Increase in 3,5,3'-triiodothyronine (T3)-binding sites in tadpole erythrocyte nuclei during spontaneous and T3-induced metamorphosis.

Abstract

The change in T3-binding sites in larval and adult type red blood cell (RBC) nuclei during metamorphosis was studied with the use of Scatchard plots. The number of binding sites per nucleus or the maximum binding capacity (MBC) in the larval type RBC (containing larval hemoglobins) increased from 240 at stage X to 410 at stage XVIII. The apparent Kd also increased from 170 pM at stage X to 260 pM at stage XVIII. At stages XX-XXI, adult type RBCs (containing adult hemoglobins) accounted for 22% of the RBC population. At stage XXII, adult RBCs increased to 54%. In the separated larval and adult RBCs from stage XXII, obtained by Percoll gradient centrifugation, the MBC in larval RBCs had increased to 930 sites/nucleus, a 4-fold increase over stage X, with a Kd of 760 pM; adult RBCs had 1150 sites/nucleus and a Kd of 1530 pM. In contrast, when T3 binding was measured in the whole RBC population from stage XXII tadpoles, the MBC and Kd in larval and adult RBC were estimated at 780 sites/nucleus with a Kd of 640 pM and 1200 sites/nucleus with Kd of 2300 pM, respectively. After metamorphosis, the larval RBCs disappeared and were replaced by adult RBCs. The MBC in adult RBC had declined to 350 sites/nucleus at stage XXV (froglet). The Kd of adult RBC decreased to 790 pM at stage XXV. When stage XVII tadpoles were given a single injection of T3, a 2-fold increase in MBC and Kd was observed after 14 days at 20 C. The values observed for T3-treated animals (870 sites/nucleus with a Kd of 660 pM) were close to the values for the larval RBCs at stage XXII in spontaneous metamorphosis. The MBC and apparent dissociation constant of control animals (400 sites/nucleus; Kd of 270 pM) were almost the same as at stage XVIII in spontaneous metamorphosis. Treatment with PRL (10 micrograms/10 g tadpole every 2 days) had no effect on the T3-binding sites of RBC nuclei. This dose is adequate to stimulate tail fin growth and inhibit hind limb growth. The same dose of PRL did not affect the T3-induced increase in the MBC and Kd of the T3-binding sites. The results are discussed in terms of the positive regulation of the number of T3 receptors by thyroid hormones. The metamorphic response in amphibia appears to require an increase in the number of T3-binding sites (Receptor Induction).