Anatase is a metastable polymorph of TiO2. In contrast to the more widely studied TiO2 rutile, O vacancies (V(O)'s) are not stable at the anatase (101) surface. Low-temperature STM shows that surface V(O)'s, created by electron bombardment at 105 K, start migrating to subsurface sites at temperatures ≥200 K. After an initial decrease of the V(O) density, a temperature-dependent dynamic equilibrium is established where V(O)'s move to subsurface sites and back again, as seen in time-lapse STM images. We estimate that activation energies for subsurface migration lie between 0.6 and 1.2 eV; in comparison, density functional theory calculations predict a barrier of ca. 0.75 eV. The wide scatter of the experimental values might be attributed to inhomogeneously distributed subsurface defects in the reduced sample.