Abstract
Background/Objectives: Adult neural stem cells retain the capacity to generate immature neuronal lineages; however, pharmacological approaches that robustly enhance neurogenic activity remain limited. To identify compounds with intrinsic activity under physiologically restrictive conditions, we aimed to screen for small molecules that promote neural stem cell proliferation in the absence of exogenous growth factors and are compatible with central nervous system drug discovery. Methods: We developed a human neural stem cell–based phenotypic screening cascade performed under growth factor–free conditions. Compound activity was evaluated in vitro by ATP-based proliferation assays, BrdU incorporation, and assessment of neurogenic marker analysis. In vivo neurogenic effects were assessed in adult rats by BrdU labeling and immunohistochemical analysis of BrdU/Nestin- and BrdU/DCX-positive cells in the subventricular zone and hippocampal subgranular zone, together with pharmacokinetic analysis to assess brain exposure. Results: Using this platform, we identified Lead-238 as a small-molecule that enhanced neural stem cell proliferation and neurogenic output in vitro. In vivo, Lead-238 increased neurogenic activity in the subventricular zone, as evidenced by elevated numbers of BrdU-positive, BrdU/DCX-positive, and BrdU/Nestin-positive cells, whereas no detectable effects were observed in the hippocampal subgranular zone. Lead-238 achieved sufficient brain exposure, and its neurogenic effects were not readily explained by established neurogenic pathways. Conclusions: These findings demonstrate that growth factor–free phenotypic screening using human neural stem cells is an effective approach for identifying compounds that enhance adult neurogenic activity and identify Lead-238 as a small molecule that increases neurogenic activity in the subventricular zone without detectable effects in the hippocampal subgranular zone.