There is concern that degradation of vision as a result of space flight may compromise both mission goals and long-term quality of life after space travel. The visual disturbances may be due to a combination of intracerebral pressure changes and exposure to ionizing radiation. The retina and the retinal vasculature play important roles in vision, yet have not been studied extensively in relationship to space travel and space radiation. The goal of the current study was to characterize oxidative damage and apoptosis in retinal endothelial cells after whole-body gamma-ray, proton and oxygen (¹⁶O) ion radiation exposure at 0.1 to 1 Gy. Six-month-old male C57Bl/6J mice were whole-body irradiated with 600 MeV/n ¹⁶O ions (0, 0.1, 0.25, 1 Gy), solar particle event (SPE)-like protons (0, 0.1, 0.25, 0.5 Gy) or ⁶⁰Co gamma rays (0, 0.1, 0.25, 0.5 Gy). Eyes were isolated for examining endothelial nitric oxide synthase (eNOS) expression and characterization of apoptosis in retina and retinal endothelial cells at two weeks postirradiation. The expression of eNOS was significantly increased in the retina after proton and ¹⁶O ion exposure. ¹⁶O ions induced over twofold increase in eNOS expression compared to proton exposure at two weeks postirradiation (P < 0.05). TUNEL assays showed dose-dependent increases in apoptosis in the retina after irradiation. Low doses of ¹⁶O ions elicited apoptosis in the mouse retinal endothelial cells with the most robust changes observed after 0.1 Gy irradiation (P < 0.05) compared to controls. Data also showed that ¹⁶O ions induced a higher frequency of apoptosis in retinal endothelial cells compared to protons (P < 0.05). In summary, our study revealed that exposure to low-dose ionizing radiation induced oxidative damage and apoptosis in the retina. Significant changes in retinal endothelial cells occur at doses as low as 0.1 Gy. There were significant differences in the responses of endothelial cells among the radiation types examined here.