Long-term neurological deficiencies resulting from hippocampal cytotoxicity induced by cranial irradiation (IR) present a challenge in the treatment of primary and metastatic brain cancers, especially in children. Previously, we showed that lithium protected hippocampal neurons from IR-induced apoptosis and improved neurocognitive function in treated mice. Here, we demonstrate accelerated repair of IR-induced chromosomal double-strand breaks (DSBs) in lithium-treated neurons. Lithium treatment not only increased IR-induced DNA-dependent protein kinase (DNA-PK) threonine 2609 foci, a surrogate marker for activated nonhomologous end-joining (NHEJ) repair, but also enhanced double-strand DNA end-rejoining activity in hippocampal neurons. The increased NHEJ repair coincided with reduced numbers of IR-induced gamma-H2AX foci, well-characterized in situ markers of DSBs. These findings were confirmed in vivo in irradiated mice. Consistent with a role of NHEJ repair in lithium-mediated neuroprotection, attenuation of IR-induced apoptosis of hippocampal neurons by lithium was dramatically abrogated when DNA-PK function was abolished genetically in SCID mice or inhibited biochemically by the DNA-PK inhibitor IC86621. Importantly, none of these findings were evident in glioma cancer cells. These results support our hypothesis that lithium protects hippocampal neurons by promoting the NHEJ repair-mediated DNA repair pathway and warrant future investigation of lithium-mediated neuroprotection during cranial IR, especially in the pediatric population.