No abstract

N-Methyl-D-aspartate (NMDA) receptors are tetrameric protein complexes composed of the glycine-binding NR1 subunit with a glutamate-binding NR2 and/or glycine-binding NR3 subunit. Tri-heteromeric receptors containing NR1, NR2, and NR3 subunits reconstitute channels, which differ strikingly in many properties from the respective glycine-and glutamate-gated NR1/NR2 complexes and the NR1/NR3 receptors gated by glycine alone. Therefore, an accurate oligomerization process of the different subunits has to assure proper NMDA receptor assembly, which has been assumed to occur via the oligomerization of homodimers. Indeed, using fluorescence resonance energy transfer analysis of differentially fluorescence-tagged subunits and blue native polyacrylamide gel electrophoresis after metabolic labeling and affinity purification revealed that the NR1 subunit is capable of forming homo-oligomeric aggregates. In contrast, both the NR2 and the NR3 subunits formed homo-and hetero-oligomers only in the presence of the NR1 subunit indicating differential roles of the subunits in NMDA receptor assembly. However, co-expression of the NR3A subunit with an N-terminal domain-deleted NR1 subunit (NR1 ⌬NTD ) abrogating NR1 and NR1-containing hetero-oligomers are readily formed, we assume that heterodimerization of the NR1 with an NR3 or NR2 subunit, which is followed by the subsequent association of two heterodimers, is the key step in determining proper NMDA receptor subunit assembly and stoichiometry.Excitatory neurotransmission in the mammalian brain is mainly mediated by ionotropic glutamate receptors (iGluRs). 4Based on pharmacological studies, iGluRs have been grouped into three distinct subfamilies: (RS)-2-amino-3-(3-hydroxy-5-methyl-4-isoxazolyl)propionic acid (AMPA) receptors, kainate receptors, and N-methyl-D-aspartic acid (NMDA) receptors (1). All iGluR subunits share a common modular design of four distinct regions: (i) an extracellular N-terminal domain (NTD) of about 400 amino acids sharing homology with the bacterial leucine-, isoleucine-, and valine-binding protein, implicated to play a role in iGluR oligomerization and modulation, (ii) an extracellular S1S2 ligand binding domain sharing homology with the bacterial glutamine-binding protein that binds agonists in a Venus-flytrap like mechanism, (iii) a membrane-associated domain composed of four membrane segments forming the ion channel, and (iv) an intracellular C-terminal domain involved in linking the receptor to the membrane scaffold and signal transduction proteins (2).The NMDA subtype of iGluRs is an obligatory hetero-oligomeric membrane protein composed of homologous NR1, NR2, and/or NR3 subunits and plays a key role in brain development, synaptic plasticity, and memory formation (1). Cloning of NMDA receptor subunits revealed that the glycine-binding NR1 subunit is a single gene product occurring in eight splice variants (a-h), whereas the glutamate-binding NR2 and the glycine-binding NR3 subunits are encoded by four (NR2A-NR2D) and two (NR3A and -B) different genes...

The N-methyl-D-aspartate (NMDA) subtype of ionotropic glutamate receptors (iGluRs) is a tetrameric protein composed of homologous NR1 and NR2 subunits, which require the binding of glycine and glutamate, respectively, for efficient channel gating. The extracellular N-terminal domains (NTDs) of iGluR subunits show sequence homology to the bacterial periplasmic leucine/isoleucine/valine binding protein (LIVBP) and have been implicated in iGluR assembly, trafficking, and function. Here, we investigated how deletion of the NR1-and NR2-NTDs affects the expression and function of NMDA receptors. Both proteolytic cleavage of the NR1-NTD from assembled NR1/ NR2 receptors and coexpression of the NTD-deleted NR1 subunit with wild-type or NTD-deleted NR2 subunits resulted in agonist-gated channels that closely resembled wild-type receptors. This indicates that the NTDs of both NMDA receptor subunits are not essential for receptor assembly and function. However, deletion of either the NR1 or the NR2 NTD eliminated high-affinity, allosteric inhibition of agonist-induced currents by Zn 2ϩ and ifenprodil, consistent with the idea that interdomain interactions between these domains are important for allosteric receptor modulation. Furthermore, by replacing the NR2A-NTD with the NR2B NTD, and vice versa, the different glycine affinities of NR1/NR2A and NR1/NR2B receptors were found to be determined by their respective NR2-NTDs. Together, these data show that the NTDs of both the NR1 and NR2 subunits determine allosteric inhibition and glycine potency but are not required for NMDA receptor assembly.

Rationale Oxytocin receptors (Oxtr) are important mediators of social learning and emotion, with bidirectional effects on fear and anxiety. Contrary to the anxiolytic actions of Oxtr in the amygdala, we recently showed that Oxtr in the lateral septum mediate the enhancement of fear conditioning by social defeat in mice. Objectives Using positive social interactions, which impair fear conditioning, here we attempted to delineate whether the role septal Oxtr in fear regulation depends on the valence of the social memory. Methods Pharmacological and genetic manipulations of lateral septal Oxtr were combined with the social buffering of fear paradigm, in which pre-exposure to nonfearful conspecifics reduces subsequent contextual fear conditioning, as revealed by decreased freezing behavior. Results Antagonism and down-regulation of Oxtr in the lateral septum abolished, while oxytocin (Oxt) administration before pre-exposure to nonfearful conspecifics facilitated the decrease of freezing behavior. Conclusions The septal oxytocin system enhances memory of social interactions regardless of their valence, reducing fear after positive and enhancing fear after negative social encounters. These findings explain, at least in part, the seemingly bidirectional role of Oxt in fear regulation.