Antarctic notothenioid fish are characterized by their evolutionary adaptation to the cold, thermostable Southern Ocean, which is associated with unique physiological adaptations to withstand the cold and reduce energetic requirements but also entails limited compensation capacities to environmental change. This study compares the capacities of mitochondrial acclimation to ocean warming and acidification between the Antarctic nototheniid Notothenia rossii and the sub-Antarctic Lepidonotothen squamifrons, which share a similar ecology, but different habitat temperatures. After acclimation of L. squamifrons to 9°C and N. rossii to 7°C (normocapnic/hypercapnic, 0.2 kPa CO2/2000 ppm CO2) for 4-6 weeks, we compared the capacities of their mitochondrial respiratory complexes I (CI) and II (CII), their P/O ratios (phosphorylation efficiency), proton leak capacities and mitochondrial membrane fatty acid compositions. Our results reveal reduced CII respiration rates in warm-acclimated L. squamifrons and cold hypercapnia-acclimated N. rossii. Generally, L. squamifrons displayed a greater ability to increase CI contribution during acute warming and after warm-acclimation than N. rossii. Membrane unsaturation was not altered by warm or hypercapnia-acclimation in both species, but membrane fatty acids of warm-acclimated L. squamifrons were less saturated than in warm normocapnia-/hypercapnia-acclimated N. rossii. Proton leak capacities were not affected by warm or hypercapnia-acclimation of N. rossii. We conclude that an acclimatory response of mitochondrial capacities may include higher thermal plasticity of CI supported by enhanced utilization of anaplerotic substrates (via oxidative decarboxylation reactions) feeding into the citrate cycle. L. squamifrons possesses higher relative CI plasticities than N. rossii, which may facilitate the usage of energy efficient NADH-related substrates under conditions of elevated energy demand, possibly induced by ocean warming and acidification. The observed adjustments of electron transport system complexes with a higher flux through CI under warming and acidification suggest a metabolic acclimation potential of the sub-Antarctic L. squamifrons, but only limited acclimation capacities for N. rossii. © 2013 Strobel et al.