TiO2 nanotubes (TiO2-NTs) are currently attracting a high interest because the intrinsic properties of TiO2 provide the basis for many outstanding functional features. Herein, we focus on the cytotoxicity and sublocation of TiO2-NTs in neural stem cells (NSCs). The cytotoxicity of TiO2-NTs is investigated using the methyl tetrazolium cytotoxicity and reactive oxygen species assay, the apoptosis assay by flow cytometry. Cell viability assay shows that TiO2-NTs inside cells are nontoxic at the low concentration. A time-dependent relationship is observed, while a dose-dependent relationship is seen only at the concentration higher than 150 mu g/ml. The uptake happens shortly after incubation with cells. TiO2-NTs can easily pass through the cell membrane and enter into the cells. The uptake amount is increased with prolonging incubation time and reach to maximum at 48 h. Transmission electron microscopy and confocal is used to study subcellular location of TiO2-NTs. It is found that TiO2-NTs traversed cell membrane and localized in many vesicles (endosomes and lysosomes) and cytoplasm. TiO2-NTs in NSCs firstly disperse or metabolism by lysosomal enzymes and then exocytosis from NSCs.