In the eurythermal cuttlefish Sepia officinalis, performance greatly depends on hearts that ensure systemic oxygen supply over a broad range of temperatures. We therefore aimed to identify adjustments in energetic cardiac capacity and underlying mitochondrial function supporting thermal acclimation that could be critical for the cuttlefish's competitive success in variable environments. Two genetically distinct cuttlefish populations were acclimated to 11°C, 16°C and 21°C, respectively. Subsequently, skinned and permeabilised heart fibres were used to assess mitochondrial functioning by means of high-resolution respirometry and a substrate-inhibitor protocol, followed by measurements of cardiac citrate synthase activity. In cuttlefish hearts, thermal sensitivity of mitochondrial substrate oxidation was high for proline and pyruvate but low for succinate. Oxygen efficiency of catabolism rose from 11°C to 21°C via shifts to oxygen-conserving oxidation of proline and pyruvate as well as via reduced proton leak. Acclimation to 21°C decreased mitochondrial complex I activity in Adriatic cuttlefish and increased complex IV activity in English Channel cuttlefish. However, compensation of mitochondrial capacities did not occur during cold acclimation to 11°C. Moreover, cold adapted English Channel cuttlefish had larger hearts with lower mitochondrial capacities than warm adapted Adriatic cuttlefish. The changes observed for substrate oxidation, mitochondrial complexes, proton leak or heart weights improve energetic efficiency and essentially seem to extend tolerance to high temperatures and reduce associated tissue hypoxia. We conclude that cuttlefish sustain cardiac performance and thus, systemic oxygen delivery over short and long-term changes of temperature and environmental conditions by multiple adjustments in cellular and mitochondrial energetics.