BACKGROUND: There is limited information regarding the effects of spaceflight on the anatomical configuration of the brain and on cerebrospinal fluid (CSF) spaces. METHODS: We used magnetic resonance imaging (MRI) to compare images of 18 astronauts’ brains before and after missions of long duration, involving stays on the International Space Station, and of 16 astronauts’ brains before and after missions of short duration, involving participation in the Space Shuttle Program. Images were interpreted by readers who were unaware of the flight duration. We also generated paired preflight and postflight MRI cine clips derived from high-resolution, three-dimensional imaging of 12 astronauts after long-duration flights and from 6 astronauts after short-duration flights in order to assess the extent of narrowing of CSF spaces and the displacement of brain structures. We also compared preflight ventricular volumes with postflight ventricular volumes by means of an automated analysis of T1-weighted MRIs. The main prespecified analyses focused on the change in the volume of the central sulcus, the change in the volume of CSF spaces at the vertex, and vertical displacement of the brain. RESULTS: Narrowing of the central sulcus occurred in 17 of 18 astronauts after long-duration flights (mean flight time, 164.8 days) and in 3 of 16 astronauts after short-duration flights (mean flight time, 13.6 days) (P<0.001). Cine clips from a subgroup of astronauts showed an upward shift of the brain after all long-duration flights (12 astronauts) but not after short-duration flights (6 astronauts) and narrowing of CSF spaces at the vertex after all long-duration flights (12 astronauts) and in 1 of 6 astronauts after short-duration flights. Three astronauts in the long-duration group had optic-disk edema, and all 3 had narrowing of the central sulcus. A cine clip was available for 1 of these 3 astronauts, and the cine clip showed upward shift of the brain. CONCLUSIONS: Narrowing of the central sulcus, upward shift of the brain, and narrowing of CSF spaces at the vertex occurred frequently and predominantly in astronauts after long-duration flights. Further investigation, including repeated postflight imaging conducted after some time on Earth is required to determine the duration and clinical significance of these change.