This paper describes the effect of geometrical changes in tube-shaped three-dimensional (3D) microenvironment on mouse neural stem cells (mNSCs) differentiating rate and neuronal alignment. We successfully fabricate three types of mNSC microtube structures by controlling the flow rate through double coaxial microfluidic device. Interestingly, we found that thin core diameter induces high neural differentiating rate and increases neuronal alignment in the 3D microenvironment. These results are significantly important information for 3D neuronal tissues formation from neural stem cells. We believe the neuronal alignment in the 3D microenvironment advances the applicability of the tube-shaped structure to tissue engineering, medical transplantation and stem cell biology.