Adult neurogenesis, a process of generating newborn neurons from adult neural stem cells, is required for brain homeostasis, cognition, and affective behaviors. Deciphering the molecular mechanisms underlying adult neurogenesis will provide valuable insights into the functional integrity of the adult brain and the etiology of neurological disorders. Here, we present an optimized protocol combining stereotactic injection of retrovirus expressing red fluorescent protein to label newborn neurons and implantation of a mini-osmotic pump to investigate newborn neuron development in adult mouse hippocampus. For complete details on the use and execution of this protocol, please refer to Tang et al. (2019).