PROJECT SUMMARY Adult neurogenesis is emerging as an important player in maintaining brain homeostasis and normal functions. The dysfunctions of neurogenesis have been associated with aging and neurological disorders, including Alzheimer’s disease (AD). The ability to systematically map the molecular dynamics of neurogenesis at single- cell resolution could serve as a foundation for a systematic effort to better understand the molecular events that give rise to abnormal cell states in aging and diseases. While the rapid advances in single-cell genomics are creating unprecedented opportunities to explore molecular heterogeneity in mammalian brains, nearly all such methods are restricted to low throughput and fail to recover the heterogeneity and dynamics of the profoundly rare cell states in adult neurogenesis (e.g., less than 0.1% of the cell population in the brain). Herein, we propose to develop novel methodologies that enable a comprehensive view of temporal-spatial dynamics of neurogenesis during aging and Alzheimer's disease (AD) in both human and mouse brains. Specifically, we will first develop a novel high-throughput, low-cost single-cell genomics approach, sciNext1000, to profile the molecular heterogeneity of four million cells from post-mortem human hippocampal samples. This approach will be powerful because we can not only quantitatively characterize the frequency of human adult hippocampal neurogenesis at single-cell resolution, but also identify the transcriptome features associated with impaired neurogenesis in aging and AD at isoform resolution. In addition, we will develop another novel single-cell genomic technique, sci-Div- seq, to enhance the detection of newborn neurons, and identify the cellular differentiation trajectories and associated transcriptomic features of adult neurogenesis in young and aged mouse brains. The resulting dataset will advance our understanding of gene regulation in neurogenesis across different neural lineages and constitute a significant step towards a comprehensive characterization of the molecular mechanism underlying neurogenesis impairment in aging. In addition to the internal molecular programs, the neurogenesis process is controlled by aspects of environmental signals from the neural stem niche. We will apply a high-throughput spatial transcriptomic strategy to identify the cellular interactions and local microenvironment involved in adult neurogenesis in both human and mouse brains. These multi-pronged approaches will open a new paradigm for understanding the global molecular programs and environmental regulation of adult neurogenesis, thereby informing potential therapeutic targets to restore cell population homeostasis in aging and brain disorders.