INTRODUCTION: In prolonged spaceflights the effect of long-term confinement on the autonomic regulation of the heart is difficult to separate from the effect of prolonged exposure to microgravity or other space-related stressors. Our objective was to investigate whether the sleep-wake variations in the autonomic control of the heart are specifically altered by long-term confinement during the 105-d pilot study of the Earth-based Mars500 project. METHODS: Before (pre), during (T1: 30, T2: 70, andT3: 100 d), and after (post) confinement, 24-h EKG records were obtained from the six crewmembers that participated in the mission. Sleep and wake periods were determined by fitting a square wave to the data. Autonomic activity was evaluated through time and frequency domain indexes of heart rate variability (HRV) analysis during wake and sleep periods. RESULTS: During confinement, wake HRV showed decreased mean heart rate and increased amplitude at all frequency levels, particularly in the very low (pre: 13.3 +/- 0.2; T1: 13.9 +/- 0.3; T2: 13.9 +/- 0.2; T3: 13.9 +/- 0.2; post: 13.2 +/- 0.2) and high (pre: 7.6 +/- 0.4; T1: 8.3 +/- 0.5; T2: 8.2 +/- 0.4; T3: 8.1 +/- 0.4; post: 7.6 +/- 0.3) frequency components (values expressed as mean +/- SE of wavelet power coefficients). Sleep HRV remained constant, while sleep-wake high frequency HRV differences diminished. DISCUSSION: The observed autonomic changes during confinement reflect an increase in parasympathetic activity during wake periods. Several factors could account for this observation, including reduced daylight exposure related to the confinement situation.