Granulocyte-colony stimulating factor (G-CSF) is a growth factor that regulates proliferation, differentiation and survival of hematopoietic progenitor cells. There is growing evidence to suggest that G-CSF exerts a powerful neuroprotective effect in different neurological disorders. However, it has remained to be elucidated if G-CSF has a direct effect on neural stem cells (NSCs). Here, we show that G-CSF could stimulate the proliferation of NSCs and promote their differentiation in vitro. Additionally, we have shown that G-CSF-induced proliferation of NSCs is associated with phosphorylation of STAT3, and the differentiation is linked to altered expression of differentiation-related genes. Remarkably, G-CSF could not initiate the differentiation of NSCs. The added roles of G-CSF in regulating proliferation and differentiation of NSCs as shown in this study would serve as a useful reference in designing new stem cell therapy strategies for promoting brain recovery and repair.