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Metabolic rates are linked to the energetic costs of different activities of an animal’s life. However, measuring the metabolic rate in free-swimming fish remains challenging due to the lack of possibilities to perform these direct measurements in the field. Thus, the calibration of acoustic transmitters with the oxygen consumption rate (MO₂) could be promising to counter these limitations. In this study, rainbow trout (<i>Oncorhynchus mykiss</i> Walbaum, 1792; n = 40) were challenged in a critical swimming test (U_crit) to (1) obtain insights about the aerobic and anaerobic metabolism throughout electromyograms; and (2) calibrate acoustic transmitters’ signal with the MO₂ to be later used as a proxy of energetic costs. After this calibration, the fish (n = 12) were implanted with the transmitter and were followed during ~50 days in an aquaculture facility, as a case study, to evaluate the potential of such calibration. Accelerometer data gathered from tags over a long time period were converted to estimate the MO₂. The MO₂ values indicated that all fish were reared under conditions that did not impact their health and welfare. In addition, a diurnal pattern with higher MO₂ was observed for the majority of implanted trout. In conclusion, this study provides (1) biological information about the muscular activation pattern of both red and white muscle; and (2) useful tools to estimate the energetic costs in free-ranging rainbow trout. The use of acoustic transmitters calibrated with MO₂, as a proxy of energy expenditure, could be promising for welfare assessment in the aquaculture industry.