Compartmentalized microfluidic chambers enable long-term maintenance and communication between human pluripotent stem cell-derived forebrain and midbrain neurons

Compartmentalized microfluidic devices are becoming increasingly popular and have proven to be valuable tools to probe neurobiological functions that are inherently difficult to study using traditional approaches. The ability of microfluidic devices to compartmentalize neurons offers considerable promise for disease modeling and drug discovery. Rodent cortical neurons/neural progenitors are commonly used in such studies but, while these cells mature rapidly, they do not possess the same receptors, ion channels and transport proteins found in human cortical neurons. Human pluripotent stem cell derived neurons offer a human phenotype, but their slow maturation offsets this phenotypic advantage, particularly over long-term culture where overgrowth and subsequent death of neurons may be a problem. In this work, we integrate the use of Matrigel as a 3D cell culture scaffold that enables high cell seeding density over a small fraction of the culture surface. This approach, in an open chamber microfluidic system, enables culture over a five-month period without the use of growth inhibitors. Matrigel was also uniquely utilized to hinder agonist diffusion across microchannels. We demonstrate the development of neuron-to-neuron communication networks by showing that electrical stimulation or the unilateral addition of agonists to one chamber resulted in activation of neurons in the adjacent chamber. Lastly, using a delayed neuron seeding strategy, we show that we can foster essentially one-way communication between separate populations of human forebrain and midbrain dopaminergic neuron containing cultures.