We have previously studied cortical interneuron development using a co-culture system in which mouse embryonic MGE-derived progenitors are plated over a “feeder culture” composed mainly of mouse cortical pyramidal neurons and glia (Xu et al., 2004). To determine whether aspects of human interneuron maturation would be accelerated in this system relative to the xenografts, NKX2.1::GFP+ cells were collected by FACS at day 32 and replated onto cultures of dissociated embryonic mouse cortex (Figure 4A). The cortical cells were isolated from E13.5 mouse embryos, a stage prior to the immigration of the ventrally-derived interneurons into the dorsal neocortex (Anderson et al., 1997). Thus, this co-culture system mimics aspects of normal development with human NKX2.1+ cells developing in the presence of the glutamatergic cortical pyramidal neurons. Studies were performed in parallel using GFP+ cells derived from each of the three SHH treatment regimens (Figure 4B–E). Roughly 80% of the GFP+ cells from the 10-18 treated “MGE-like” cultures, versus 40% from the 2-18 “hypothalamic-like” and 15% from the 6-18 culture, expressed GABA. Likewise, the 6-18 culture that is enriched for telencephalic (FOXG1+) cells but lacks expression of the interneuron precursor marker LHX6 (Figure 3D), was enriched for choline acetyltransferase (ChAT) neurons.
