A simplified method for long-term maintenance of human induced pluripotent stem-cell derived neural networks

In-vitro models of neuronal networks have become a powerful tool for modeling network activity in the human brain. The exploration of network properties has largely been made possible via microelectrode arrays (MEAs). However, addressing certain tissue engineering challenges remains imperative for their long-term utilization. Maintaining human neural assemblies on glass MEAs is difficult as cells often clump, peel, and spontaneously detach from the surface. Such difficulties in long-term human neuronal network culture has prompted utilization of rat astrocytes, conditioned media, and genetically modified neurons. These methods may not accurately capture the micro-environment of the brain and require significant expertise, making them less universally applicable across diverse studies. Our method produced four successful human induced pluripotent stem cell-derived neuron-astrocyte co-culture with >90 days viability without xeno sources or genetic modification. The presence of mature neurons was confirmed with live staining. Recordings taken from one culture exhibited neuronal spiking activity at an amplitude of ∼200 µV and evidence of network-wide activity. Synchronized bursting was also observed at a rate of 0.34 - 1.08 Hz in recording data. This demonstrates the feasibility of an easily implementable solution for long-term culturing of active neural networks.