Although high-altitude exposure can lead to neurocognitive impairment, even upon return to sea level, it remains unclear the extent to which brain volume and regional cerebral vascular reactivity ( CVR) are altered following high-altitude exposure. The purpose of this study was to simultaneously determine the effect of 3 weeks at 5050 m on: (1) structural brain alterations; and (2) regional CVR after returning to sea level for 1 week. Healthy human volunteers ( n = 6) underwent baseline and follow-up structural and functional magnetic resonance imaging ( MRI) at rest and during a CVR protocol (end-tidal PCO2 reduced by −10, −5 and increased by +5, +10, and +15 mmHg from baseline). CVR maps (% mmHg−1) were generated using BOLD MRI and brain volumes were estimated. Following return to sea level, whole-brain volume and gray matter volume was reduced by 0.4 ± 0.3% ( P < 0.01) and 2.6 ± 1.0% ( P < 0.001), respectively; white matter was unchanged. Global gray matter CVR and white matter CVR were unchanged following return to sea level, but CVR was selectively increased ( P < 0.05) in the brainstem (+30 ± 12%), hippocampus (+12 ± 3%), and thalamus (+10 ± 3%). These changes were the result of improvement and/or reversal of negative CVR to positive CVR in these regions. Three weeks of high-altitude exposure is reflected in loss of gray matter volume and improvements in negative CVR.