A novel chitosan scaffold with micro- and nano-hybrid structures was proposed in this study. The hemispheric array of the barrier layer of an anodic aluminum oxide (AAO) film was used as the substrate. Microelectromechanical systems and nickel electroforming techniques were integrated for fabricating chitosan scaffolds with different micro/nanohybrid structures. Nerve cells were then cultured on the conduits. It was demonstrated that the scaffold with pure microstructures can guide the nerve cells to grow along the ridges of the microstructure and some cells to grow across the groove in between two ridges of the microstructure. It was also shown that the scaffold with microscale ridges and nanopatterns on the groove between two ridges can more effectively guide the cells to grow along the ridges, thus enhancing the proliferation of nerve cells.