A model incubation system containing purified thyroid peroxidase (TPO) was used to study the mechanism of action of the thioureylene anti-thyroid drugs--propylthiouracil (PTU), methylmercapto imidazole (MMI) and carbimazole. Two general types of experiments were performed: a) measurement of the inhibitory effects of the drugs on TPO-catalyzed iodination and on TPO-catalyzed oxidation of guaiacol, and b) studies of the metabolism of PTU and MMI by the TPO model system. The major observations can be summarized as follows: 1) The thioureylene drugs are potent inhibitors of TPO-catalyzed iodination of protein and tyrosine. Their potency increases greatly as the concentration of I- decreases. 2) The thioureylene drugs are also potent inhibitors of TPO-catalyzed oxidation of guaiacol, a reaction that does not involve iodide. 3) MMI and PTU are readily oxidized in the model incubation system when iodide is present but not in the absence of iodide. The rate of oxidation increased as the iodide concentration was increased from 10 to 100 muM. 4) Oxidation of PTU and MMI by the model incubation system is inhibited by relatively slight increases in the concentration of PTU and MMI. These drugs are capable of inhibiting their own and each other's metabolism. 5) Inhibition of iodination is competitively antagonized by iodide at low drug concentrations, but not at higher drug concentrations. 6) Inhibition of iodination by MMI and PTU may be either reversible (low ratio of drug to iodide), or irreversible (higher ratio of drug to iodide). In reversible inhibition the iodination is inhibited for a period which may be as brief as 2 min or as long as 20 min, but thereafter, iodination begins, and there is escape from inhibition. During the lag-period there is extensive metabolism of the drug. In the case of irreversible inhibition of iodination is inhibited completely or almost completely for 60 min, and drug oxidation during this period is relatively low. 7) Irreversible inhibition may be transformed into reversible inhibition by increasing the concentration of TPO or the concentration of iodide. However, increasing the concentration of H2O2 or of tyrosine does not overcome irreversible inhibition. On the basis of these findings and of current views concerning the mechanism of enzymatic iodination, a scheme is proposed for the mechanism of inhibition by thioureylene drugs of TPO-catalyzed iodination of protein and tyrosine.

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We have developed sensitive and reliable radioimmunoassays for T4 and T3 in amphibian plasma and have used these procedures to measure plasma T4 and T3 levels in spontaneously developing Rana catesbeiana tadpoles at various stages of metamorphosis. During premetamorphosis circulating levels of both T4 and T3 were below the limits of detection of the RIA procedures (T4 less than 50 ng/100 ml, T3 less than 5 ng/100 ml). A gradual rise in plasma T3 and T4 became apparent during prometamorphosis, and at the onset of metamorphic climax the levels of both T4 and T3 increased sharply. Peak levels for both hormones were observed in the middle of metamorphic climax (stage XXIII). The circulating T3 level reached a mean peak of 78 ng/100 mg, at least 15 times greater than the level during premetamorphosis. The peak T4 level was 0.5 microgram/100 ml, about a 10-fold increase over the premetamorphosis level. The surge in thyroid hormone secretion lasted only for several days, and during the latter half of metamorphic climax there was a fairly rapid decrease in plasma T4 and T3. By 2 days post-climax the levels had declined to about 20% of their peak values. Free T4 and T3 levels in plasma followed the same general pattern as the total hormone levels during the various stages of tadpole development. In adult R. catesbeiana, plasma T4 and T3 levels were surprisingly low. Similarly low values were observed in Bufo marinus and in Rana pipiens. The very low levels of circulating T4 and T3 both in premetamorphosis tadpoles and in adults suggest that thyroid hormones in anuran Amphibia may be of importance only during the period of metamorphosis.

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We measured thyroxine 5'-deiodinase I (T(4)5'D-I) activity in thyroid, liver, and kidney and thyroxine 5'-deiodinase II (T(4)5'D-II) activity in brown adipose tissue (BAT) in rats on a low-iodine diet (LID) to test the possibility that increased deiodinase activity in these tissues might contribute to the maintenance of ther serum 3,5,3'-triiodothyronine (T3) level. Control rats received LID plus KI. Experiments were also performed with LID and LID plus KI rats exposed to cold. T(4)5'D-I activity was greatly increased in the thyroids of LID rats but not in liver or kidney. We consider it likely that increased thyroxine (T4)-to-T3 conversion in the greatly enlarged thyroids of LID rats contributed to the maintenance of serum T3. T(4)5'D-II activity in BAT was markedly increased in LID rats and was further greatly increased on cold exposure. However, we were unable to demonstrate an increase in uncoupling protein mRNA levels in BAT in response to cold in LID rats. We attribute this to the very low serum T4 level, which limits substrate availability. This factor also makes it unlikely that BAT contributes to maintenance of serum T3 in LID rats.