Background: Emerging evidence highlighted vascular injury in aggravating radiation-induced brain injury (RIBI), a common complication of radiotherapy. This study aimed to delineate the pathological feature of cerebral small vessel and investigate the functional roles of Notch signaling in RIBI. Methods: Brain tissue and functional MRI from RIBI patients were collected and analyzed for radiation-induced vasculopathy. A RIBI mouse model was induced by a single dose of 30-Gy cranial irradiation. Vascular morphology, pulsatility, and reactivity to pharmacological interventions, such as nimodipine and 20-hydroxyeicosa-6(Z),15(Z)-dienoic acid, were monitored by 2-photon imaging in mice at 6 weeks postirradiation. Western blot, real-time quantitative PCR, immunofluorescence staining, and behavioral tests were performed. The effect of N-[N-(3, 5-difluorophenacetyl)-l-alanyl]-s-phenylglycinet-butyl ester, a Notch inhibitor, was used to investigate the vascular pathogenesis of RIBI mouse model. Results: Morphologically, radiation resulted in vascular malformation featured by focal contractile rings together with general stenosis. Functionally, radiation also led to hypoperfusion, attenuated vascular pulsatility, and decreased dilation to nimodipine and 20-hydroxyeicosa-6(Z),15(Z)-dienoic acid. Mechanically, Notch activation and increased expression of α-SMA protein were found in both surgical specimens of RIBI patients and the irradiated mice. Importantly, Notch inhibition by N-[N-(3, 5-difluorophenacetyl)-l-alanyl]-s-phenylglycinet-butyl ester significantly alleviated cerebral hypoperfusion, vasculopathy, and cognitive deficits in the RIBI mouse model. Conclusions: Radiation-induced cerebral vasculopathy showed bead-like shape and increased contractile state. Inhibition of Notch signaling by N-[N-(3, 5-difluorophenacetyl)-l-alanyl]-s-phenylglycinet-butyl ester effectively attenuated vasculopathy and relieved cognitive impairment, suggesting Notch signaling as a therapeutic target for the treatment of RIBI.