Gonadal steroid hormones are known to alter the expression of proopiomelanocortin (POMC) mRNA in neurons of the arcuate nucleus (ARC). These neurons send projections to the medial preoptic area (MPOA), wherein mu-opiate receptor density is cyclical and gonadal steroid hormone-dependent. Although beta-endorphin-(beta-Endo) content in the MPOA is known to vary across the estrous cycle, the effect of gonadal hormones on the distribution and density of beta-Endo-like immunoreactive (IR) fiber density in the preoptic area is unknown. In the present study, immunohistochemical staining was used to investigate the effects of gonadal steroid hormone treatment on beta-Endo-like IR fibers in the MPOA of ovariectomized (OVX) female rats. The density of beta-Endo-like IR fibers was low in the MPOA of OVX rats, but increased slightly following treatment with 17 beta-estradiol (E2) or 3 h after subsequent progesterone (P) injection. However, beta-Endo-like IR fiber density increased significantly 27 h after E2P treatment, and remained elevated 51 h after E2P treatment in the periventricular zone and in the medial portion of the medial preoptic nucleus, although the general distribution of fibers was unchanged. These results suggest that the density of MPOA beta-Endo innervation is normally gonadal steroid hormone-dependent and that the medial MPOA contains greater opioid tone than the lateral MPOA regardless of the hormonal state. Furthermore, since beta-Endo-like IR fiber density remained elevated even though gonadal hormone levels decreased, additional factors might modulate the release or turnover of beta-Endo in the MPOA during normal estrous cycling.