Uptake and binding of thyroxine and triiodothyronine by rat diaphragm in vitro

Abstract

Hemidiaphragms from weanling Sprague-Dawley rats were incubated in Krebs-Ringer buffer containing 1.3 x 10^–7 m l-thyroxine or l-triiodothyronine labeled with radioiodine. Triiodothyronine uptake was about twice as rapid as thyroxine uptake. The data were consistent with a kinetic analysis based on a two-step process: 1) simple diffusion into the tissue (rate-limiting step) and 2) binding to cellular proteins (a process that occurs very rapidly compared to diffusion). The diffusion constants were calculated to be k_d(T₄) = 0.153 cm³/min and k_d(T₃) = 0.352 cm³/min. The data suggest a single species of tissue-binding site with identical tissue-binding capacity and binding affinity for both triiodothyronine and thyroxine. Maximum binding was found to be 28 x 10^–4 µm/g tissue and the affinity constant for binding was 1.4 x 10⁴ g/µm. The difference in the rate of accumulation of thyroxine and triiodothyronine appears to be due only to the difference in the diffusion rate. A direct rate equation for the accumulation of thyroid hormone was derived. Points calculated by means of this equation fit remarkably well with the experimental data.