XB-ART-56935
Nat Chem Biol
2020 Feb 01;162:188-196. doi: 10.1038/s41589-019-0449-5.
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Biased modulators of NMDA receptors control channel opening and ion selectivity.
Perszyk RE
,
Swanger SA
,
Shelley C
,
Khatri A
,
Fernandez-Cuervo G
,
Epplin MP
,
Zhang J
,
Le P
,
Bülow P
,
Garnier-Amblard E
,
Gangireddy PKR
,
Bassell GJ
,
Yuan H
,
Menaldino DS
,
Liotta DC
,
Liebeskind LS
,
Traynelis SF
.
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Allosteric modulators of ion channels typically alter the transitions rates between conformational states without changing the properties of the open pore. Here we describe a new class of positive allosteric modulators of N-methyl D-aspartate receptors (NMDARs) that mediate a calcium-permeable component of glutamatergic synaptic transmission and play essential roles in learning, memory and cognition, as well as neurological disease. EU1622-14 increases agonist potency and channel-open probability, slows receptor deactivation and decreases both single-channel conductance and calcium permeability. The unique functional selectivity of this chemical probe reveals a mechanism for enhancing NMDAR function while limiting excess calcium influx, and shows that allosteric modulators can act as biased modulators of ion-channel permeation.
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R01 MH109026 NIMH NIH HHS, R01 HD082373 NICHD NIH HHS , R01 NS065371 NINDS NIH HHS , R35 NS111619 NINDS NIH HHS
GO keywords: ion channel activity
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Figure 2. EU1622-1 reduces single channel conductance for GluN1/GluN2B NMDARs. (A) Unitary currents were recorded from an outside-out patch excised from an HEK cell co-transfected with GluN1, GluN2B, and GFP. The holding potential was −80 mV and channel openings are shown as downward by convention. Responses were produced by the application of external solution with 0.1 mM glutamate plus 0.05 mM glycine (0.5 mM Ca2+). NMDARs showed reduced unitary current amplitude when 50 μM EU1622-1 was co-applied with the agonist (lower panel) compared to vehicle (upper panel). (C) Composite amplitude histograms determined by time course fitting were analyzed by fitting with the sum of multiple Gaussian components by maximum likelihood for vehicle (open bars, solid line, left y-axis) and 30–50 μM EU1622-1 (filled bars, dashed line, right y-axis). Supplementary Table 8 provides a summary of chord conductance. (D) Composite open duration histograms for the patch shown and fitted by the sum of two exponential components displayed similarly to (C). Mean open time is given in Supplementary Table 8; individual fitted time constants ± standard deviation and area for the fits were: vehicle Tau1 0.10 ± 0.0028 ms, 12%, Tau2 2.3 ± 0.31 ms, 88%, EU1622-1 Tau1 0.54 ± 0.40 ms, 26%, Tau2 3.2 ± 0.93 ms, 74% (n = 3 patches). (E) Examples of direct sublevel transitions in the presence of EU1622-1 are shown, marked by arrowheads. | |
Figure 3. EU1622-14 prolongs the response deactivation time course and reduces channel conductance of NMDARs. (A) Representative GluN1/GluN2B NMDAR current responses during rapid application of 1/0.1 mM glutamate/glycine for 10 ms (triangle) in 100 μM glycine (white bar) with vehicle or EU1622-14 (30 μM, Compound 2). Horizontal triangles show peak amplitudes. An expanded, normalized view is shown on the right to highlight the deactivation time course of the responses. (B) Peak amplitude (left) and τweighted of the deactivation (right) of rapid glutamate applications (10 ms) with and without EU1622-14 for all diheteromeric NMDARs (see Supplementary Table 4). (C) Representative slow perfusion stimulated GluN1/GluN2B responses with glutamate and glycine co-applied with vehicle or E1622-14. The NMDAR-mediated increase in variance can be seen from the high-pass filtered current (grey, below). (D) Current-variance plot of the rise and decay of the low concentration agonist application from (C). (E) Mean conductance of all diheterometic NMDARs with and without EU1622-14 (50 μM). For summary data see Supplementary Table 4. For all panels, * p<0.05 by paired t-test, with the Holm-Bonferroni correction to control for the family wise error rate in (B). | |
Figure 4. EU1622-14 reduces native NMDAR conductance. Unitary currents activated by 500 μM NMDA and 100 μM glycine (−80 mV) in outside-out patches were from cortical neurons in 1.25 (A) or 0.5 mM Ca2+ (B); scale bars are 2 s (A1), 100 ms (A2) and 50 ms (A3). The dashed line is 5 pA. “C” indicates closed state and arrowheads in A3 show sublevel transitions in EU1622-14. Vehicle was 0.25% DMSO. (C,D) Amplitudes determined (6,369 vehicle, 14,406 EU1622-14) from 6 patches in 1.25 mM Ca2+ or 4 patches (3,473 vehicle, 6,639 EU1622-14) in 0.5 mM Ca2+ were fitted to the sum of 2–3 Gaussians (smooth line is the probability density function). The amplitude ± standard deviation and area for the fits (1.25 mM Ca2+) were: vehicle Amp1 −4.13 ± 0.36 pA, 85%, Amp2 −3.52 ± 0.83 pA, 15%, EU1622-14 Amp1 −3.45 ± 0.59 pA, 71% Amp2 −2.58 ± 0.22 pA, 23% Amp3 −2.16 ± 0.57 pA, 6 %. The amplitude ± standard deviation and area for the fits (0.5 mM Ca2+) were: vehicle Amp1 −4.79 ± 0.35 pA, 73% Amp2 −4.16 ± 0.75 pA, 27%, EU1622-14 Amp1 −4.23 ± 0.55 pA, 79%, Amp2 −3.17 ± 0.26 pA, 16% Amp3 −2.27 ± 0.65 pA, 4%. | |
Figure 5. EU1622-14 reduces Ca2+ permeation through NMDARs. (A) Representative I-V curves for GluN1/GluN2A responses to maximally-effective agonist co-applied with vehicle (left) or EU1622-14 (right) in the low/high Ca2+. A high monovalent method was used to determine the relative Ca2+/monovalent permeability ratio. The fitted curve is a fourth-order polynomial. (B) The average reversal potentials are given for GluN1/GluN2A and GluN1/GluN2B (vehicle vs. EU1622-14). (C) The mean ΔReversal potentials (high Ca2+ minus low Ca2+) are given. (D) The average Ca2+ permeability ratio to monovalent ions is from the Lewis equation. *p<0.05 (One-way ANOVA, post-hoc t-test of drug vs vehicle, Bonferroni correction for multiple comparisons); F3,28 = 6.91. For summary data see Supplementary Table 9. | |
Figure 6. EU1622-14 biasedly modulates Na+ and Ca2+ influx through NMDARs. (A-C) Simultaneous patch-clamp/imaging recordings from cortical neuronal cultures illustrate a significant augmentation of total current with a non-significant increase in the Ca2+ fluorescence (stimulated by 500/100 μM NMDA/glycine) due to EU1622-14 modulation. (A) Time-lapse imaging of a Ca2+-sensitive, cell impermeant dye (Calbryte 590 potassium-salt, images were taken at a 1s interval). Representative vehicle and EU1622-14 fluorescence B, left) and current (C, left) responses to 500/100 μM NMDA/glycine co-applied with vehicle (equivalent volume DMSO) or EU1622-14 (50 μM) following a 1 min pre-application of vehicle or drug. Summary graphs are shown on the right of each figure panel. A significant increase in the peak current amplitude (p = 0.0004, unpaired t-test) was detected in the presence of EU1622-14 (50 μM) but not a significant increase in the slope of the Ca2+-fluorescence signal (p = 0.09, unpaired t-test). (D-G) Simultaneous fluorescence monitoring of Na+ and Ca2+ influx using specific cell permeable dyes illustrates selective modulation by EU1622-14. (D) Intracellular Na+ and Ca2+ levels were measured in cortical neuronal cultures utilizing time-lapse imaging of Na+- and Ca2+-sensitive dyes. 500/100 μM NMDA/glycine were co-applied in vehicle (0.5% DMSO) or EU1622-14 (50 μM) following a 1 min pre-application of vehicle or drug. Images were acquired every 4 s. Boxed regions in (E) are shown in (D). (F-G) Cellular fluorescence responses to NMDAR stimulation in the presence of vehicle or EU1622-14. Agonist solution perfusion (indicate by the arrow, 500/100 μM NMDA/glycine, with vehicle or EU1622-14) resulted in a rise in the CoroNa AM dye (F, Na+ response) and the Calbryte 590 AM dye (G, Ca2+ response). For each trial, the slope of the rise of the respective signals were measured for all neurons in the camera view, then averaged to obtain one experimental data point (shown in the plot on the right). Scale bars denote 50 μm. |
References [+] :
Bassetti,
Somatomammotrophic cells in GH-secreting and PRL-secreting human pituitary adenomas.
1990, Pubmed
Bassetti, Somatomammotrophic cells in GH-secreting and PRL-secreting human pituitary adenomas. 1990, Pubmed
Belinsky, Role of DNA methylation in the activation of proto-oncogenes and the induction of pulmonary neoplasia by nitrosamines. 1990, Pubmed
Burnashev, Fractional calcium currents through recombinant GluR channels of the NMDA, AMPA and kainate receptor subtypes. 1995, Pubmed
Choi, Excitotoxic cell death. 1992, Pubmed
Chopra, A single-channel mechanism for pharmacological potentiation of GluN1/GluN2A NMDA receptors. 2017, Pubmed , Xenbase
Collingridge, The NMDA receptor as a target for cognitive enhancement. 2013, Pubmed
Coyle, Converging evidence of NMDA receptor hypofunction in the pathophysiology of schizophrenia. 2003, Pubmed
Davies, The mitotic response of thymus-derived cells to antigenic stimulus. 1967, Pubmed
Degreef, Miconazole nitrate in the treatment of dermatomycoses. 1975, Pubmed
Dravid, Activation of recombinant NR1/NR2C NMDA receptors. 2008, Pubmed
Erreger, Subunit-specific agonist activity at NR2A-, NR2B-, NR2C-, and NR2D-containing N-methyl-D-aspartate glutamate receptors. 2007, Pubmed , Xenbase
Freedman, Compensation for auditory re-arrangement in hand-ear coordination. 1970, Pubmed
Gibb, A structurally derived model of subunit-dependent NMDA receptor function. 2018, Pubmed
Gonzalez, NMDARs in neurological diseases: a potential therapeutic target. 2015, Pubmed
Hackos, Positive Allosteric Modulators of GluN2A-Containing NMDARs with Distinct Modes of Action and Impacts on Circuit Function. 2016, Pubmed
Hansen, Implementation of a fluorescence-based screening assay identifies histamine H3 receptor antagonists clobenpropit and iodophenpropit as subunit-selective N-methyl-D-aspartate receptor antagonists. 2010, Pubmed , Xenbase
Heresco-Levy, Double-blind, placebo-controlled, crossover trial of D-cycloserine adjuvant therapy for treatment-resistant schizophrenia. 1998, Pubmed
Hu, Human GRIN2B variants in neurodevelopmental disorders. 2016, Pubmed
Ikonomidou, Why did NMDA receptor antagonists fail clinical trials for stroke and traumatic brain injury? 2002, Pubmed
Ingram, New pharmacological strategies for cognitive enhancement using a rat model of age-related memory impairment. 1994, Pubmed
Jatzke, Voltage and concentration dependence of Ca(2+) permeability in recombinant glutamate receptor subtypes. 2002, Pubmed
Javitt, Management of negative symptoms of schizophrenia. 2001, Pubmed
Karakas, Crystal structure of a heterotetrameric NMDA receptor ion channel. 2014, Pubmed
Kazi, Asynchronous movements prior to pore opening in NMDA receptors. 2013, Pubmed
Khatri, Structural determinants and mechanism of action of a GluN2C-selective NMDA receptor positive allosteric modulator. 2014, Pubmed , Xenbase
Kleinberg, Biochemistry of the dental plaque. 1970, Pubmed
Kokosov, [Principles of treatment of the infectious-allergic form of bronchial asthma]. 1976, Pubmed
Krüger, [Ischemic occlusion of the central retinal vein and protein C deficiency]. 1991, Pubmed
Lee, NMDA receptor structures reveal subunit arrangement and pore architecture. 2014, Pubmed , Xenbase
Lester, Channel kinetics determine the time course of NMDA receptor-mediated synaptic currents. 1990, Pubmed
Letourneau, The selection of physicians. 1966, Pubmed
Lewis, Ion-concentration dependence of the reversal potential and the single channel conductance of ion channels at the frog neuromuscular junction. 1979, Pubmed
MacGowan, Screening of urines with dipstrips: does it reduce workload and consumable costs? 1990, Pubmed
Mizuta, Developmental expression of NMDA receptor subunits and the emergence of glutamate neurotoxicity in primary cultures of murine cerebral cortical neurons. 1998, Pubmed
Mullasseril, A subunit-selective potentiator of NR2C- and NR2D-containing NMDA receptors. 2010, Pubmed
Nakano, [Nausea and vomiting]. 1973, Pubmed
Neher, Correction for liquid junction potentials in patch clamp experiments. 1992, Pubmed
Ogden, Contribution of the M1 transmembrane helix and pre-M1 region to positive allosteric modulation and gating of N-methyl-D-aspartate receptors. 2013, Pubmed , Xenbase
Ogden, Molecular Mechanism of Disease-Associated Mutations in the Pre-M1 Helix of NMDA Receptors and Potential Rescue Pharmacology. 2017, Pubmed , Xenbase
Paoletti, Glycine-independent and subunit-specific potentiation of NMDA responses by extracellular Mg2+. 1995, Pubmed , Xenbase
Parsons, Extrasynaptic NMDA receptor involvement in central nervous system disorders. 2014, Pubmed
Pepe, Glucose-6-phosphate dehydrogenase activity in the endometrium and myometrium of the rat uterus during the estrous cycle and progestation. 1972, Pubmed
Perszyk, An NMDAR positive and negative allosteric modulator series share a binding site and are interconverted by methyl groups. 2018, Pubmed , Xenbase
Perszyk, GluN2D-Containing N-methyl-d-Aspartate Receptors Mediate Synaptic Transmission in Hippocampal Interneurons and Regulate Interneuron Activity. 2016, Pubmed , Xenbase
Premkumar, Identification of a high affinity divalent cation binding site near the entrance of the NMDA receptor channel. 1996, Pubmed , Xenbase
Rapaport, [Use of instrumental conditioning in Skinner's box as test of adaptation to environmental hyperthermia in young rats]. 1972, Pubmed
Rosenmund, The tetrameric structure of a glutamate receptor channel. 1998, Pubmed
Sandberg, Intolerance to lactose in Negro children. 1971, Pubmed
Sapkota, Mechanism and properties of positive allosteric modulation of N-methyl-d-aspartate receptors by 6-alkyl 2-naphthoic acid derivatives. 2017, Pubmed , Xenbase
Schade, D-Cycloserine in Neuropsychiatric Diseases: A Systematic Review. 2016, Pubmed
Schewe, A pharmacological master key mechanism that unlocks the selectivity filter gate in K+ channels. 2019, Pubmed , Xenbase
Schrezenmeir, The role of oxygen supply in islet transplantation. 1993, Pubmed
Siegler Retchless, A single GluN2 subunit residue controls NMDA receptor channel properties via intersubunit interaction. 2012, Pubmed
So, Haemoptysis due to unsuspected foreign body penetration of the oesophagus. 1983, Pubmed
Sobolevsky, Subunit-specific contribution of pore-forming domains to NMDA receptor channel structure and gating. 2007, Pubmed , Xenbase
Strong, The Structure-Activity Relationship of a Tetrahydroisoquinoline Class of N-Methyl-d-Aspartate Receptor Modulators that Potentiates GluN2B-Containing N-Methyl-d-Aspartate Receptors. 2017, Pubmed
Swanger, A Novel Negative Allosteric Modulator Selective for GluN2C/2D-Containing NMDA Receptors Inhibits Synaptic Transmission in Hippocampal Interneurons. 2018, Pubmed , Xenbase
Talukder, Specific sites within the ligand-binding domain and ion channel linkers modulate NMDA receptor gating. 2010, Pubmed , Xenbase
Traynelis, Glutamate receptor ion channels: structure, regulation, and function. 2010, Pubmed
Traynelis, Software-based correction of single compartment series resistance errors. 1998, Pubmed
Traynelis, Getting the most out of noise in the central nervous system. 1998, Pubmed
Twomey, Structural Mechanisms of Gating in Ionotropic Glutamate Receptors. 2018, Pubmed
Wang, A novel NMDA receptor positive allosteric modulator that acts via the transmembrane domain. 2017, Pubmed
Watanabe, DRPEER: a motif in the extracellular vestibule conferring high Ca2+ flux rates in NMDA receptor channels. 2002, Pubmed , Xenbase
Willett, ABC of major trauma. Management of limb injuries. 1990, Pubmed
Wollmuth, Different mechanisms of Ca2+ transport in NMDA and Ca2+-permeable AMPA glutamate receptor channels. 1998, Pubmed
Woodhull, Ionic blockage of sodium channels in nerve. 1973, Pubmed
Wyllie, Single-channel currents from recombinant NMDA NR1a/NR2D receptors expressed in Xenopus oocytes. 1996, Pubmed , Xenbase
Yuan, Ionotropic GABA and Glutamate Receptor Mutations and Human Neurologic Diseases. 2015, Pubmed
Zheng, Selectivity changes during activation of mutant Shaker potassium channels. 1997, Pubmed , Xenbase
Zhu, Mechanism of NMDA Receptor Inhibition and Activation. 2016, Pubmed , Xenbase