To investigate a possible role of TSH in the regulation of its own receptor, a sensitive assay of [125I]TSH tropin binding to rat thyroid membranes was used. With 1 mM MgCl2 in the buffer, Scatchard analysis of displacement of TSH gave a curvilinear plot with a high affinity, low capacity (K1, 3.4 nM; Q1, 3.1 pmol/microgram) and a low affinity, high capacity binding site (K2, 0.54 microM; Q2, 1.2 X 0.1 nmol/microgram). Feeding rats propylthiouracil led to a decrease in [125I]TSH binding (expressed either per U protein or per wet wt of tissue) that was related to the duration of treatment. Evaluation by Scatchard analysis showed that this was due to a loss of binding sites, e.g. a 50-60% decrease after 1 month of propylthiouracil treatment; affinity was actually slightly increased in the goitrous tissue. This change in the number of TSH-binding sites was readily reversed in association with the suppression of TSH in vivo either by injections of T3 for 3 days or more by feeding a normal diet for 1 month. Thus, the data are compatible with TSH, that is in high concentration in the serum of rats fed propylthiouracil, exerting a downregulatory influence on its own receptors.