Breast-feeding reduces fertility, and this seems to be related, in part, to an enhancement of the sensitivity of the GnRH system to the negative feedback effects of estradiol related to suckling. Previously, we showed that short-term treatment with small doses of estradiol delivered transdermally suppress plasma gonadotropin concentrations in breast-feeding women. We have now monitored the effects on ovarian function of longer-term low-dose estradiol treatment using plasma inhibin B and inhibin A concentrations and ultrasonography. Breast-feeding women (n = 45) using barrier methods of contraception were enrolled at 6 weeks postpartum and followed up to 18 weeks PP. Nineteen women agreed to being randomized to wear either an estrogen [transdermal estradiol supplementation (TES); n = 10; Estraderm, 50 microg/24 h] or a placebo (PL; n = 9) patch for 12 weeks, whereas the remaining 26 women acted as untreated controls. TES did not significantly increase plasma estradiol concentrations. Plasma FSH levels decreased from 6.1+/-0.8 U/L to 3.3+/-0.6 U/L after 2 weeks of treatment (P < 0.01) and were lower in the TES group compared with the PL group at all times during the treatment (at least P < 0.05). Plasma LH concentrations in the TES group were lower than in the PL group after 4, 6, 8, and 10 weeks of estrogen treatment (at least P < 0.05). Throughout the study, no ovarian follicles detected by ultrasound were greater than 10 mm in diameter. Nevertheless, after 2 weeks of treatment, plasma inhibin B concentrations were significantly lower in the TES group than in the PL group (15.5+/-5.8 vs. 64.9+/-11.1 ng/L; P < 0.01) and remained significantly (P < 0.01) suppressed throughout the treatment, suggesting a suppression of the functional ovarian activity during TES. Inhibin A levels remained low in all groups (3-45 ng/L) but were suppressed further by TES treatment with no levels greater than 7 ng/L. We conclude that low-dose estradiol treatment given as TES suppresses ovarian activity as measured by inhibins B and A by reducing the secretion of LH and FSH during breast-feeding for several weeks. This supports the concept that suckling-induced suppression of the GnRH system is associated with an enhancement of the negative effects of estradiol on the hypothalamic GnRH system. Furthermore, because the contraceptive efficacy of breast-feeding is complicated by the unpredictable early return of ovarian activity in some women, TES could be the basis for the development of a novel contraceptive for breast-feeding women.