["The Journal of Physiology, EarlyView. ", "\nAbstract figure legend The mechanisms underpinning improved maximal oxygen uptake following sprint interval training (SIT) with post‐exercise blood flow restriction (BFR), compared to SIT alone, remain unclear. This study examined the effects of 6 week SIT with (BFR; n = 12) or without (CON; n = 8) post‐exercise BFR on V̇O2peak${{\\dot{V}}_{{{{\\mathrm{O}}}_2}{\\mathrm{peak}}}}$, lactate thresholds, citrate synthase (CS) activity and mitochondrial respiration in physically active males. There were time × condition interactions for CS activity (P = 0.011) and CS activity‐corrected CIIE (P = 0.047) and CIVE (P = 0.010), which increased following BFR (12.1%, P = 0.040; 74.3%, P = 0.030; 64.4%, P = 0.002, respectively) but not in CON (−4.6%, P = 0.053; 9.9% P = 0.460; –7.4%, P = 0.664, respectively). There were no between‐group differences in any performance marker changes (P ≥ 0.176). The addition of BFR to a 6 week SIT program increased mitochondrial content and uncoupled respiration.\n\n\n\n\n\n\n\n\n\nAbstract\nSprint interval training (SIT) combined with post‐exercise blood flow restriction (BFR) can augment adaptive signalling responses in skeletal muscle. However, mitochondrial adaptations to SIT with BFR are not well‐understood. This study examined the effects of a 6 week SIT program with or without post‐exercise BFR on skeletal muscle mitochondrial content and respiratory function, alongside physiological performance markers. Physically active males (n = 20; 25.3 ± 5.9 years; V̇O2peak${{\\dot{V}}_{{{{\\mathrm{O}}}_2}{\\mathrm{peak}}}}$, 52.5 ± 4.6 mL·min−1·kg−1) completed a SIT intervention (repeated 30 s sprints interspersed with 4.5 min of rest) with (BFR; n = 12) or without (CON; n = 8) post‐exercise BFR. Baseline and post‐training V̇O2peak${{\\dot{V}}_{{{{\\mathrm{O}}}_2}{\\mathrm{peak}}}}$ and lactate thresholds were measured and muscle biopsies obtained for determination of citrate synthase (CS) activity and mitochondrial respiration [O2 flux during leak (L), ADP‐stimulated oxidative phosphorylation (P) and uncoupled maximal electron transfer (E) states through mitochondrial complexes I–IV (CI–IV)]. There were time × condition interactions for CS activity (P = 0.011) and CS activity‐corrected CIIE (P = 0.047) and CIVE (P = 0.010), which increased following BFR (12.1%, P = 0.040; 74.3%, P = 0.030; 64.4%, P = 0.002, respectively) but not in CON (−4.6%, P = 0.053; 9.9% P = 0.460; −7.4%, P = 0.664, respectively). There were no between‐group differences in the changes in V̇O2peak${{\\dot{V}}_{{{{\\mathrm{O}}}_2}{\\mathrm{peak}}}}$ or any other performance markers (P ≥ 0.176). The addition of BFR to a 6 week SIT program increased mitochondrial content and uncoupled respiration in physically active males, which may have implications for improving skeletal muscle oxidative metabolism.\n\n\n\n\n\n\n\n\n\nKey points\n\nSprint interval training (SIT) with post‐exercise BFR (SIT+BFR) augments the exercise stimulus without limiting intensity and is shown to improve maximal oxygen uptake in athletes compared to SIT alone. However, the mechanisms underpinning this response remain unclear.\nThis study is the first to investigate the effects of SIT+BFR on citrate synthase (CS) activity and mitochondrial respiratory parameters.\nWe demonstrate that a 6 week SIT+BFR intervention in physically active males, increased CS activity and uncoupled mitochondrial respiration compared to SIT alone. However, improvements in performance determinants in response to SIT did not differ between groups.\nThese findings provide novel insight into the mitochondrial bioenergetic potential of BFR, when combined with SIT, and can inform recommendations for exercise training interventions.\n\n\n"]