Intrinsic mechanisms of neural stem and progenitor cell aging and the age-related decline in adult hippocampal neurogenesis

Adult hippocampal neurogenesis (AHN) continues throughout life but declines with age. Neural stem cells (NSCs) in the dentate gyrus (DG) generate new neurons through tightly regulated intrinsic programs supported by niche cells and systemic factors. Aging reduces AHN by decreasing NSC and neural precursor cell (NPC) numbers and impairing their function. To uncover intrinsic mechanisms driving this decline, we performed single-cell RNA sequencing (scRNA-seq) of murine DG cells from 1–18 months, using wild-type and NSC/NPC-reporter mice to enrich rare precursor populations in aged samples. We also analysed NSC-specific cyclin D2-knockout mice to assess the role of cyclin D2-dependent proliferation in NSC aging. A protocol for extracting and storing high-quality single-cell suspensions compatible with scRNA-seq was first established. A pilot study confirmed that methanol fixation with saline sodium citrate rehydration reliably preserves adult DG cells. Transcriptomic analysis identified eight precursor populations with marked age-related depletion, strongest in active and differentiating cells. Primed and active NSCs showed early aging signatures, while quiescent NSCs remained more stable. Aging down-regulated proliferation, oxidative phosphorylation and protein turnover pathways, indicating deeper quiescence, and up-regulated oxidative stress, apoptosis, senescence and ferroptosis genes, reflecting disrupted energy and redox balance. Early inflammatory signalling appeared in primed NSCs, suggesting contributions from resident and peripheral immune cells in the aging DG. A late shift toward astrocytic differentiation was also observed. Although cyclin D2 loss reduced NSC proliferation, it did not cause age-related NSC depletion, indicating additional mechanisms underlie NSC aging. Overall, these findings reveal intrinsic pathways contributing to precursor aging and AHN decline, providing insight for strategies to maintain neurogenesis and cognitive function in aging.