The development of eye pathology is a serious concern for astronauts who spend time in deep space. Microgravity is a major component of the spaceflight environment which could have adverse effects on ocular health. The use of centrifugation to exert forces that partially or fully mimic Earth-level gravity in space is a possible countermeasure to mitigate the effects of microgravity on the eye. Therefore, we subjected mice on the International Space Station (ISS) to microgravity (0 G) or artificial gravity by centrifugation at 0.33 G, 0.67 G, and 1 G, and then performed RNA sequencing (RNA-seq) on optic nerve and retinal tissue after returning them to Earth alive. We find that the microgravity environment induces transcriptomic changes in the optic nerve and retina consistent with an increased oxidative stress load, inflammation, apoptosis, and lipid metabolic stress. We also find that adding artificial gravity on board the ISS attenuates the transcriptomic response to microgravity in a dose-dependent manner. Such attenuation may effectively protect from and mitigate spaceflight-induced detrimental effects on ocular tissue.