Quiescence enables long-term maintenance of adult neural stem cells and exists on a continuum from deep to shallow states, yet the transcriptional programs guiding transitions along this spectrum remain poorly defined. Using single-cell transcriptomics and genetic models, we show that Ascl1 is essential for neural stem cell activation, promoting exit from deep quiescence and inducing Mycn to drive progression through shallow quiescence toward proliferation. In gliomas, quiescent or slow-cycling tumour stem cells are a major source of therapy resistance and recurrence. Using newly developed in vitro models, we show that glioma stem cells share conserved gene expression trajectories with neural stem cells during activation, including potential therapeutic targets. This analysis also reveals that key features of quiescence—such as the "poised" transcriptional state—are conserved in glioma stem cells, uncovering an additional therapeutic vulnerability in this treatment-refractory population.