The present study sought to determine whether water deprivation increases Fos immunoreactivity, a neuronal marker related to synaptic activation, in sympathetic- regulatory neurons of the hypothalamic paraventricular nucleus (PVN). Fluorogold (4%, 50 nl) and cholera toxin subunit B (0.25%, 20 - 30 nl) were microinjected into the spinal cord (T-1 - T-3) and rostral ventrolateral medulla (RVLM), respectively. Rats were then deprived of water but not food for 48 h. Water deprivation significantly increased the number of Fos-positive nuclei throughout the dorsal, ventrolateral, and lateral parvocellular divisions of the PVN ( water deprived, 215 +/- 23 cells; control, 45 +/- 7 cells, P < 0.01). Moreover, a significantly greater number of Fos-positive nuclei were localized in spinally projecting ( 11 +/- 3 vs. 2 +/- 1 cells, P < 0.025) and RVLM-projecting ( 45 +/- 7 vs. 7 +/- 1 cells, P < 0.025) neurons of the PVN in water-deprived vs. control rats, respectively. The majority of these double-labeled neurons was found in the ventrolateral and lateral parvocellular divisions of the ipsilateral PVN. Interestingly, a significantly greater percentage of RVLM-projecting PVN neurons were Fos positive compared with spinally projecting PVN neurons in the ventrolateral (25.8 +/- 0.7 vs. 8.0 +/- 1.5%, respectively, P < 0.01) and lateral ( 23.4 +/- 2.1 vs. 5.0 +/- 0.9%, respectively, P > 0.01) parvocellular divisions. In addition, we analyzed spinally projecting neurons of the RVLM and found a significantly greater percentage were Fos positive in water-deprived rats than in control rats ( 26 +/- 3 vs. 3 +/- 1%, respectively; P < 0.001). Collectively, the present findings indicate that water deprivation evokes a distinct cellular response in sympathetic- regulatory neurons of the PVN and RVLM.