Activation of Group II Metabotropic Glutamate Receptors Increases Proliferation but does not Influence Neuronal Differentiation of a Human Neural Stem Cell Line

Abstract

The multiple functions of glutamate include regulation of neural development and stem cells. While the importance of the ionotropic glutamate receptors is well-established, less is known about the role of metabotropic glutamate receptors (mGluRs). In this study, we examined the effects of pharmacological activation and inhibition of mGluR2/3 on proliferation, differentiation and viability of a human neural stem cell line. Immunofluorescence staining revealed the presence of mGluR2/3 receptors on both proliferating and differentiating stem cells, including cells differentiated into β-tubulin III-positive immature neurons and glial fibrillary acidic protein-positive astrocytes. Stimulation of mGluR2/3 receptors during cell propagation using the agonist (2S,2′R,3′R)-2-(2′,3′-dicarboxycyclopropyl) glycine (DCG-IV) increased total cell numbers significantly (60% compared to untreated controls). This effect could be inhibited by the specific antagonist (2S)-2-Amino-2-[(1S,2S)-2-carboxycycloprop-1-yl]-3-(xanth-9-yl) propanoic acid (LY341495). The antagonist alone had no effect. No significant decrease in cell death was found following mGluR2/3 stimulation, suggesting that the observed elevation in cell number was not related to cell viability. Subsequent differentiation of the cells resulted in a slight decrease in β-tubulin III-positive neurons (5.2–3.2% of total cells) for DCG-IV pre-treated cultures. Treatment with DCG-IV and LY342495 during cell differentiation alone had no such effect. Western blot analysis revealed that the active, dimeric form of mGluR2/3 was mainly present on the proliferating cells, which may explain our findings. This study emphasizes the importance of glutamate and mGluRs on regulation of human neural stem cells and suggests a significant role of mGluR2/3 during cell proliferation.