3 and T-2 were found in bile. Significant proportions of unconjugated T-4 and T-3 also 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) T-4 and T-3 undergo sulfation, and in contrast to mammals, are excreted in significant amounts in the bile, (iii) 3,3'-T-2, a prominent deiodination product of T-3, is excreted as both glucuronide and sulfate conjugates, and (iv) the isolated hepatocyte system is appropriate for studying aspects of TH metabolism in trout. (C) 1996 Academic Press, Inc.