Kinetic evidence suggesting two mechanisms for iodothyronine 5'-deiodination in rat cerebral cortex.

Abstract

Enzymatic 5'-deiodination of 3,3',5'-triiodothyronine (rT3) and 3,3',5,5'-tetraiodothyronine (thyroxine, T4) was studied in microsomal preparations of rat cerebral cortex. Evidence was obtained for the existence of two thiol-dependent 5'-deiodinase entities. One of these predominates in tissue from euthyroid and long-term hypothyroid rats, is specific for rT3, follows "ping-pong" kinetics with dithiothreitol as the cosubstrate, and is inhibited by propylthiouracil (PrSUra) and iodoacetate. Inhibition by PrSUra is uncompetitive with rT3 and competitive with dithiothreitol. These properties are shared with the 5'-deiodinase activity of liver and kidney. The activity of a second type of 5'-deiodinase is highest in cerebral cortex from short-term hypothyroid rats, prefers T4 to rT3 as the substrate, is insensitive to PrSUra and iodoacetate, and follows "sequential" reaction kinetics. A similar PrSUra-insensitive 5'-deiodinase activity is also found in pituitary but is not detectable in liver and kidney; it seems, therefore, characteristic oftissues in which local T4 to 3,3',5-triiodothyronine (T3) conversion supplies a major portion of the total intracellular T3.The main secretory product of the thyroid gland, 3,3',5,5'-tetraiodothyronine (thyroxine, T4) is converted to the biologically more potent 3,3',5-triiodothyronine (T3) in many tissues of the body by a process termed 5'-deiodination. There appears to be a differentiation between tissues such as the liver and the kidneys, where most of the T3 produced is returned to the circulation, and tissues such as the brain and the pituitary, where the T4 to T3 conversion seems to serve local purposes (1). Another distinction between the deiodinase activities at these loci is the effect ofchanges in thyroid status. Thus, hypothyroidism is associated with decreased T4 to T3 conversion in rat liver and kidney, whereas increased conversion rates are observed in the pituitary and the brain (2)(3)(4)(5). Not only are the changes opposite but also they occur more rapidly in the central organs (6,*).3,3',5'-Triiodothyronine (reverse T3, rT3), an inactive intermediary product in the metabolism of T4, is in fact a better substrate for the enzymes that convert T4 to T3 in the liver and the kidney (7)(8)(9). The enzymes are located in membrane-derived fractions ofthese tissues, depend on thiols for activity, and are effectively inhibited by derivatives of 2-thiouracil-e.g., 6-propyl-2-thiouracil (PrSUra) (7-15).A third difference with T4 5'-deiodinase activities ofthe liver and the kidney is that conversion of T4 to T3 in the pituitary and the brain is not affected by PrSUra in vivo or in vitro (3,4,16,17). However, brain rT3 5'-deiodination is sensitive to PrSUra (18, t pooled tissue from 10 rats (approximately 5 g wet weight) was homogenized in 5-6 vol (vol/wt) of ice-cold 0.32 M sucrose, 10 mM Hepes (pH 7.0), containing 10 mM dithiothreitol (DTT), using a hand-driven, all-glass homogenizer. The homogenate was centrifuged at 4°C for 15 min at 15,000 X g. The pellet was wa...

Induction of type II iodothyronine 5'‐deiodmase and mitochondrial uncoupling protein in brown adipocytes differentiated in cell culture

“…The Km for T4 of the 5'D induced in cultured adipocytes by norepinephrine and dbcAMP was around 10-9 M, similar to microsomes of cerebral cortex [3] or infranatants of BAT [3] from hypothyroid adult rats. The 5'D of cultured cells was also insensitive to 1 mM propylthiouracil (lower than 10°70 inhibition at 0.2 nM and 2 nM T4 as substrate).…”

“…5'D activity was assayed according to [3] with the following modifications: the incubation mixture (200 tzl) contained 100 mM sodium phosphate (pH 7.0), i mM EDTA, 10 mM DTT, 0.15 nM [t231]T4 (-50000 cpm of L-[3',5'-I25I]T4, > 44TBq/g; from IZIN-TA, Hungary), 2.25 nM unlabeled "1"4 and 1 mM 6-npropyl-2-thiouracil. After 1 h at 37°C, 50 #1 of 2°70 BSA was added followed by 350 #1 of ice-cold 10070 TCA.…”

“…After 1 h at 37°C, 50 #1 of 2°70 BSA was added followed by 350 #1 of ice-cold 10070 TCA. Released 123I-was separated from iodothyronines on Dowex 50W-X2 columns and counted as previously described [3]. The tracer was purified by paper electrophoresis before each assay and blank values (no protein) were less than 1 °70 of the radioactivity added.…”

“…In contrast to type I enzyme present in liver and kidney, type II 5'D is insensitive to propylthiouracil, exhibits different kinetics and has low gm values for T4 and reverse T3 in the nanomolar range [3,4].…”

The effects of thyroid state on beta-adrenergic and serotonergic receptors in rat brain

“…Furthermore, it has been demonstrated that the control and kinetics of 11 Address correspondence to: George A. Mason,203 BSRC (220H), UNC Department of Psychiatry, Chapel Hill, NC 27514, U.S.A. 261 the 5'-monodeiodinase which converts T 4 to T 3 , found in almost every peripheral organ (type I), is different from the enzyme (type II) present in brain, brown fat and pituitary (Visser et al, 1982). These enzymatic differences may be responsible for the marked differences in changes which occur in central and peripheral levels of T 3 during starvation, for example, in which peripheral T 3 is reduced and brain T 3 relatively unchanged (Larsen et al, 1981).…”