Unidirectional diphenylalanine nanotubes for dynamically guiding neurite outgrowth

Neural networks have been culturedin vitroto investigate brain functions and diseases, clinical treatments for brain damage, and device development. However, it remains challenging to form complex neural network structures with desired orientations and connectionsin vitro. Here, we introduce a dynamic strategy by using diphenylalanine (FF) nanotubes for controlling physical patterns on a substrate to regulate neurite-growth orientation in cultivating neural networks. Parallel FF nanotube patterns guide neurons to develop neurites through the unidirectional FF nanotubes while restricting their polarization direction. Subsequently, the FF nanotubes disassemble and the restriction of neurites disappear, and secondary neurite development of the neural network occurs in other direction. Experiments were conducted that use the hippocampal neurons, and the results demonstrated that the cultured neural networks by using the proposed dynamic approach can form a significant cross-connected structure with substantially more lateral neural connections than static substrates. The proposed dynamic approach for neurite outgrowing enables the construction of oriented innervation and cross-connected neural networksin vitroand may explore the way for the bio-fabrication of highly complex structures in tissue engineering.