Peripherally derived macrophages infiltrate the brain after bone marrow transplantation and during central nervous system (CNS) inflammation. It was initially suggested that these engrafting cells were newly derived microglia and that irradiation was essential for engraftment to occur. However, it remains unclear whether brain-engrafting macrophages (beMϕs) acquire a unique phenotype in the brain, whether long-term engraftment may occur without irradiation, and whether brain function is affected by the engrafted cells. In this study, we demonstrate that chronic, partial microglia depletion is sufficient for beMϕs to populate the niche and that the presence of beMϕs does not alter behavior. Furthermore, beMϕs maintain a unique functional and transcriptional identity as compared with microglia. Overall, this study establishes beMϕs as a unique CNS cell type and demonstrates that therapeutic engraftment of beMϕs may be possible with irradiation-free conditioning regimens.