The regulatory mechanism of cytosolic sulfation of T3 has been studied in rat liver. Sulfation of T3 is sexually differentiated in adult rats of Sprague-Dawley (SD), Fisher 344, and ACI strains. In SD strain, the male animals showed 4 times higher sulfating activity than did the females. The specific activity was decreased by hypophysectomy of male adult rats, but was not affected in the females. Thus, the sex-difference was abolished in the hypophysectomized condition. Supplement of human GH intermittently twice daily for 7 days, to mimic the male secretory pattern, increased T3 sulfating activity in both sexes of hypophysectomized rats, whereas continuous infusion to mimic a female secretory pattern had no appreciable effect. Cytosolic sulfation of T3 was decreased by 25 to 30% by thyroidectomy or propylthiouracil treatment of male adult rats, and was restored by the supplementation of T3 (50 micrograms/kg daily for 7 days) to thyroidectomized rats. Administration of T3 in hypophysectomized rats almost completely restored the sulfating activity in the males and increased the activity in the females. Cytosolic T3 sulfation was inhibited by the addition of known inhibitors of phenol sulfotransferase, pentachlorophenol or 2,6-dichloro-4-nitrophenol. These results indicate a role of pituitary GH in hepatic sulfation of thyroid hormones in rats. The data obtained also raise the possibility that GH may modify the effect of thyroid hormones on the pituitary by a feed-back mechanism through changing the level of a sex-dominant phenol sulfotransferase(s) in rat livers. T3 was also sulfated in hepatic cytosols of mouse, hamster, rabbit, dog, monkey, and human.(ABSTRACT TRUNCATED AT 250 WORDS)