Double immunostaining for c-fos and oxytocin (OXY) was used to study the topography and time course of the metabolic activation of the hypothalamic oxytocinergic system upon osmotic stress in the male rat. Animals injected i.p. with hypertonic saline expressed c-fos-like immunoreactivity (FLI) in the paraventricular (PVN), periventricular (PEV) and supraoptic (SON) hypothalamic nuclei, and in the preoptic and retrochiasmatic regions, as early as 30 min after stimulation and up to 6 h, while these areas were mostly devoid of staining in isotonic saline-injected animals. The activation of the oxytocinergic system peaked at 30 min and declined at different rates in the PVN and in the SON after 90 min. The maximal percentage of OXY neurons expressing FLI upon osmotic stress was about 80% in the SON, PEV and LSN, 60% in the PVN and 50% in the medial preoptic area. Activated OXY neurons were found in both the magnocellular and parvocellular divisions of the system. These data show that OXY nuclei in the rat hypothalamus are differentially activated by osmotic stress. They also suggest a role of OXY in the central as well as in the humoral response to changes in plasma osmolarity.