We examined the differentiation potential of murine neural stem cells (NSCs) grown in vitro and transplanted into intact and irradiated recipients. NSCs were isolated from neonatal Balb/c mice using the neurosphere assay. On in vitro differentiation assays, NSCs produced beta-III tubulin(+) neurons, glial fibrillary acidic protein (GFAP(+)) astrocytes, and O4(+) oligodendrocytes. After neural grafting to histocompatible adult mice, NSCs gave rise to neuronal and glial cells. When cells were transplanted in the form of solid neurospheres, they reached terminal differentiation and spatial arrangements that mimicked the three-dimensional organization of nervous tissue. To create conditions that would allow us to assess the potential for generation of nonneural cells, NSCs were intravenously injected into irradiated mice. Transplantation of NSCs stimulated hematopoiesis because the number of colony-forming units of granulocyte-monocyte lineage (CFU-GM) colonies isolated from the spleen and bone marrow of transplanted mice was greater than that from irradiated, nontransplanted animals. Moreover, transplanted cells tagged with beta-galactosidase were identified in the thymus of animals grafted with labelled NSCs. NSCs harvested from the neurosphere assay produced viable and transplantable cells. In vitro differentiation assays and neural grafting confirmed the multipotency of NSCs and their commitment to generate neuronal and glial cells. Following intravenous injection of NSCs, the transplanted cells colonized hematopoietic and lymphatic organs, facilitating hematopoiesis in irradiated animals.