Identification of Neural Stem Cells from Postnatal Mouse Auditory Cortex In Vitro

Auditory signals are processed in multiple central nervous system structures, including the auditory cortex (AC). Development of stem cell biology provides the opportunity to identify neural stem cells (NSCs) in the central nervous system. However, it is unclear whether NSCs exist in the AC. The aim of this study is to determine the existence of NSCs in the postnatal mouse AC. To accomplish this aim, postnatal mouse AC tissues were dissected and dissociated into singular cells and small cell clumps, which were suspended in the culture medium to observe neurosphere formation. The spheres were examined by quantitative real-time polymerase chain reaction and immunofluorescence to determine expression of NSC genes and proteins. In addition, AC-spheres were cultured in the presence or absence of astrocyte-conditioned medium (ACM) to study neural differentiation. The results show that AC-derived cells were able to proliferate to form neurospheres, which expressed multiple NSC genes and proteins, including SOX2 and NESTIN. AC-derived NSCs (AC-NSCs) differentiated into cells expressing neuronal and glial cell markers. However, the neuronal generation rate is low in the culture medium containing nerve growth factor, similar to 8%. To stimulate neuronal generation, AC-NSCs were cultured in the culture medium containing ACM. In the presence of ACM, similar to 29% AC-NSCs differentiated into cells expressing neuronal marker class III beta-tubulin (TUJ1). It was observed that the length of neurites of AC-NSC-derived neurons in the ACM group was significantly longer than that of the control group. In addition, synaptic protein immunostaining showed significantly higher expression of synaptic proteins in the ACM group. These results suggest that ACM is able to stimulate neuronal differentiation, extension of neurites, and expression of synaptic proteins. Identifying AC-NSCs and determining effects of ACM on NSC differentiation will be important for the auditory research and other neural systems.