Prospective identification of functionally distinct stem cells and neurosphere-initiating cells in adult mouse forebrain

Neurosphere formation is commonly used as a surrogate for neural stem cell (NSC) function but the relationship between neurosphere-initiating cells (NICs) and NSCs remains unclear. We prospectively identified, and isolated by flow cytometry, adult mouse lateral ventricle subventricular zone (SVZ) NICs as Glast^midEGFR^highPlexinB2^highCD24-/^lowO4/PSA-NCAM-/lowTer119/CD45- (GEPCOT) cells. They were highly mitotic and short-lived in vivo based on fate-mapping with Ascl1^CreERT2 and Dlx1^CreERT2. In contrast, pre-GEPCOT cells were quiescent, expressed higher Glast, and lower EGFR and PlexinB2. Pre-GEPCOT cells could not form neurospheres but expressed the stem cell markers Slc1a3-CreER^T, GFAP-CreER^T2, Sox2^CreERT2, and Gli1^CreERT2 and were long-lived in vivo. While GEPCOT NICs were ablated by temozolomide, pre-GEPCOT cells survived and repopulated the SVZ. Conditional deletion of the Bmi-1 polycomb protein depleted pre-GEPCOT and GEPCOT cells, though pre-GEPCOT cells were more dependent upon Bmi-1 for Cdkn2a (p16^Ink4a) repression. Our data distinguish quiescent NSCs from NICs and make it possible to study their properties in vivo.