Monolayer Differentiation

Stem cells will be plated at high density and treated with dual SMAD inhibitors (Dorsomorphin and SB-431542) for 14 days in 3N media essentially as described (Shi, Y., et al., Human Cerebral Cortex Development from Pluripotent Stem Cells to Functional Excitatory Synapses. Nature Neurosci., 2012. 15(3): p. 477-486). Cells will be passed and cultured for 5-7 days in 3N media supplemented with FGF to form neural rosettes. Rosettes will be manually isolated and plated in 3N media for neural differentiation and cultured either as rosettes or dispersed neural progenitor cells for an additional 3 weeks. At this point the cultures will contain neurons, glia and undifferentiated neuronal progenitor cells.