Integrated single-nucleus transcriptomics, chromatin accessibility profiling, spatial transcriptomics, and live imaging reveal that human ganglionic eminences contain molecularly distinct progenitor populations including outer radial glia-like cells (GE-oRG) that undergo mitotic somal translocation, organize into a trabecular architecture with periphery-to-center differentiation gradients in the MGE that form doublecortin-enriched cellular nests (DENs), and include a spatially segregated CRABP1+/ANGPT2+ domain that contributes to both cortical and striatal interneurons, with PCDH19 serving as a key regulator of primate-specific DEN formation.

The embryonic medial and lateral ganglionic eminences (MGE, LGE) are the principal sources of most neurons and glia for the basal ganglia. In primates, the MGE has a distinctive cytoarchitecture characterized by doublecortin enriched cellular nests (DENs), yet the architectonic organization underlying DEN formation, the molecular heterogeneity of ganglionic eminence progenitors and their lineage relationships, remain poorly understood. Here, using paired single-nucleus transcriptomics and chromatin accessibility profiling of the three GEs, we identify distinct progenitor populations, delineate their gene regulatory networks, and reconstruct their lineage trajectories. Live imaging reveals a unipolar outer radial glia-like population (GE-oRG) that undergoes mitotic somal translocation. Spatial transcriptomics identifies a distinct CRABP1+/ANGPT2+ domain within the MGE. Integrated spatial and electron microscopy demonstrates a periphery-to-center gradient of differentiation in the MGE. Leveraging DEN-forming MGE organoids derived from PCDH19 knockout human pluripotent stem cell lines, we identify the protocadherin, PCDH19, as a key regulator of DEN formation.