Orexins and their receptors OX₁ and OX₂ regulate energy balance and sleep-wake cycle. We studied the expression of prepro-orexin (PPO), OX₁ and OX₂ in brain and pituitary under the influence of the hormonal status in adult rats. Primarily, PPO, OX₁ and OX₂ expression was determined in Sprague-Dawley female cycling rats during proestrus and in males. Animals were sacrificed at two hours intervals. Anterior (AH) and mediobasal (MBH) hypothalamus, anterior pituitary (P) and frontoparietal cortex (CC), were homogenized in TRIzol and mRNAs obtained for screening of PPO, OX₁, OX₂ expression by semiquantitative RT-PCR. Main findings were confirmed and extended to all days of the cycle by quantitative real-time RT-PCR. Hormones and food consumption were determined. Finally, OX₁, OX₂ and PPO were measured by real-time RT-PCR in tissues collected at 19:00 h of proestrus after treatments at 14:00 h with ovulation blocking agents, Cetrorelix or Pentobarbital. OX₁ and OX₂ expression increased at least three-fold in AH, MBH and P, but not in CC, between 17:00 h and 23:00 h of proestrus, without variations in estrus, diestrus or males. PPO in AH and MBH showed a four-fold or higher increase only during proestrous afternoon. Cetrorelix or Pentobarbital prevented increases of OX₁ and OX₂ only in the pituitary and blunted gonadotropin surges, but left OX₁, OX₂ and PPO brain expression unchanged. Reproduction, energy balance and sleep-wake cycle are integrated. Here we demonstrate that in the physiological neuroendocrine condition leading to ovulation, information to the orexinergic system acts in hypothalamus and pituitary by different mechanisms.