Previous studies have shown that Senegalese sole is partially euryhaline in the juvenile phase, being able to adapt to a wide range of salinities in a short-time period, due to changes at the osmoregulatory and metabolic level. This study aimed to assess the effects of acclimation of sole to a wide range of salinities, with a special emphasis on the role of plasma amino acids during this process. Sole juveniles were acclimated for 2 weeks to different salinities: 5, 15, 25, 38, and 55 g L−1. Plasma levels of cortisol, glucose, osmolality, and free amino acids were assessed at the end. Changes in plasma levels of cortisol, glucose, and amino acids indicate that fish reared at 5 and 55 g L−1 were facing extra energy costs. Amino acids seem to play an important role during salinity acclimation, either as energy sources or as important osmolytes for cell volume regulation.