The Drosophila central nervous system (CNS) develops from a heterogeneous population of neural stem cells (neuroblasts), yet the genes regulating neuroblast determination remain unknown. The segmentation gene wingless is regionally expressed in the neuroectoderm from which neuroblasts develop. A conditional wingless mutation is used to inactivate CNS function without affecting segmentation. The stripe of wingless-expressing neuroectoderm generates apparently normal neuroblasts after wingless inactivation; however, adjacent anterior and posterior neuroectoderm requires wingless nonautonomously for subsequent neuroblast determination and formation. Loss of wingless results in the absence or duplication of identified neuroblasts, highlighting its role in generating neuroblast diversity in the CNS.