To characterize the differentiating neurons for known markers of the pallium, sub-pallium, GE (including CGE, MGE and LGE) and cortical layers, RNA expression levels from days 5, 8, 18 and 49 (direct contact) of differentiation were analyzed. During maturation, neuronal precursors showed a gradual increase in the expression of most ganglionic eminence markers tested (Fig 2A–2C). CGE progenitor marker CoupTF2 [33] showed a slight increase during early differentiation (day 8) and CGE marker PROX1 [34] showed increased expression between day 5 and day 18 (Fig 2A). Although expression of MGE marker NKX2.1 was not induced, other MGE markers LHX6 and SATB1, respectively involved in migration of neurons from the ventral telencephalon [35] and in enhancing terminal differentiation of parvalbumin (PV) and somatostatin (SST) interneurons [36], were increasingly expressed at day 18 and 49 of differentiation (Fig 2B). Expression of LGE progenitor markers MEIS2 [37] and GSX2, which regulate LGE progenitor maturation [38] showed increases in expression at day 49 and day 18, respectively (Fig 2C). The sub-pallium marker ASCL1 stayed at a constant expression level whereas the pre-synaptic proteins VGAT and VGLUT1 showed an increased expression over time (Fig 2D).
