We investigated the underlying molecular mechanisms by which post-exercise cold-water immersion (CWI) may alter key markers of mitochondrial biogenesis following both a single session and six weeks of sprint interval training (SIT). Nineteen males performed a single SIT session, followed by one of two 15-min recovery conditions: cold-water immersion (COLD; 10°C) or a passive room-temperature control (CON; 23°C). Sixteen of these participants also completed six weeks SIT, each session followed immediately by their designated recovery condition. Four muscle biopsies were obtained in total, three during the single SIT session (pre-exercise, post-recovery, and 3 h post-recovery), and one 48h after the last SIT session. Following a single SIT session, phosphorylated (p-) AMPK, p-p38 MAPK, p-p53 and PGC1α mRNA were all increased (P < 0.05). Post-exercise CWI had no effect on these responses. Consistent with the lack of a response following a single session, regular post-exercise CWI had no effect on PGC-1α or p53 protein content. Six weeks of SIT increased peak aerobic power, VO_2peak, maximal uncoupled respiration (complexes I and II), and 2-km time-trial performance (P < 0.05). However, regular CWI had no effect on changes in these markers, consistent with the lack of response in the markers of mitochondrial biogenesis. While these observations suggest CWI is not detrimental to endurance adaptations following six weeks of SIT, they question whether post-exercise CWI is an effective strategy to promote mitochondrial biogenesis and improvements in endurance performance.