scholarly journals Competitive Inhibition at the Glycine Site of the N -Methyl-d-aspartate Receptor by the Anesthetics Xenon and Isoflurane

2007 ◽  
Vol 107 (5) ◽  
pp. 756-767 ◽  
Author(s):  
Robert Dickinson ◽  
Brian K. Peterson ◽  
Paul Banks ◽  
Constantinos Simillis ◽  
Juan Carlos Sacristan Martin ◽  
...  
Keyword(s):  
Nmda Receptor ◽  
Nmda Receptors ◽  
Competitive Inhibition ◽  
Inhibitory Effect ◽  
Glycine Site ◽  
Aspartate Receptor ◽  
Mechanism Of Inhibition ◽  
Anesthetic Gases ◽  
Mutant Receptors ◽  
Grand Canonical

Background Inhibition of N-methyl-D-aspartate (NMDA) receptors by anesthetic gases and vapors may play an important role in anesthesia and neuroprotection. However, the site of action of these agents on the NMDA receptor is unknown. The authors show that xenon and isoflurane compete for the binding of the coagonist glycine on the NMDA receptor NR1 subunit. Methods Using a novel application of grand canonical Monte Carlo simulations, the authors predict the binding site of xenon on NMDA receptors. They test this prediction using electrophysiology on recombinant NMDA receptors. Results The authors' modeling predicts that xenon binds at the glycine site of the NMDA receptor. The authors show that inhibition of NMDA receptors by xenon and isoflurane increases as glycine concentration is decreased, consistent with the prediction of competitive inhibition at the glycine site. Lineweaver-Burk analysis shows that isoflurane inhibition seems purely competitive with glycine, but for xenon, there is an additional component of noncompetitive inhibition. The loss of inhibitory effect of xenon and isoflurane in mutant NR1(F639A)/NR2A receptors is explained by increased glycine affinity of the mutant receptors, and inhibition is restored at low glycine concentrations. Conclusions Xenon and isoflurane inhibit NMDA receptors by binding at the same site as the coagonist glycine. This finding may have important implications for general anesthesia and neuroprotection. Neuroprotectants that act at the glycine site of the NMDA receptor antagonists are well tolerated in patients, being devoid of psychotomimetic side effects, and the mechanism of inhibition may play a role in their clinical profile.

scholarly journals The induction of N -methyl-D-aspartate receptor-dependent long-term potentiation

2003 ◽  
Vol 358 (1432) ◽  
pp. 635-641 ◽  
Author(s):  
Graham L. Collingridge
Keyword(s):  
Nmda Receptor ◽  
Nmda Receptors ◽  
Long Term Potentiation ◽  
Personal Reflection ◽  
Aspartate Receptor ◽  
Term Potentiation

The role of N -methyl-D-aspartate (NMDA) receptors in the induction of long-term potentiation (LTP) was established during the 1980s. In this article I present a personal reflection upon the role that my colleagues and I played in the discovery of the mechanism of induction of NMDA receptor-dependent LTP.

Evidence That Xenon Does Not Produce Open Channel Blockade of the NMDA Receptor

2008 ◽  
Vol 99 (4) ◽  
pp. 1983-1987 ◽  
Author(s):  
Henry U. Weigt ◽  
Oliver Adolph ◽  
Michael Georgieff ◽  
Eva M. Georgieff ◽  
Karl J. Föhr
Keyword(s):  
Nmda Receptor ◽  
Nmda Receptors ◽  
Open Channel ◽  
Channel Blocker ◽  
Channel Blockers ◽  
Cell Recording ◽  
Neuro2a Cells ◽  
Induced Currents ◽  
Mutant Receptors ◽  
Open Channel Blocker

Previous studies had not excluded the possibility that the mechanism by which Xenon (Xe) blocks N-methyl-d-aspartate (NMDA) receptors might be that of an open-channel blocker. We tested this possibility on mutant NMDA receptors carrying an alanine (A) to cysteine (C) mutation located within the SYTANLAAF-motif of the third transmembrane region (TM3). This mutation was shown to yield constitutively open ion channels after modification with a thiol-modifying reagent. We expressed such mutant channels in Neuro2A cells and recorded glutamate (50 μM)-induced currents in the whole cell recording mode. Although Xe (3.5 mM) blocked the currents through the wild-type receptor NR1-1a/NR2A and NR1-1a/NR2B by ∼40% and those through the mutant receptors NR1-1a/NR2A(A650C) or NR1-1a/NR2B(A651C) by ∼30%, it was unable to block the currents through the methane thiosulfonate etyhlammonium-modified mutant receptors. On the other hand, established open-channel blockers of the NMDA receptor such as MK-801 (1 μM) or Mg ions (Mg2+; 1 mM) were able to block these permanently open channels. These results suggest that Xe does not act as a classical open-channel blocker at the NMDA receptor.

scholarly journals Identification of Two Mutations (F758W and F758Y) in the N -methyl-D-aspartate Receptor Glycine-binding Site that Selectively Prevent Competitive Inhibition by Xenon without Affecting Glycine Binding

2012 ◽  
Vol 117 (1) ◽  
pp. 38-47 ◽  
Author(s):  
Scott P. Armstrong ◽  
Paul J. Banks ◽  
Thomas J. W. McKitrick ◽  
Catharine H. Geldart ◽  
Christopher J. Edge ◽  
...  
Keyword(s):  
Amino Acids ◽  
Nmda Receptor ◽  
Binding Site ◽  
Competitive Inhibition ◽  
Site Directed Mutagenesis ◽  
Glycine Site ◽  
General Anesthetic ◽  
Hek 293

Background Xenon is a general anesthetic with neuroprotective properties. Xenon inhibition at the glycine-binding site of the N-Methyl-D-aspartate (NMDA) receptor mediates xenon neuroprotection against ischemic injury in vitro. Here we identify specific amino acids important for xenon binding to the NMDA receptor, with the aim of finding silent mutations that eliminate xenon binding but leave normal receptor function intact. Methods Site-directed mutagenesis was used to mutate specific amino-acids in the GluN1 subunit of rat NMDA receptors. Mutant GluN1/GluN2A receptors were expressed in HEK 293 cells and were assessed functionally using patch-clamp electrophysiology. The responses of the mutant receptors to glycine and anesthetics were determined. Results Mutation of phenylalanine 758 to an aromatic tryptophan or tyrosine left glycine affinity unchanged, but eliminated xenon binding without affecting the binding of sevoflurane or isoflurane. Conclusions These findings confirm xenon binds to the glycine site of the GluN1 subunit of the NMDA receptor and indicate that interactions between xenon and the aromatic ring of the phenylalanine 758 residue are important for xenon binding. Our most important finding is that we have identified two mutations, F758W and F758Y, that eliminate xenon binding to the NMDA receptor glycine site without changing the glycine affinity of the receptor or the binding of volatile anesthetics. The identification of these selective mutations will allow knock-in animals to be used to dissect the mechanism(s) of xenon's neuroprotective and anesthetic properties in vivo.

scholarly journals Identification of an N-Methyl-d-aspartate Receptor in Isolated Nervous System Mitochondria

2012 ◽  
Vol 287 (42) ◽  
pp. 35192-35200 ◽  
Author(s):  
Amit S. Korde ◽  
William F. Maragos
Keyword(s):  
Cell Death ◽  
Nmda Receptor ◽  
Nmda Receptors ◽  
Calcium Uptake ◽  
Immunogold Electron Microscopy ◽  
Mitochondrial Calcium ◽  
Nmda Receptor Subunit ◽  
Tissue Sections ◽  
Aspartate Receptor ◽  
Nr2a Subunit

NMDA ionotropic glutamate receptors gate the cytoplasmic influx of calcium, which may, depending on the intensity of the stimulus, subserve either normal synaptic communication or cell death. We demonstrate that when isolated mitochondria are exposed to calcium and NMDA agonists, there is a significant increase in mitochondrial calcium levels. The agonist/antagonist response studies on purified mitochondria suggest the presence of a receptor on mitochondria with features similar to plasma membrane NMDA receptors. Immunogold electron microscopy of hippocampal tissue sections revealed extensive localization of NR2a subunit immunoreactivity on mitochondria. Transient transfection of neuronal GT1-7 cells with an NR1-NR2a NMDA receptor subunit cassette specifically targeting mitochondria resulted in a significant increase in mitochondrial calcium and neuroprotection against glutamate-induced cell death. Mitochondria prepared from GT1-7 cells in which the NR1 subunit of NMDA receptors was silenced demonstrated a decrease in calcium uptake. Our findings are the first to demonstrate that mitochondria express a calcium transport protein that shares characteristics with the NMDA receptor and may play a neuroprotective role.

Glycine at the NMDA receptor in cat visual cortex: saturation and changes with age

1996 ◽  
Vol 75 (1) ◽  
pp. 311-317 ◽  
Author(s):  
D. Czepita ◽  
N. W. Daw ◽  
S. N. Reid
Keyword(s):  
Visual Cortex ◽  
Nmda Receptor ◽  
Nmda Receptors ◽  
Visual Response ◽  
Glycine Site ◽  
Positive Evidence ◽  
Cat Visual Cortex ◽  
Site Of Action

1. Saturation of the glycine site at the N-methyl-D-aspartate (NMDA) receptor in cat visual cortex was tested by iontophoresing D-serine, and the contribution of this site to the visual response was tested by iontophoresing 7-chlorokynurenic acid (7-Cl-KYNA). Animals were tested at ages 3 wk, 6 wk, 5 mo, and adult. 2. In at least 40% of cells (24/57), D-serine increased the response significantly, showing that the glycine site was not saturated. However, the increase was rarely > 100%. 3. The amount by which D-serine increased the response did not vary significantly with age. 4. In five cases there was positive evidence for saturation of the glycine site. D-Serine did not increase the response significantly; however, when D-serine was applied on top of 7-Cl-KYNA, it did increase the response compared with that seen with 7-Cl-KYNA alone. These cases were all in 3-wk-old animals. In other cases (28/57), D-serine did not increase the response significantly, but we could not be absolutely certain that the D-serine had reached the site of action. 5. The effect of 7-Cl-KYNA was largest in 3-wk animals, in agreement with previous findings that the effect of D-2-amino-5-phosphonovalerate (APV) is largest at this age. 6. These results may provide a small part of the explanation for the finding that the NMDA receptor-mediated component of the visual response decreases between 3 and 6 wk of age, while the number of NMDA receptors is increasing. However, the magnitude of the results make it likely that other factors are more important.

Competitive Inhibition of NMDA Receptor–Mediated Currents by Extracellular Calcium Chelators

2000 ◽  
Vol 84 (2) ◽  
pp. 693-697 ◽  
Author(s):  
Nansheng Chen ◽  
Timothy H. Murphy ◽  
Lynn A. Raymond
Keyword(s):  
Nmda Receptor ◽  
Nmda Receptors ◽  
Dose Response ◽  
Response Curve ◽  
Competitive Inhibition ◽  
Response Shift ◽  
Dose Response Curve ◽  
Hek 293 ◽  
Electrophysiological Recordings ◽  
The Right

Calcium chelators have been widely used in electrophysiological recordings of N-methyl-d-aspartate (NMDA) receptor–mediated currents, as well as in studies of excitotoxicity. Intracellularly applied calcium chelators are known to inhibit, at least in part, such calcium-dependent processes as calmodulin-dependent inactivation, calcineurin-dependent desensitization, and rundown of NMDA receptors. On the other hand, the functional consequences and potential nonspecific effects of extracellularly applied chelators have not been extensively investigated. In whole-cell patch-clamp recordings from human embryonic kidney (HEK) 293 cells transiently transfected with recombinant NMDA receptors, we found that addition of calcium chelators such as EGTA shifted the glutamate dose-response curve to the right, from an EC50 for NR1A/NR2A of 8 μM in 1.8 mM Ca2+ to ∼24 μM in a solution containing nominal 0 Ca2+/5 mM EGTA and further to ∼80 μM in 20 mM EGTA. A similar shift in glutamate dose-response was observed for NR1A/NR2B currents. This dose-response shift was not due to a decrease in extracellular Ca2+ concentration because there was no change in the glutamate EC50 at Ca2+concentrations ranging from 10 mM to nominal 0/200 μM EGTA. Moreover, addition of 5 mM EGTA fully chelated with 6.8 mM Ca2+ did not produce any shift in the glutamate dose-response curve. We propose that calcium chelators, containing four free carboxyl moieties, competitively inhibit glutamate binding to NMDA receptors.

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