Thyroid hormone metabolism in primary cultures of fetal rat brain cells

Abstract

The metabolism of thyroxine 3,5,3′,5′-tetraiodothyronine, (T 4) and 3,5,3′-triiodothyronine (T 3) was studied in primary cultures of dispersed fetal rat brain cells. Cultured brain cells actively metabolized both T 4 and T 3 by enzyme catalyzed deiodination reactions which increase (type II 5′-deiodinase) or decrease (type I 5′-deiodinase and type III 5-deiodinase) the bioactivity of thyroid hormone. Homogenates of cultured brain cells showed both type I and type II 5′-deiodinating activities and these two enzymes tended to differ in their time course of appearance. Cultures exposed to 10 μM cytosine arabinoside for 16 h showed up to a 70% reduction in type I activity without decreasing the type II enzyme suggesting that the type II enzyme is associated with non-dividing neuronal cells. The pre-dominant pathway for T 4 and T 3 metabolism in situ was tyrosyl-ring or type III 5′-deiodination which followed first order kinetics with a t 1/2 of 70 min. T 4 to T 3 conversion by the type II enzyme was consistently observed after correcting for the degradation of newly formed T 3 by the type III enzyme. In situ, both type II and type III enzymes were thiol-dependent and both activities were inhibited by iopanoic acid. Type III 5-deiodination of T 4 produced 34 fmol 3,3,5′-triiodothyronine (rT 3)/h per 10 6 cells at 10 mM dithiothreitol (DTT) and 97 fmol of rT 3/h per 10 6 cells at 50 mM DTT. T 3 production by the II enzyme was 1.2 and 4.4 fmol of T 3/h per 10 6 cells at 10 and 50 mM DTT, respectively. Thyroid hormone deficient culture conditions increased type II enzyme activity by 4–5-fold within 48 h and this was prevented in a dose-dependent fashion by supplementing the media with increasing amounts of T 3. These data indicate that primary cultures of dispersed brain cells mimic the intact cerebral cortex with respect to the metabolism of thyroid hormone and the regulatory mechanisms which defend cerebrocortical T 3 levels. The vigorous metabolism of both T 4 and T 3 by these cultures may explain some of the difficulties in demonstrating thyroid hormone-dependent biochemical changes at physiologically relevant levels of thyroid hormone.