The purpose of the present study was to assess in the gilthead sea bream (Sparus aurata) the impact of different environmental salinities on the energy metabolism of tissues directly involved in the osmoregulatory process such as gills and kidney, and those probably related indirectly such as liver and brain. Therefore, several osmoregulatory parameters, the activities of several key enzymes, and levels of metabolites were measured in those tissues of fish acclimated to brackish water (BW, 12 p.p.t.), seawater (SW, 38 p.p.t) and hyper saline water (HSW, 55 p.p.t.) for 14 days. Plasma osmolality, and levels of sodium and chloride presented a clear relationship with environmental salinities, with fish acclimated to HSW showing higher values than those acclimated to BW. The results of the metabolic parameters clearly pointed to a general activation of energy metabolism in liver, gills, kidney and brain under different osmotic conditions. In liver, an enhancement of glycogenolytic and glycolytic potential was observed in fish acclimated to BW and HSW compared with those in SW. In plasma, an increased availability of metabolites such as glucose, lactate, and protein were observed in parallel with the increase in salinity. In gills, an increased Na⁺/K⁺-ATPase activity, a clear decrease in the capacity for use of exogenous glucose and the pentose phosphate pathway, as well as an increased glycolytic potential were observed in parallel with the increased salinity. In kidney, Na⁺/K⁺-ATPase activity and lactate levels increased in HSW, whereas the capacity for the use of exogenous glucose decreased in BW- and HSW-acclimated fish compared with SW-acclimated fish. In brain, fish acclimated to BW or HSW displayed an enhancement in their potential for glycogenolysis, use of exogenous glucose, and glycolysis compared with SW-acclimated fish. Also in brain, lactate and ATP levels decreased in parallel with the increase in salinity. The data are discussed in the context of energy expenditure associated with osmotic acclimation to different envinronmental salinities in fish euryhaline species.