We studied the basal and thyrotropin-releasing hormone (TRH) (50 nM) induced thyrotropin (TSH) release in isolated hemipituitaries of ovariectomized rats treated with near-physiological or high doses of 17-beta-estradiol benzoate (EB; sc, daily for 10 days) or with vehicle (untreated control rats, OVX). One group was sham-operated (normal control). The anterior pituitary glands were incubated in Krebs-Ringer bicarbonate medium, pH 7.4, at 37 degrees C in an atmosphere of 95% O2/5% CO2. Medium and pituitary TSH was measured by specific RIA (NIDDK-RP-3). Ovariectomy induced a decrease (P < 0.05) in basal TSH release (normal control = 44.1 +/- 7.2; OVX = 14.7 +/- 3.0 ng/ml) and tended to reduce TRH-stimulated TSH release (normal control = 33.0 +/- 8.1; OVX = 16.6 +/- 2.4 ng/ml). The lowest dose of EB (0.7 microgram/100 g body weight) did not reverse this alteration, but markedly increased the pituitary TSH content (0.6 +/- 0.06 microgram/hemipituitary; P < 0.05) above that of OVX (0.4 +/- 0.03 microgram/hemipituitary) and normal rats (0.46 +/- 0.03 microgram/hemipituitary). The intermediate EB dose (1.4 micrograms/100 g body weight) induced a nonsignificant tendency to a higher TSH response to TRH compared to OVX and a lower response compared to normal rats. Conversely, in the rats treated with the highest dose (14 micrograms/100 g body weight), serum 17-beta-estradiol was 17 times higher than normal, and the basal and TRH-stimulated TSH release, as well as the pituitary TSH content, was significantly (P < 0.05) reduced compared to normal rats and tended to be even lower than the values observed for the vehicle-treated OVX group, suggesting an inhibitory effect of hyperestrogenism. In conclusion, while reinforcing the concept of a positive physiological regulatory role of estradiol on the TSH response to TRH and on the pituitary stores of the hormone, the present results suggest an inhibitory effect of high levels of estrogen on these responses.