Intravital imaging identifies long-term self-renewing stem cells in the adult mouse hippocampus

Neural stem cells (NSCs) generate new neurons throughout life in the mammalian hippocampus. However, the potential for long-term self-renewal of individual NSCs within the adult brain remains unclear. We used chronic in vivo 2-photon microscopy and followed single NSCs that were genetically labeled through conditional recombination driven by the regulatory elements of the stem cell-expressed genes GLI Family Zinc Finger 1 (Gli1) or Achaete-scute homolog 1 (Ascl1). Through intravital imaging of NSCs and their progeny we identify a population of Gli1-targeted NSCs showing long-term self-renewal in the adult hippocampus. In contrast, once activated, Ascl1-targeted NSCs undergo limited proliferative activity before they becoming exhausted. Using protein expression profiling and single-cell RNA sequencing (scRNA-seq), we show that Gli1- and Ascl1-targeted cells have highly similar yet distinct transcriptional profiles, supporting the existence of heterogeneous NSC populations with diverse behavioral properties. Thus, we here provide the cellular framework for how functional diversity of NSCs enables the generation of new neurons in the adult hippocampus. Overall design: Comparison between Gli1- and Ascl1-targeted tdTomato+ cells. 5 days or 12 weeks after tamoxifen injection, tdTomato+ cells were isolated from the dentate gyrus of 3 mice per group per timepoint. Single cells were FACS sorted into 384 -well plates. Seven 384-well plates were processed according to the SmartSeq2 protocol using a microliter pipetting robot and sequenced on an Illumina HiSeq2500 or HiSeq4000.