Injury to the adult mammalian CNS results in reactive changes among the glial cells surrounding the site of damage. Recently, an unusual class of glial cells has been identified within the intact adult rat cerebellum on the basis of the expression of the NG2 chondroitin-sulfate proteoglycan (Levine and Card, 1987). To determine whether the cells that express the NG2 proteoglycan show reactive changes after injury, small puncture lesions were made into the cerebelli of adult rats, and changes among astrocytes, microglia and NG2-positive cells were examined using immunohistochemical staining with cell type-specific marker antibodies. Beginning at 24 hr after lesion, NG2-positive cells immediately adjacent to the lesion site bound the anti-NG2 antibodies more heavily than cells within the undamaged areas of the cerebellum. This increase in anti-NG2 immunoreactivity was transient, reaching a maximum at 7 d postlesion and declining slowly thereafter. The increase in anti-NG2 immunoreactivity was accompanied by an increase in the levels of mRNA encoding the NG2 core protein as demonstrated by in situ hybridization. NG2-positive cells adjacent to the lesion site incorporated 3H-thymidine into their nuclei beginning at 24 hr postlesion and increased in number. Concurrent with these changes, microglia became activated and increased in number, monocytes invaded the damaged tissue, and an astrocytic scar formed. These observations demonstrate that the cells that express the NG2 proteoglycan are a reactive cell type that responds to brain injury. The increased expression of the NG2 chondroitin-sulfate proteoglycan may contribute to the failure of damaged CNS axons to regenerate successfully.