The metabolite of dihydrotestosterone, 5α-androstane-3α,17β-diol (3α-Diol), is a potent inhibitor of estrogen-induced gonadotropin and prolactin secretion and lordosis behavior in the female rat. This study examined whether 3α-Diol can counteract the ultrastructural changes which are known to occur in the ventromedial nucleus (VMN) of the hypothalamus following estrogen treatment. Ovariectomized rats were treated with estradiol (E<sub>2</sub>; n = 7), 3α-Diol (n = 5), E<sub>2</sub> and 3α-Diol (n = 6), or received control (n = 6) treatments. E<sub>2</sub> was administered in subcutaneous capsules for two discontinuous 2-hour periods separated by 5 h, a ‘pulsed’ treatment regimen known to mimic the timing of endogenous E<sub>2</sub> action and to influence neuronal ultrastructure in the VMN. Animals given 3α-Diol received subcutaneous injections (6 mg/kg) 3 h prior to each implantation of E<sub>2</sub> or empty capsules. Control animals received vehicle 3 h prior to implantation of blank capsules. Animals were perfused 24 h after initial hormone treatment and neurons from the ventrolateral portion of the VMN were examined using electron microscopy. Separately, both E<sub>2</sub> and 3α-Diol treatment increased somal and nuclear size, altered somal and nuclear shape, and increased the numbers of lysosomes present in the cytoplasm above control levels. E<sub>2</sub> treatment resulted in increased stacking of the rough endoplasmic reticulum while 3α-Diol treatment resulted in an unusual plexiform rough endoplasmic reticulum distribution. In contrast, combined treatment with E<sub>2</sub> and 3α-Diol resulted in cells which were similar to ovariectomized control cells on these measures. All steroid treatments decreased the amount of heterochromatin present within the nucleus compared to that seen in controls. Thus, 3α-Diol influences the ultrastructural characteristics of neurons within the VMN in a manner somewhat though not altogether similar to E<sub>2</sub>. However, 3α-Diol given in combination with E<sub>2</sub> counteracts or prevents the actions of E<sub>2</sub> within these same neurons.