The anatomical and temporal expression of N-methyl-D-aspartate receptor (NMDAR) subunit NMDAR3 (NR3) in the mammalian brain using a novel monoclonal anti-NR3 antibody

Abstract

NR3 is a novel, developmentally regulated NMDA receptor (NMDAR) subunit that was previously known as NMDAR-Like (NMDAR-L) or χ-1 subunit. The discovery of NR3 suggested a role for this novel subunit in neurogenesis as expression of NR3 transcripts peaked at the end of the first postnatal week and was reduced to a low level in adulthood. The regulatory role in neural development was further supported by the evidence that mice in which NR3 was ablated by gene targeting showed increased NMDA-induced currents and increased density of dendritic spines in early postnatal cerebrocortical neurons. These data suggest an inhibitory role of NR3 in NMDAR functions and that it may be involved in the development of synaptic elements by modulating NMDAR activity. To date, the localization of NR3 at the protein level is still unknown. To examine the expression of NR3 protein in the brain is essential to explore the regulatory role of NR3 in the context of NMDAR functions. Therefore, a novel monoclonal anti-NR3 antibody was generated and characterized to delineate the anatomical and temporal expression of NR3 protein in the mammalian brain. The monoclonal anti-NR3 was found to be specific for NR3 in immunocytochemistry, immunoprecipitation and Western blotting. NR3 protein was found to peak at postnatal day 8 (P8), and to decrease gradually from P12 to adulthood in rat central nervous system. NR3 protein was heavily expressed in all areas of the isocortex, a substantial portion of nuclei in the amygdala, in selective cell layers and nuclei in the hippocampal formation, the thalamus, the brainstem, the cerebellar cortex and the spinal cord. The information provided in this study will be useful for the future research of NMDARs, by contributing to the understanding on the role of NR3 and the possible stoichiometry of NMDARs in certain neuronal circuits