Autofluorescence is a biomarker of neural stem cell activation state

Neural stem cells (NSCs) must exit quiescence to produce neurons; however, our understanding of this process remains constrained by the technical limitations of current technologies. Fluorescence lifetime imaging (FLIM) of autofluorescent metabolic cofactors has been used in other cell types to study shifts in cell states driven by metabolic remodeling that change the optical properties of these endogenous fluorophores. Using this non-destructive, live-cell, and label-free strategy, we found that quiescent NSCs (qNSCs) and activated NSCs (aNSCs) have unique autofluorescence profiles. Specifically, qNSCs display an enrichment of autofluorescence localizing to a subset of lysosomes, which can be used as a graded marker of NSC quiescence to predict cell behavior at single-cell resolution. Coupling autofluorescence imaging with single-cell RNA sequencing, we provide resources revealing transcriptional features linked to deep quiescence and rapid NSC activation. Together, we describe an approach for tracking mouse NSC activation state and expand our understanding of adult neurogenesis. [Display omitted] •Autofluorescence is a live-cell and label-free marker of NSC quiescence•Lysosome-localized autofluorescence is enriched in quiescent NSCs•NSC autofluorescence predicts cell behavior in vitro and ex vivo•Autofluorescence imaging paired with scRNA sequencing can reveal quiescence substates Classical approaches for determining neural stem cell activation state are technically limited, often involving confounding exogenous reagents or sample destruction through lysis or fixation. Overcoming this barrier, Morrow et al. developed a non-destructive, live-cell, and label-free approach for distinguishing quiescent and activated neural stem cells through autofluorescence imaging.