Fig 2: Colocalization of NRXN1 immunoreactivity (red) with phenotypic markers (green) by immunofluorescence. At 8 PCW NRXN1 immunoreactivity was present in cytoplasm/membranes around PAX6-positive radial glia, particularly near the apical ventricular surface (A) but showed little colocalization with SYP in the pSP at either 8 PCW (B) or 12 PCW (D) nor with GAP43 in growing axons of the IZ (C). However, there was strong colocalization (orange–yellow) with TBR1 in the postmitotic neurons of the CP. Scale bars = 100 µm.

Fig 3: Cells generated in the VP belong to different subpopulations of both pallial and subpallial origin. Immunohistochemistry (IHC) of telencephalic coronal sections from a representative cultured embryo (E11.5 + 24 h in culture) with antibodies to different markers: (A–D) pallial marker Tbr1, (E–H) subpallial marker Calbindin and (I–L) Reelin, marker of CR cells. (M,N) Proportion of CFDA+ cells at E10.5 + 24 h in culture (M) and E11.5 + 24 h in culture (N) stages. At E10.5 + 24 h in culture, approximately 3.6% of CFDA-labeled cells expressed Tbr1, 3.1% expressed Calbindin, and 28.7% expressed Reelin (M) In E11.5 + 24 h in culture, 6% of labeled cells expressed Tbr1, 2.8% expressed Calbindin and 26.4% expressed Reelin (N). Scale bar: 50 μm.

Fig 4: Formation and validation of the human cortical organoid under hypoxic conditions. (A) Representative bright-field pictures of the cortical organoids generated on the chip within the hypoxic environment. Scale bars, 500 μm. (B) The immunofluorescence images of NPCs (SOX2 and NESTIN), proliferative marker Ki67, cortical layer (TBR1, SATB2, and CTIP2), differentiated neurons (TUJ1), postsynapse (αPSD95 and drebrin), Wnt signaling pathway (TCF7L2), and inhibitory neurons (GABA) within 40-d cortical organoids. Scale bars, 100 μm (in “Merge”) and 50 μm (in “Enlarged”).

Fig 5: miR-210 Promotes Premature Differentiation in Neural Progenitors(A) Illustration of in utero electroporation timeline for tissue collection. Electroporation was performed at E13.5 and tissue analyzed at E17.5.(B–E) Overexpression of miR-210 increased the early-born neuronal marker TBR1 (B and C) and decreased the late-born neuronal marker SATB2 (D and E) in GFP-positive cells. Sp-210 decreased TBR1 expression in GFP-positive cells (C).VZ, ventricular zone; SVZ, subventricular zone; IZ, intermediate zone; CP, cortical plate.Scale bar, 20 μm. Data are presented as means ± SEM; n = 5 individual brains for all constructs; p values in relation to control (∗p < 0.05, ∗∗p < 0.01). N.S., not significant.