Effects of severe hypercapnia have been extensively studied in marine fishes, while the knowledge on the impacts of moderately elevated CO2 levels and their combination with warming is scarce. Here we investigate ion regulation mechanisms and energy budget in gills from Atlantic cod acclimated long-term to elevated PCO2 levels (2,500 μatm) and temperature (18°C). Isolated perfused gill preparations were established to determine gill thermal plasticity during acute exposures (10-22°C) and in vivo costs of Na(+)/K(+)-ATPase activity and of protein and RNA synthesis. Maximum enzyme capacities of F1Fo-ATPase, H(+)-ATPase and Na(+)/K(+)-ATPase were measured in vitro in crude gill homogenates. After whole animal acclimation to elevated PCO2 and/or warming, branchial oxygen consumption responded more strongly to acute temperature change. The fractions of gill respiration allocated to protein and RNA synthesis remained unchanged. In gills of fish CO2-exposed at both temperatures, energy turnover associated with Na(+)/K(+)-ATPase activity was reduced by 30 percent below rates of the control group. This contrasted in vitro capacities of Na(+)/K(+)-ATPase, which remained unchanged under elevated CO2 at 10°C, and earlier studies which had found a strong upregulation under more severe hypercapnia. F1Fo-ATPase capacities increased in hypercapnic gills at both temperatures, whereas Na(+)/K(+)ATPase and H(+)-ATPase capacities only increased in response to elevated CO2 and warming indicating the absence of thermal compensation under CO2. We conclude that in vivo ion-regulatory energy demand is lowered under moderately elevated CO2 levels despite the stronger thermal response of total gill respiration and the upregulation of F1Fo-ATPase. This effect is maintained at elevated temperature.