In the Telencephalon, GluN2C NMDA Receptor Subunit mRNA is Predominately Expressed in Glial Cells and GluN2D mRNA in Interneurons
N-methyl-d-aspartate receptors (NMDARs) are widely distributed in the brain with high concentrations in the telencephalon where they modulate synaptic plasticity, working memory, and other functions. While the actions of the predominate GluN2 NMDAR subunits, GluN2A and GluN2B are relatively well understood, the function of GluN2C and GluN2D subunits in the telencephalon is largely unknown. To better understand the possible role of GluN2C subunits, we used fluorescence in situ hybridization (FISH) together with multiple cell markers to define the distribution and type of cells expressing GluN2C mRNA. Using a GluN2C-KO mouse as a negative control, GluN2C mRNA expression was only found in non-neuronal cells (NeuN-negative cells) in the hippocampus, striatum, amygdala, and cerebral cortex. For these regions, a significant fraction of GFAP-positive cells also expressed GluN2C mRNA. Overall, for the telencephalon, the globus pallidus and olfactory bulb were the only regions where GluN2C was expressed in neurons. In contrast to GluN2C, GluN2D subunit mRNA colocalized with neuronal and not astrocyte markers or GluN2C mRNA in the telencephalon (except for the globus pallidus). GluN2C mRNA did, however, colocalize with GluN2D in the thalamus where neuronal GluN2C expression is found. These findings strongly suggest that GluN2C has a very distinct function in the telencephalon compared to its role in other brain regions and compared to other GluN2-containing NMDARs. NMDARs containing GluN2C may have a specific role in regulating l-glutamate or d-serine release from astrocytes in response to l-glutamate spillover from synaptic activity.
KeywordsN-methyl-d-aspartate receptor l-glutamate Astrocyte Cortex Hippocampus mRNA GluN2C GluN2D
This study was funded by the National Institutes of Health grants MH60252, NS104705, and GM110768 and the Edna Ittner Pediatric Research Foundation. The authors thank Dr. Andres Buonanno and colleagues for having provided the GluN2C-KO mouse line, Dr. Masahiko Watanabe for helpful discussions, Robin Taylor for expert graphical assistance, and Dr. Emily Harrison for technical advice.
- 6.Ishii T, Moriyoshi K, Sugihara H, Sakurada K, Kadotani H, Yokoi M, Akazawa C, Shigemoto R, Mizuno N, Masu M (1993) Molecular characterization of the family of the N-methyl-d-aspartate receptor subunits. J Biol Chem 268:2836–2843Google Scholar
- 9.Zhang Y, Buonanno A, Vertes RP, Hoover WB, Lisman JE (2012) NR2C in the thalamic reticular nucleus; effects of the NR2C knockout. PLoS ONE 7, e41908Google Scholar
- 17.Angerer LM, Angerer RC (1992) In situ hybridization to cellular RNA with radiolabeled RNA probes. In: Wilkinson DG (ed) In situ hybridization: a practical approach. Oxford University Press, Oxford, pp 15–32Google Scholar
- 26.Perszyk RE, DiRaddo JO, Strong KL, Low CM, Ogden KK, Khatri A, Vargish GA, Pelkey KA, Tricoire L, Liotta DC, Smith Y, McBain CJ, Traynelis SF (2016) GluN2D-Containing N-methyl-d-Aspartate Receptors Mediate Synaptic Transmission in Hippocampal Interneurons and Regulate Interneuron Activity. Mol Pharmacol 90:689–702CrossRefGoogle Scholar
- 29.Lozovaya N, Gataullina S, Tsintsadze T, Tsintsadze V, Pallesi-Pocachard E, Minlebaev M, Goriounova NA, Buhler E, Watrin F, Shityakov S, Becker AJ, Bordey A, Milh M, Scavarda D, Bulteau C, Dorfmuller G, Delalande O, Represa A, Cardoso C, Dulac O, Ben-Ari Y, Burnashev N (2014) Selective suppression of excessive GluN2C expression rescues early epilepsy in a tuberous sclerosis murine model. Nat Commun 5:4563CrossRefGoogle Scholar
- 32.Mullen RJ, Buck CR, Smith AM (1992) NeuN, a neuronal specific nuclear protein in vertebrates. Development 116:201–211Google Scholar
- 33.Zhang Y, Chen K, Sloan SA, Bennett ML, Scholze AR, O’Keeffe S, Phatnani HP, Guarnieri P, Caneda C, Ruderisch N, Deng S, Liddelow SA, Zhang C, Daneman R, Maniatis T, Barres BA, Wu JQ (2014) An RNA-sequencing transcriptome and splicing database of glia, neurons, and vascular cells of the cerebral cortex. J Neurosci 34:11929–11947CrossRefGoogle Scholar
- 50.Cahoy JD, Emery B, Kaushal A, Foo LC, Zamanian JL, Christopherson KS, Xing Y, Lubischer JL, Krieg PA, Krupenko SA, Thompson WJ, Barres BA (2008) A transcriptome database for astrocytes, neurons, and oligodendrocytes: a new resource for understanding brain development and function. J Neurosci 28:264–278CrossRefGoogle Scholar
- 52.Costa BM, Feng B, Tsintsadze TS, Morley RM, Irvine MW, Tsintsadze V, Lozovaya NA, Jane DE, Monaghan DT (2009) N-methyl-d-aspartate (NMDA) receptor NR2 subunit selectivity of a series of novel piperazine-2,3-dicarboxylate derivatives: preferential blockade of extrasynaptic NMDA receptors in the rat hippocampal CA3-CA1 synapse. J Pharmacol Exp Ther 331:618–626CrossRefGoogle Scholar
- 55.Krupp JJ, Vissel B, Heinemann SF, Westbrook GL (1996) Calcium-dependent inactivation of recombinant N-methyl-d-aspartate receptors is NR2 subunit specific. Mol Pharmacol 50:1680–1688Google Scholar