The central nervous system is a dense, layered, 3D interconnected network of populations of neurons,
and thus recapitulating that complexity for in vitro CNS models requires methods that can create
defined topologically-complex neuronal networks. Several three-dimensional patterning approaches
have been developed but none have demonstrated the ability to control the connections between
populations of neurons. Here we report a method using AC electrokinetic forces that can guide,
accelerate, slow down and push up neurites in un-modified collagen scaffolds. We present a means to
create in vitro neural networks of arbitrary complexity by using such forces to create 3D intersections
of primary neuronal populations that are plated in a 2D plane. We report for the first time in vitro basic
brain motifs that have been previously observed in vivo and show that their functional network is highly
decorrelated to their structure. This platform can provide building blocks to reproduce in vitro the
complexity of neural circuits and provide a minimalistic environment to study the structure-function
relationship of the brain circuitry.

T. Honegger, M. I. Thielen, S. Feizi, N. E. Sanjana, J. Voldman, Sci. Rep. 2016, 6, 28384.

Full text here