Background The correlation between brain injury and high-altitude (HA) exposure duration (Dur_HA) as well as peripheral oxygen saturation (SpO(2)) remains unclear. Purpose To evaluate the global and regional brain volume differences between HA immigrants and sea-level residents, and the relationship between brain volume with Dur_HA and SpO(2). Material and Methods Structural magnetic resonance imaging (MRI) scans were acquired in 33 healthy male HA immigrants (HA group) and 33 matched sea-level male residents (SL group). Differences in global gray matter volume (GMV), white matter volume (WMV), brain parenchyma volume (BV), total intracranial volume (TIV), and the volume-fraction (the ratio of GMV/TIV, WMV/TIV, BV/TIV) were assessed. Regional gray matter differences were investigated using voxel-based morphology analysis. The volume of clusters with GM loss were calculated as the volume of volume of interest (V_VOI). Student's t-test and partial correlation were adopted for statistic calculation. Results Compared to the SL group, the HA immigrants had larger WMV (P = 0.015), smaller ratio of GMV/WMV (P = 0.022), and regional gray matter loss in bilateral basal ganglion, limbic system, midbrain, and vermis (cluster size >100 voxels, family-wise error corrected at P = 0.01). The global GMV, BV, and V_VOI confined to vermis had negative correlations with the Dur_HA (r = -0.369, P = 0.049; r = -0.380, P = 0.042; and r = -0.471, P = 0.010. Neither global nor regional brain volume correlated with SpO(2). Conclusion Global and regional brain are affected by long-term HA exposure, and global and regional gray matter have a time-dependent volume loss.