Direct Radioimmunoassay of Nuclear 3,5,3′ Triiodothyronine in Rat Anterior Pituitary

P. R. Larsen; S. Z. Bavli; M. Castonguay; R. Jove

doi:10.1172/JCI109713

Direct Radioimmunoassay of Nuclear 3,5,3′ Triiodothyronine in Rat Anterior Pituitary

P. R. Larsen, S. Z. Bavli, M. Castonguay, and R. Jove

Published March 1, 1980 - More info

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Abstract

Previous tracer studies have suggested that 5′-monodeiodination of l-thyroxine (T₄) in anterior pituitary may contribute a substantial portion of specifically bound nuclear 3,5,3′ l-triiodothyronine (T₃) in this tissue in rats. To evaluate this possibility, a radioimmunoassay for nuclear T₃ in individual anterior pituitaries was developed. Animals received [¹²⁵I]T₃ 60 min before removal of the anterior pituitary and isolation of the nuclei by differential centrifugation. This allowed calculation of the nuclear:serum T₃ ratio and comparison of expected with measured T₃. T₃ was extracted in ethanol, dried, and reconstituted in assay buffer. In untreated hypothyroid rats, anterior pituitary nuclear T₃ was 0.18 ± 0.06 pg/μg DNA which was 0.13 pg/μg DNA greater than expected from the serum T₃ concentration and the pituitary nuclear:serum [¹²⁵I]T₃ ratio. In 10 hypothyroid rats given a single bolus of 400 ng T₃/100 g body wt., the nuclear T₃ by radioimmunoassay was 1.0 ± 0.06 pg/μg DNA, whereas that expected from the T₃ specific activity calculations was 0.85 pg/μg DNA (P < 0.025). Serum T₄ concentrations in these rats were < 0.25 μg/dl but the nuclear T₃ derived from as little as 0.2 μg/dl T₄ could explain a large portion of these small discrepancies between observed and measured nuclear T₃. In 29 normal rats, anterior pituitary nuclear T₃ was 0.63±0.04 pg/μg DNA, whereas that expected from the serum T₃ concentration (55±2 ng/dl) was 0.23±0.02 pg/μg DNA (P < 0.001). Total pituitary T₃ based on this measurement was 92±6 pg. Because the maximal nuclear binding capacity for T₃ in rat anterior pituitary is 0.77 pg/μg DNA, these results suggest there is 82% occupancy of these nuclear receptors. The requirement for normal serum concentrations of both T₄ and T₃ to achieve normal nuclear T₃ saturation in anterior pituitary is in marked contrast to the situation in liver, kidney, and heart muscle which appear to require only a normal serum T₃. As a consequence, the anterior pituitary can monitor both serum T₄ and T₃ and respond appropriately to changes in their concentrations.