hiPSC-derived neurons had on average larger currents than hESC-derived neurons, suggestive of a higher maturation state, as reported previously [49]. When spontaneous calcium transients were measured, few ROIs increased activity upon TTX wash-out, likely due to a rebound effect of this compound [50]. Bicuculline administration changed calcium activity indicating functional neurons, as GABAergic neurons are excitatory during development. Kuijlaars et al.[51] recently presented protocols of hiPSC-derived cortical neuronal cells in co-culture with primary human astrocytes. Interestingly, they found increased maturation of co-culture systems around day 80 compared to neuronal monocultures by studying synchronized calcium influx activity. This confirms that astrocytes are of utmost importance to generate functional neuronal networks. Further studies should show when our co-culture systems show synchronized activity. Overall, our immunocytochemical and electrophysiological analyses indicate derivation of neuronal networks with mixed glutamatergic and GABAergic neuronal activity.
