Simple Summary:& nbsp;Cortisol is a key stress hormone in teleosts. Cortisol exerts its effects through genomic-and membrane-initiated mechanisms, however, the role of the latter in long-term stress responses is unknown. Here, we treated Sparus aurata with cortisol or cortisol-BSA (exclusive inductor to membrane-initiated effects) to emulate a long-term stress situation. We found that cortisol, but not cortisol-BSA, promotes energy substrate mobilization in the liver, together with the regulation of metabolism-related genes. We suggest that genomic cortisol actions exclusively participate in metabolic responses during prolonged treatment using cortisol in S. aurata. This study contributes to the current knowledge on cortisol's involvement in stress responses in fish.Cortisol is the main glucocorticoid hormone promoting compensatory metabolic responses of stress in teleosts. This hormone acts through genomic and membrane-initiated actions to exert its functions inside the cell. Experimental approaches, using exogenous cortisol administration, confirm the role of this hormone during short (minutes to hours)- and long-term (days to weeks) responses to stress. The role of membrane-initiated cortisol signaling during long-term responses has been recently explored. In this study, Sparus aurata were intraperitoneally injected with coconut oil alone or coconut oil containing cortisol, cortisol-BSA, or BSA. After 3 days of treatment, plasma, liver, and skeletal muscle were extracted. Plasma cortisol, as well as metabolic indicators in the plasma and tissues collected, and metabolism-related gene expression, were measured. Our results showed that artificially increased plasma cortisol levels in S. aurata enhanced plasma glucose and triacylglycerols values as well as hepatic substrate energy mobilization. Additionally, cortisol stimulated hepatic carbohydrates metabolism, as seen by the increased expression of metabolism-related genes. All of these responses, observed in cortisol-administered fish, were not detected by replicating the same protocol and instead using cortisol-BSA, which exclusively induces membrane-initiated effects. Therefore, we suggest that after three days of cortisol administration, only genomic actions are involved in the metabolic responses in S. aurata.