Estradiol (E₂) applied topically twice weekly to mouse skin at doses as low as 1 nmol inhibited hair growth by blocking the transition of the hair follicle from the resting phase (telogen) to the growth phase (anagen). In contrast, application of ≤10 nmol of other steroids produced limited inhibition. Topical treatment with the estrogen receptor (ER) antagonist ICI-182780 reversed the effects of E₂, and when applied alone, ICI-182780 caused a telogen-to-anagen transition. Both E₂ and ICI-182780 were highly effective at their site of application but not at distant sites, indicating the direct rather than secondary systemic nature of their effects. Western analysis detected a 65-kDa ER-α immunoreactive dermal protein, and Northern analysis revealed the presence of a 6.7-kb ER-α mRNA. A ribonuclease protection assay confirmed the presence of ER-α transcripts but failed to detect ER-β transcripts. These findings implicate a skin-specific ER-α pathway in the regulation of the hair follicle cycle.