5-Monodeiodination of T4 and T3 and 5'-monodeiodination of T4 and rT3 were studied in brain homogenates of male Sprague-Dawley rats, aged 1-60 days. Portions of the homogenates were incubated with the substrates at 37 C for 30 min. The reaction products were estimated by specific RIAs. All of the four reactions were dependent upon time, temperature, pH, and upon the concentrations of substrate, thiol, and tissue protein. Maximal reactions were obtained between 40 and 160 mM dithioerythritol. T4 5'-deiodination proceeded optimally at pH 7.4 and 0.4 microM substrate, the other reactions at pH 8.5 and 10 microM substrate. The four reactions were inactivated by heat (56 C, 30 min) and inhibited by 10(-5) M iopanoic acid. Only rT3 5'-deiodination was inhibited by 3 X 10(-4) M propylthiouracil (greater than 95%). In cerebellum, basal ganglia, brainstem, and hypothalamus both T4 and T3 5-deiodinase activity were very high in perinatal rats [up to 5.56 pmol/(min X mg protein) in hypothalamus], and decreased rapidly with age. In cortex and olfactory bulb these enzyme activities were low after birth, followed by an increase during the growth spurt [up to 632 fmol/(min X mg protein) in olfactory bulb]. T4 and rT3 5'-deiodinase activity in all brain regions studied were at their lowest in perinatal rats. During and after the growth spurt an increase was observed [up to 457 fmol/(min X mg protein) in cerebellum]. The reciprocal course of 5- and 5'-deiodination between birth and growth spurt in most of the brain regions studied might lead to a reduced intracellular thyromimetic activity during the perinatal period.