Phenolic and tyrosyl ring deiodination of iodothyronines in rat brain homogenates

Abstract

Conversion of thyroxine (T4) to 3,5,3'-triiodothyronine (T3) in rat brain has recently been shown in in vivo studies. This process contributes a substantial fraction of endogenous nuclear T3 in the rat cerebral cortex and cerebellum. Production of T4 metabolites besides T3 in the brain has also been suggested. To determine the nature of these reactions, we studied metabolism of 0.2 - 1.0 nM [125I]T4 and 0.1 - 0.3 nM [131I]T3 in whole homogenates and subcellular fractions of rat cerebral cortex and cerebellum. Dithiothreitol (DTT) was required for detectable metabolic reactions: 100 mM DTT was routinely used. Ethanol extracts of incubation mixtures were analyzed by paper chromatography in t-amyl alcohol: hexane: ammonia and in 1-butanol:acetic acid. Rates of production of iodothyronines from T4 and T3 were greater at pH 7.5 than at 6.4 or 8.6 and greater at 37°C than at 22° or 4°C. Lowering the pH, reducing the protein or DTT concentrations, and preheating homogenates to 100°C all increased excess I- production but reduced iodothyronine production. In cerebral cortical homogenates from normal rats, products of T4 degradation were as follows (percent added T4 ± SEM in nine experiments): T3, 1.9 ± 0.5%; 3,3',5'-triiodothyronine (rT3), 34.0 ± 2.4%; 3,3'-diiodothyronine (3,3'-T2), 5.8 ± 1.6%; 3'-iodothyronine (3'-T1), ≤ 2.5%; and excess I-, 4.7 ±1.2%. In the same experiments, products of T3 degradation were 3,3'-T2, 63.3±5.5%, and 3'-T1, 12.6±1.4%. Cerebral cortical homogenates from hyperthyroid rats and normals were similar in regard to T4 to T3 deiodination. In contrast, in cerebral cortical homogenates from hypothyroid rats, phenolic ring deiodination rates were increased and tyrosyl ring deiodination rates were decreased compared with normals. T4 to T3 conversion rates in cerebellar homogenates were greater than rates in cerebral cortical homogenates from the same normal rats and less than rates in cerebellar homogenates from hypothyroid rats. T4 and T3 tyrosyl ring deiodination rates were greatly diminished in cerebelar homogenates compared with cerebral cortical homogenates in normal and hypothyroid rats. High-speed (1,000 - 160,000 g) pellets from cerebral cortical homogenates were enriched in phenolic and tyrosyl ring deiodinating activities relative to cytosol. Fractional conversion of T4 to T3 was inhibited by T4, iopanoic acid, and rT3, but not by T3. Tyrosyl ring deiodinatin reactions were inhibited by T3, T4, and iopanoic acid, but not by rT3. These studies demonstrate separate phenolic and tyrosyl ring iodothyronine deiodinase enzymes in rat brain. The brain phenolic ring deiodinase serves in vivo as a T4 5'-deiodinase and closely resembles anterior pituitary T4 5'-deiodinase in physiological and biochemical characteristics. The physiological significance of the tyrosyl ring iodothyronine deiodinase enzyme is unclear; it shares several properties with rat hepatic T4 5-deiodinase.