| Summary: | 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.
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