Activation of the leukemia inhibitory factor (LIF) receptor has been reported to promote gliogenesis and also to support neural stem cell (NSC) renewal. To investigate this paradox, we isolated NSCs and generated neurospheres from embryonic mice either wild-type, heterozygous, or homozygous null for LIF receptor (LIFR)-β. Exogenous LIF abrogated neurosphere formation and promoted expression of GFAP by all cells in wild-type and heterozygous cultures. LIF also stimulated a twofold increase in the number of multipotential clones generated from these cultures in comparison with those pretreated with EGF and FGF-2 (E+F) alone. In contrast, the clonogenicity of low-density cultures of LIFR knockout cells was reduced in comparison with that of wild-type cells grown in E+F and was unaffected by LIF. Thus, although LIFR signaling is not necessary for NSC self-renewal, it enhances both the clonogenicity and the expression of GFAP by these multipotential cells.