Ionotropic glutamate receptor triggered cell signaling in Schwann cells

We previously demonstrated that rat Schwann cells (SCs) express the N-methyl-D-aspartate receptor (NMDA-R). In this study, we performed PCR to identify specific subunits of ionotropic glutamate receptors in cultured rat SCs. The NMDA-R-associated subunits: NR1, NR2B, NR2C, NR2D, and NR3B were present at readily detected levels in SCs. We also detected the AMPA Receptor subunits: GluA1, GluA2, and GluA3, and the Kainate Receptor subunits: GluK2, GluK3, GluK4, and GluK5. To test whether SC ionotropic glutamate receptors trigger cell-signaling, cells were exposed to glutamate for 15 min. Protein phosphorylation was assessed using the R&D Systems Phospho-protein Proteome Profiler. Substantial phosphorylation events were noted, including ERK1/2 and c-Jun. The PI3K-Akt pathway was prominently activated, including the downstream kinase, p70 S6 kinase. Activation of Akt may have been supported by mTORC2. The transcription factor, CREB was phosphorylated as well. Proteome Profiler experiments were performed using human SCs and a strikingly similar signaling response to glutamate was noted. Glutamate did not induce SC death at concentrations up to 1.0 mM. In Transwell cell migration experiments, glutamate significantly promoted SC migration (p<0.05). The effects of glutamate on SC migration were enhanced by adding glycine (50 nM). To determine which ionotropic glutamate receptor is responsible for the activity of glutamate in SC signaling, we studied ERK1/2 phosphorylation, as a representative cell-signaling event, by immunoblot analysis. ERK1/2 phosphorylation was substantially inhibited, by more than 50%, by MK801 or by silencing expression of the NR1 subunit of the NMDA-R. MK801 also inhibited SC migration induced by glutamate. These results suggest that the NMDA-R plays a prominent role in eliciting SC responses to glutamate and may participate in SC transdifferentiation that is essential for nerve repair.