Autophagy is an evolutionary conserved cellular degradation system implicated in maintaining health and promoting longevity. Few human data exist investigating the autophagic response to exercise; however, acute moderate-intensity, continuous exercise (MICT) has been shown to stimulate autophagy in skeletal muscle. Presently, it is unknown whether high-intensity interval training (HIIT) exercise induces autophagy. PURPOSE: The purpose of this study was to compare the autophagy response of an acute bout of HIIT exercise (treadmill running) to MICT exercise in human skeletal muscle. METHODS: Using a crossover design, ten recreationally-active males (n=5) and females (n=5) performed a bout of MICT (60 minutes at 55% of max velocity [Vmax]]) and HIIT (6 bouts of 1 minute at 100% Vmax and 1 minute at 3 MPH, followed by 5 minutes at 3 MPH, followed by 6 bouts of 1 minute and 100% Vmax and 1 minute at 3 MPH). Muscle biopsies from the vastus lateralis were taken pre- and 3 hours post-exercise. Exercise bouts were separated by ≥72 hours and performed after abstaining from alcohol for ≥24 hours and food and caffeine for ≥8 hours. Subjects also refrained from food, energy-containing beverages, and caffeine during the 3-hour post-exercise period prior to the muscle biopsy. Muscle tissue was analyzed for protein expression of markers of autophagy (LC3I, LC3II) and autophagy signaling (p38MAPK) via western blot analysis. RESULTS: No differences were detected for LC3I, LC3II, and p38MAPK protein content measured 3 hours post-exercise compared to pre-exercise in both HIIT and MICT bouts (p>0.05). LC3II:LC3I ratio increased 3 hours post-exercise in HIIT (162.4 ±: 45.9%), which was significantly higher than MICT at 3 hours post-exercise which decreased from pre-exercise (48.8± 9.4%; p<0.05). CONCLUSION: Our findings show that despite discrepant durations and intensities, HIIT stimulates autophagy in human skeletal muscle, however, in a distinct fashion compared to MICT. Our data also add to the current literature demonstrating that autophagy is activated by continuous (≥ 60 minutes), moderate-intensity (55 - 70% VO2max) exercise.