Sex differences in systemic morphine analgesia occur with male rodents displaying significantly greater analgesic magnitudes and potencies than females. Neonatal androgenization, and to a lesser degree, adult ovariectomy enhance systemic morphine analgesia in female rats, implicating both organizational and activational effects of gonadal hormones. The neuroanatomical circuits sensitive to sex-related hormones by which females display a smaller opiate analgesic effect is not clear, but the ventromedial (VMH) and medial preoptic (MPOA) hypothalamic nuclei are critical in the monitoring of estradiol and other sex hormone levels. To assess the contribution of these nuclei to sex and adult gonadectomy differences in systemic morphine analgesia, intact male, intact female and adult ovariectomized (OVEX) female rats received bilateral saline (SAL) or ibotenic acid (IBO) microinjections into either the VMH or MPOA. Following surgeries, baseline tail-flick latencies over 120 minutes (min) were assessed over 4 days in all nine groups with intact females tested in the estrus phase of their cycle. All animals then received an ascending series of morphine (1.0, 2.5, 5.0, 7.5, 10.0mg/kg) injections 30min prior to the tail-flick test time course with 8-12 day inter-injection intervals between doses. Baseline latencies failed to differ between SAL-treated intact males and females, but were significantly higher in SAL-treated OVEX females. Both VMH IBO and MPOA IBO lesions increased baseline latencies in intact male and female rats, but not in OVEX females. SAL-treated intact males (ED(50)=4.0mg/kg) and SAL-treated OVEX females (ED(50)=3.5mg/kg) displayed significantly greater potencies of systemic morphine analgesia than SAL-treated intact females (ED(50)=6.3mg/kg), confirming previous gender and gonadectomy differences. Neither VMH IBO (ED(50)=3.7 mg/kg) nor MPOA IBO (ED(50)=4.1mg/kg) males differed from SAL-treated males in the potency of systemic morphine analgesia. In contrast, VMH IBO (ED(50)=4.1mg/kg) and MPOA IBO (ED(50)=3.5mg/kg) intact females displayed significantly greater potencies in systemic morphine analgesia than SAL-treated intact females. However, VMH IBO OVEX (ED(50)=3.5mg/kg) and MPOA IBO OVEX (ED(50)=3.9 mg/kg) failed to differ from SAL-treated OVEX females in the potency of systemic morphine analgesia. The magnitudes of systemic morphine analgesia as measured by Maximum Percentage Effect values displayed similar patterns, but lesser degrees, of effects. These data suggest that VMH and MPOA nuclei act to tonically inhibit endogenous pain-inhibitory circuits in the intact female, but not intact male brain, and that removal of circulating gonadal hormones by OVEX and/or excitotoxic destruction of these estrogen receptor accumulating nuclei disinhibit the female analgesic response to systemic morphine.