In utero electroporation uncovers a distinct population of early dentate gyrus progenitors

The dentate gyrus of the hippocampus develops through complex cellular migrations and differentiations, which have been primarily characterized using genetic lineage tracing approaches. Through systematic application of in utero electroporation across developmental stages, we found that labeling was most effective at embryonic day 12.5 (E12.5), as earlier stages resulted in embryonic lethality while later stages showed markedly reduced efficiency. To directly compare these cells with genetically-defined progenitor populations, we established a novel dual-visualization system, combining electroporation with transgenic reporter mice ( Gfap -GFP). This approach revealed striking differences between two distinct populations: Gfap -GFP+ cells maintain undifferentiated neural stem/progenitor characteristics with persistent Sox2 expression, while E12.5-labeled cells predominantly differentiate into Prox1-positive granule cells by E18.5. These early-labeled cells display characteristic migration patterns, exclusively following an outside-in trajectory to establish the initial framework of the granule cell layer, without reaching the tertiary dentate matrix. In contrast, Gfap -GFP+ cells populate the tertiary dentate matrix and serve as a sustained progenitor reservoir. Molecular marker analysis reveals sequential expression of Sox2, Tbr2, and Prox1, demonstrating progressive differentiation during migration. Our direct comparison identifies a functionally distinct subset of early progenitors that rapidly differentiate, revealing previously unrecognized temporal and functional heterogeneity in dentate development. This study demonstrates how stage-specific in utero electroporation uncovers diverse progenitor populations potentially underrepresented by existing genetic approaches, providing new insights into the cellular diversity that shapes hippocampal structure and function. Clinical trial number: not applicable