Background: Body position alters aspects of pulmonary function in health and disease. Although studies have assessed positional effects on the heart and lungs, little is known about positional changes in gas exchange parameters at high altitude. We hypothesized that following ascent, supine positioning would cause lower oxygen saturation than sitting, partially due to decreased ventilation and increased partial pressure of end-tidal carbon dioxide (Petco2). Materials and Methods: Twenty-eight healthy subjects were studied at sea level and following gradual ascent to 5150 m. After 10 minutes of sitting rest, subjects were studied for 5 minutes each in the sitting, supine, and prone positions with the order randomly assigned. Pulse oximeter oxygen saturation (SpO2), minute ventilation (VE), end-tidal O2 (Peto2) and Petco2, oxygen consumption, and CO2 production were continuously measured. Alveolar ventilation (VA) was calculated from the measured parameters. Results: At high altitude, VE was not affected by body position (12.96 ± 3.09 and 11.54 ± 3.45 L/min in the sitting and supine positions, respectively, p = 0.255). Petco2 increased from sitting to supine (22.8 ± 3.1 to 23.5 ± 3.3 mm Hg, p < 0.005), but VE and Petco2 were not different between the supine and prone positions. Calculated VA was not significantly affected by body position at either sea level or high altitude. SpO2 decreased from 81.3% ± 4.4% sitting to 78.8% ± 6.0% supine (p = 0.025), with a mean positional SpO2 difference of 2.5% ± 4.9% (95% confidence interval 0.6%-4.4%). SpO2 was not different between the supine and prone positions. Twenty-two of 28 subjects had lower SpO2 supine compared with sitting. Conclusions: These results extend earlier low-altitude studies and demonstrate the importance of postural regulation in different environments. As 79% of subjects had lower SpO2 while supine than sitting, control of body position is necessary for SpO2 comparisons at altitude to be meaningful.