Structures and properties of seven isoforms of the NMDA receptor generated by alternative splicing

doi:10.1016/0006-291X(92)91701-Q

Biochemical and Biophysical Research Communications

Volume 185, Issue 3, 30 June 1992, Pages 826-832

https://doi.org/10.1016/0006-291X(92)91701-Q Get rights and content

Abstract

We here report the existence of 6 additional isoforms of the NMDA receptor generated via alternative splicing by molecular analysis of cDNA clones isolated from a rat forebrain cDNA library. These isoforms possess the structures with an insertion at the extracellular amino-terminal region of deletions at two different extracellular carboxylterminal regions, or those formed by combinations of the above insertion and deletions. One of the deletions results in the generation of a new carboxyl-terminal sequence. All these isoforms possess the ability to induce electrophysiological responses to NMDA and respond to various antagonists selective to the NMDA receptor in the Xenopus oocyte expression system. In addition, a truncated form of the NMDA receptor also exists that contains only the extreme amino-terminal sequence of this protein molecule. These data indicate that the NMDA receptor consists of heterogeneous molecules that differ in the extracellular sequence of the amino- and carboxyl-terminal regions.

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This article is part of the special issue on ‘Glutamate Receptors - NMDA Receptors'.
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NMDA receptors are ionotropic glutamate receptors that mediate excitatory neurotransmission. The diverse functions of these receptors are tuned by deploying different combinations of GluN1 and GluN2 subunits (GluN2A-D) to form either diheteromeric NMDA receptors, which contain two GluN1 and two identical GluN2 subunits, or triheteromeric NMDA receptors, which contain two GluN1 and two distinct GluN2 subunits. Here, we characterize PTC-174, a novel positive allosteric modulator (PAM) of receptors containing GluN2C or GluN2D subunits. PTC-174 potentiates maximal current amplitudes by 1.8-fold for diheteromeric GluN1/2B receptors and by > 10-fold for GluN1/2C and GluN1/2D receptors. PTC-174 also potentiates responses from triheteromeric GluN1/2B/2D and GluN1/2A/2C receptors by 4.5-fold and 1.7-fold, respectively. By contrast, PTC-174 produces partial inhibition of responses from diheteromeric GluN1/2A and triheteromeric GluN1/2A/2B receptors. PTC-174 increases potencies of co-agonists glutamate and glycine by 2- to 5-fold at GluN1/2C and GluN1/2D receptors, and NMDA receptor activation facilitates allosteric modulation by PTC-174. At native NMDA receptors in GluN2D-expressing subthalamic nucleus neurons, PTC-174 increases the amplitude of responses to NMDA application and slows the decay of excitatory postsynaptic currents (EPSCs) evoked by internal capsule stimulation. Furthermore, PTC-174 increases the amplitude and slows the decay of EPSCs in hippocampal interneurons, but has not effect on the amplitudes of NMDA receptor-mediated EPSCs in hippocampal CA1 pyramidal neurons. Thus, PTC-174 provides a useful new pharmacological tool to investigate the molecular pharmacology and physiology of GluN2C- and GluN2D-containing NMDA receptors.
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NMDA receptor (NMDAR) antagonists such as ketamine, can reproduce many of the symptoms of schizophrenia. A reliable indicator of NMDAR channel blocker action in vivo is the augmentation of neuronal oscillation power. Since the coordinated and rhythmic activation of neuronal assemblies (oscillations) is necessary for perception, cognition and working memory, their disruption (inappropriate augmentation or inhibition of oscillatory power or inter-regional coherence) both in psychiatric conditions and with NMDAR antagonists may reflect the underlying defects causing schizophrenia symptoms. NMDAR antagonists and knockout (KO) mice were used to evaluate the role of GluN2C and GluN2D NMDAR subunits in generating NMDAR antagonist-induced oscillations. We find that basal oscillatory power was elevated in GluN2C-KO mice, especially in the low gamma frequencies while there was no statistically significant difference in basal oscillations between WT and GluN2D-KO mice. Compared to wildtype (WT) mice, NMDAR channel blockers caused a greater increase in oscillatory power in GluN2C-KO mice and were relatively ineffective in inducing oscillations in GluN2D-KO mice. In contrast, preferential blockade of GluN2A- and GluN2B-containing receptors induced oscillations that did not appear to be changed in either KO animal. We propose a model wherein NMDARs containing GluN2C in astrocytes and GluN2D in interneurons serve to detect local cortical excitatory synaptic activity and provide excitatory and inhibitory feedback, respectively, to local populations of postsynaptic excitatory neurons and thereby bidirectionally modulate oscillatory power.
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Citation Excerpt :
In addition to variations in LTP, differences in neurotransmitter activity, particularly the glutamatergic (Nakanishi, 1994; Swanson et al., 2005; Gos et al., 2009) and GABAergic (Saba et al., 2011) systems require investigation to understand brain function. A variety of NMDA receptor subunits have been identified: the ubiquitously expressed NR1 subunit; a family of four distinct NR2 subunits (A, B, C, and D); and two NR3 subunits (Moriyoshi et al., 1991; Sugihara et al., 1992; Das et al., 1998). All NMDARs appear to function as heteromeric assemblies composed of multiple NR subunits (Das et al., 1998).
FKBP5 (FKBP51) is a glucocorticoid receptor (GR) binding protein, which acts as a co-chaperone of heat shock protein 90 (HSP90) and negatively regulates GR. Its association with mental disorders has been identified, but its function in disease development is largely unknown. Long-term potentiation (LTP) is a functional measurement of neuronal connection and communication, and is considered one of the major cellular mechanisms that underlies learning and memory, and is disrupted in many mental diseases. In this study, a reduction in LTP in Fkbp5 knockout (KO) mice was observed when compared to WT mice, which correlated with changes to the glutamatergic and GABAergic signaling pathways. The frequency of mEPSCs was decreased in KO hippocampus, indicating a decrease in excitatory synaptic activity. While no differences were found in levels of glutamate between KO and WT, a reduction was observed in the expression of excitatory glutamate receptors (NMDAR1, NMDAR2B and AMPAR), which initiate and maintain LTP. The expression of the inhibitory neurotransmitter GABA was found to be enhanced in Fkbp5 KO hippocampus. Further investigation suggested that increased expression of GAD65, but not GAD67, accounted for this increase. Additionally, a functional GABAergic alteration was observed in the form of increased mIPSC frequency in the KO hippocampus, indicating an increase in presynaptic GABA release. Our findings uncover a novel role for Fkbp5 in neuronal synaptic plasticity and highlight the value of Fkbp5 KO as a model for studying its role in neurological function and disease development.
Mechanism and properties of positive allosteric modulation of N-methyl-D-aspartate receptors by 6-alkyl 2-naphthoic acid derivatives
2017, Neuropharmacology
The theory that N-methyl-d-aspartate receptor (NMDAR) hypofunction is responsible for the symptoms of schizophrenia is well supported by many pharmacological and genetic studies. Accordingly, positive allosteric modulators (PAMs) that augment NMDAR signaling may be useful for treating schizophrenia. Previously we have identified several NMDAR PAMs containing a carboxylic acid attached to naphthalene, phenanthrene, or coumarin ring systems. In this study, we describe several functional and mechanistic properties of UBP684, a 2-naphthoic acid derivative, which robustly potentiates agonist responses at each of the four GluN1a/GluN2 receptors and at neuronal NMDARs. UBP684 increases the maximal l-glutamate/glycine response while having minor subunit-specific effects on agonist potency. PAM binding is independent of agonist binding, and PAM activity is independent of membrane voltage, redox state, and the GluN1 exon 5 N-terminal insert. UBP684 activity is, however, markedly pH-dependent, with greater potentiation occurring at lower pHs and inhibitory activity at pH 8.4. UBP684 increases channel open probability (P_o) and slows receptor deactivation time upon removal of l-glutamate, but not glycine. The structurally related PAM, UBP753, reproduced most of these findings, but did not prolong agonist removal deactivation time. Studies using cysteine mutants to lock the GluN1 and GluN2 ligand-binding domains (LBDs) in the agonist-bound states indicate that PAM potentiation requires GluN2 LBD conformational flexibility. Together, these findings suggest that UBP684 and UBP753 stabilize the GluN2 LBD in an active conformation and thereby increase P_o. Thus, UBP684 and UBP753 may serve as lead compounds for developing agents to enhance NMDAR activity in disorders associated with NMDAR hypofunction.

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Structures and properties of seven isoforms of the NMDA receptor generated by alternative splicing

Abstract

Trends Pharmacol. Sci

Trends Pharmacol. Sci

J. Biol. Chem

Neuron

Neuron

Cell

Annu. Rev. Pharmacol. Toxicol