The regulatory effects of thyroid hormone on adenohypophysial substance P (SP) were studied in heterotopically implanted anterior pituitaries. Three or four anterior pituitaries from 21-day-old rat pups were implanted under the renal capsule in 175- to 200-g adult rats. The donor and recipient animals were sex matched. One week after implantation, animals were thyroidectomized or sham operated. A separate group of animals received daily T4 treatment (1.5 g/100 g, ip). After 2 weeks, the native and heterotopic pituitaries were assayed for SP, TSH, PRL, and LH. Thyroidectomy resulted in a 3- to 10-fold increase in the SP concentration in both the heterotopic and native pituitaries compared to euthyroid values. T4 treatment suppressed the SP levels in the heterotopic pituitaries of the thyroidectomized rats. In contrast to the reduction of TSH concentrations in native pituitaries in thyroidectomized animals vs. controls, TSH concentrations in the heterotopic pituitaries of thyroidectomized rats were approximately 10 times greater than those in euthyroid animals. PRL concentrations were unaffected by hypothyroidism in native and heterotopic pituitaries. Thyroidectomy resulted in a decrease in LH concentrations in the native anterior pituitary, without affecting LH concentrations in the implanted pituitary. These findings indicate that a direct link from the hypothalamus to the anterior pituitary is not required for the pituitary SP response to hypothyroidism.