The prefrontal cortex (PFC) is a brain region featured with working memory function. The exact mechanism of how working memory operates within the PFC circuitry is unknown, but persistent neuronal firing recorded from prefrontal neurons during a working memory task is proposed to be the neural correlate of this mnemonic encoding. The PFC appears to be specialized for sustaining persistent firing, with N-methyl-D-aspartate (NMDA) receptors, especially slow-decay NR2B subunits, playing an essential role in the maintenance of sustained activity and normal working memory function. However, the NR2B subunit serves as a double-edged sword for PFC function. Because of its slow kinetics, NR2B endows the PFC with not only “neural psychic” properties, but also susceptibilities for neuroexcitotoxicity and psychiatric disorders. This review aims to clarify the interplay among working memory, the PFC, and NMDA receptors; demonstrate the importance of the NR2B subunit in the maintenance of persistent activity; understand the risks and vulnerabilities of how NR2B is related to the development of neuropsychiatric disorders; identify gaps that currently exist in our understanding of these processes; and provide insights regarding future directions that may clarify these issues. We conclude that the PFC is a specialized brain region with distinct delayed maturation, unique neuronal circuitry, and characteristic NMDA receptor function. The unique properties and development of NMDA receptors, especially enrichment of NR2B subunits, endows the PFC with not only the capability to generate sustained activity for working memory, but also serves as a major vulnerability to environmental insults and risk factors for psychiatric disorders.

Schizophrenia (SCZ) is characterized not only by psychosis, but also by working memory and executive functioning deficiencies, processes that rely on the prefrontal cortex (PFC). Because these cognitive impairments emerge prior to psychosis onset, we investigated synaptic function during development in the neurodevelopmental methylazoxymethanol acetate (MAM) model for SCZ. Specifically, we hypothesize that NMDAR hypofunction is attributable to reductions in the NR2B subunit through aberrant epigenetic regulation of gene expression, resulting in deficient synaptic physiology and PFC-dependent cognitive dysfunction, a hallmark of SCZ. Using Western blot and whole-cell patch-clamp electrophysiology, we found that the levels of synaptic NR2B protein are significantly decreased in juvenile MAM animals, and the function of NMDARs is substantially compromised. Both NMDA-mEPSCs and synaptic NMDA-eEPSCs are significantly reduced in prelimbic PFC (plPFC). This protein loss during the juvenile period is correlated with an aberrant increase in enrichment of the epigenetic transcriptional repressor REST and the repressive histone marker H3K27me3 at the Grin2b promoter, as assayed by ChIP-qPCR. Glutamate hypofunction has been a prominent hypothesis in the understanding of SCZ pathology; however, little attention has been given to the NMDAR system in the developing PFC in models for SCZ. Our work is the first to confirm that NMDAR hypofunction is a feature of early postnatal development, with epigenetic hyper-repression of the Grin2b promoter being a contributing factor. The selective loss of NR2B protein and subsequent synaptic dysfunction weakens plPFC function during development and may underlie early cognitive impairments in SCZ models and patients.

In primates, working memory function depends on activity in a distributed network of cortical areas that display different patterns of delay task-related activity. These differences are correlated with, and might depend on, distinctive properties of the neurons located in each area. For example, layer 3 pyramidal neurons (L3PNs) differ significantly between primary visual and dorsolateral prefrontal (DLPFC) cortices. However, to what extent L3PNs differ between DLPFC and other association cortical areas is less clear. Hence, we compared the properties of L3PNs in monkey DLPFC versus posterior parietal cortex (PPC), a key node in the cortical working memory network. Using patch clamp recordings and biocytin cell filling in acute brain slices, we assessed the physiology and morphology of L3PNs from monkey DLPFC and PPC. The L3PN transcriptome was studied using laser microdissection combined with DNA microarray or quantitative PCR. We found that in both DLPFC and PPC, L3PNs were divided into regular spiking (RS-L3PNs) and bursting (B-L3PNs) physiological subtypes. Whereas regional differences in single-cell excitability were modest, B-L3PNs were rare in PPC (RS-L3PN:B-L3PN, 94:6), but were abundant in DLPFC (50:50), showing greater physiological diversity. Moreover, DLPFC L3PNs display larger and more complex basal dendrites with higher dendritic spine density. Additionally, we found differential expression of hundreds of genes, suggesting a transcriptional basis for the differences in L3PN phenotype between DLPFC and PPC. These data show that the previously observed differences between DLPFC and PPC neuron activity during working memory tasks are associated with diversity in the cellular/molecular properties of L3PNs.Significance statementIn the human and non-human primate neocortex, layer 3 pyramidal neurons (L3PNs) differ significantly between dorsolateral prefrontal (DLPFC) and sensory areas. Hence, L3PN properties reflect, and may contribute to, a greater complexity of computations performed in DLPFC. However, across association cortical areas, L3PN properties are largely unexplored. We studied the physiology, dendrite morphology and transcriptome of L3PNs from macaque monkey DLPFC and posterior parietal cortex (PPC), two key nodes in the cortical working memory network. L3PNs from DLPFC had greater diversity of physiological properties and larger basal dendrites with higher spine density. Moreover, transcriptome analysis suggested a molecular basis for the differences in the physiological and morphological phenotypes of L3PNs from DLPFC and PPC.

Schizophrenia (SCZ) is a neurodevelopmental psychiatric disorder, in which cognitive function becomes disrupted at early stages of the disease. Although the mechanisms underlying cognitive impairments remain unclear, N-methyl-D-aspartate receptors (NMDAR) hypofunctioning in the prefrontal cortex (PFC) has been implicated. Moreover, cognitive symptoms in SCZ are usually unresponsive to treatment with current antipsychotics and by onset, disruption of the dopamine system, not NMDAR hypofunctioning, dominates the symptoms. Therefore, treating cognitive deficits at an early stage is a realistic approach. In this study, we tested whether an early treatment targeting mGluR2 would be effective in ameliorating cognitive impairments in the methylazoxymethanol acetate (MAM) model of SCZ. We investigated the effects of an mGluR2 agonist/mGluR3 antagonist, LY395756 (LY39), on the NMDAR expression and function in juveniles, as well as cognitive deficits in adult rats after juvenile treatment. We found that gestational MAM exposure induced a significant decrease in total protein levels of the NMDAR subunit, NR2B, and a significant increase of pNR2BTyr1472 in the juvenile rat PFC. Treatment with LY39 in juvenile MAM-exposed rats effectively recovered the disrupted NMDAR expression. Furthermore, a subchronic LY39 treatment in juvenile MAM-exposed rats also alleviated the learning deficits and cognitive flexibility impairments when tested with a cross-maze based set-shifting task in adults. Therefore, our study demonstrates that targeting dysfunctional NMDARs with an mGluR2 agonist during the early stage of SCZ could be an effective strategy in preventing the development and progression in addition to ameliorating cognitive impairments of SCZ.

Targeting group II metabotropic glutamate receptors (mGluR2/3) has been proposed to correct the dysfunctional glutamatergic system, particularly NMDA receptor (NMDAR) hypofunction, for treatment of schizophrenia. However, how activation of mGluR2/3 affects NMDAR function in adult animals remains elusive. Here we show the effects of LY395756 (LY39), a compound acting as both an mGluR2 agonist and mGluR3 antagonist, on the NMDAR expression and function of normal adult rat prefrontal cortex (PFC) as well as working memory function in the MK801 model of schizophrenia. We found that in vivo administration of LY39 significantly increased the total protein levels of NMDAR subunits and NR2B phosphorylation in the PFC, along with the amplitude of NMDAR-mediated miniature excitatory postsynaptic currents (mEPSC) in the prefrontal cortical neurons. Moreover, LY39 also significantly increased mTOR and pmTOR expression, but not ERK1/2, Akt, and GSK3β, suggesting an activation of mTOR signaling. Indeed, the mTOR inhibitor rapamycin, and actinomycin-D, a transcription inhibitor, blocked the enhanced effects of LY39 on NMDAR-mEPSCs. These results indicate that LY39 regulates NMDAR expression and function through unidentified mTOR-mediated protein synthesis in the normal adult rat PFC. However, this change is insufficient to affect working memory function in normal animals, nor to reverse the MK801-induced working memory deficit. Our data provide the first evidence of an in vivo effect of a novel compound that acts as both an mGluR2 agonist and mGluR3 antagonist on synaptic NMDAR expression and function in the adult rat PFC, although its effect on PFC-dependent cognitive function remains to be explored.