Purpose We examined how heat exposure affects muscle and cerebral oxygenation kinetics using statistical parametric mapping (SPM) during repeated treadmill sprints. Methods Eleven recreationally active males performed three sets of five 5-s sprints with 25 s of recovery and 3 min between sets in hot (38 °C) and temperate (25 °C) conditions. Continuous measurements of muscle (vastus lateralis) and cerebral (prefrontal cortex) tissue oxygenation were obtained using near-infrared spectroscopy. One-way ANOVA SPMF statistics were applied to pooled sprint data, with each condition including 15 time-series (three sets of five sprints) combined. Each time-series included the sprint and subsequent recovery phases. Results Muscle tissue saturation index further decreased in hot compared to temperate condition, from the middle of the 5-s sprint phase (~ 2.9 s) until the end of the recovery period (p textless 0.001), while total hemoglobin concentration was significantly higher in the early phase of recovery (from 5.1 to 11.8 s, p = 0.003). Cerebral tissue saturation index decreased from 0.7 s to 13.0 s (p textless 0.001) in the heat. Total hemoglobin concentration was lower in hot condition during both the sprint phase and the initial third of the recovery (from 0 to 11.7 s, p textless 0.001), as well as during the recovery (from 20.5 to 24.8 s, p = 0.007). Conclusion Adding heat stress to repeating treadmill sprints further lowered muscle oxygenation levels during both the sprint and recovery phases, and limited cerebral tissue perfusion during the sprint and the initial recovery phases. The use of SPM for continuous analysis of near-infrared spectroscopy data provides new insights beyond summary statistics.