Glutamate influences neuronal differentiation of neural stem cells from the injured cortex of adult rats

Objective: To explore the influence of glutamate on the neuronal differentiation of neural stem cells (NSCs) from the injured cortex of adult rats. Methods: NSCs were isolated and cultured from the injured cortex after traumatic brain injury (TBI), cells were identified by immunofluorescence staining. NSCs were divided into four groups: control, Glutamate, Glutamate+MK-801 (0 h), Glutamate+MK-801 (24 h). Cells were measured by immunofluorescence, TUNEL assay, real-time PCR, Western blot on 1, 3, 7, 14 day in vitro. Results: NSCs from the injured cortex were BrdU and nestin double-labeled, and could differentiate into MAP-2*. neurons, GFAP(+) astrocytes, and CNP+ oligodendrocytes. Glutamate promoted NSCs to differentiate into DCX+ neuronal progenitor cells (NPCs) at the initial stage; however it subsequently enhanced cellular apoptosis. N-methyl-D-aspartic acid (NMDA) receptor antagonist MK-801 could counteract the effect of glutamate when they were treated together. Interestingly, if MK-801 was added 24 h after glutamate treatment, the number of NPCs was increased and it was higher than that in the Glutamate group on day 3 and day 7 in vitro. On day 14, many MAP-2(+) mature neurons with large cell bodies and abundant processes were observed in the Glutamate+MK-801 (24 h) group. In addition, the expression of NR1 in the Glutamate+MK-801 (24 h) group were gradually increased, peaked on day 7 and then remained at a stable level. Conclusion: Glutamate promotes neuronal differentiation of NSCs at the initial stage, but it also induces neuronal apoptosis during differentiation, which may be related to the gradual increase of NMDA receptor expression during the development of neurons.