Interaction of 6-cyano-7-nitroquinoxaline-2,3-dione with the N-methyl-D-aspartate receptor-associated glycine binding site

R. A J Lester, M. L. Quarum, J. D. Parker, E. Weber, Craig Jahr

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Abstract

The interaction of newly described antagonist of the non-NMDA glutamate receptor 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX) with the glycine site of the NMDA receptor complex has been investigated. In whole-cell patch recordings from hippocampal neurons maintained in culture, currents induced by N-methyl-D-aspartate (NMDA) were dependent on extracellular glycine. Responses to both NMDA (30 μM) and kainate (20 μM) were reduced by CNQX (10-30 μM). The antagonism by CNQX of NMDA, but not kainate, receptor-mediated responses could be reversed by increasing the concentration of glycine in the external medium. Glycine concentration-response curves constructed in the presence of 30 μM NMDA were shifted to the right by CNQX, suggesting that CNQX was competing with glycine for the glycine binding site. However, even at high concentrations of glycine (300 μM) the maximal NMDA current obtained in the presence of CNQX (10-30 μM) was not restored to control levels. Because CNQX had no effect on responses produced by supramaximal concentrations of NMDA (500 μM) and glycine (300 μM), it is suggested that CNQX also interacts with the NMDA recognition site. The antagonism of currents induced by NMDA was not dependent on the membrane potential, and the rapid onset and offset of the block suggested that there was little or no use dependence. Radioligand binding experiments were performed using [3H]glycine to label the strychnine-insensitive glycine regulatory site of the NMDA receptor complex in guinea pig brain frontal cortex membranes. CNQX displaced [3H]glycine binding in a concentration-dependent manner (IC50 = 5.7 μM). Scatchard analysis of the inhibition showed a decrease in the affinity (increase in K(d)) of [3H]glycine binding, but no change in the number of binding sites (B(max)) in the presence of 5 μM CNQX, suggesting a competitive interaction. These data provide evidence that CNQX antagonizes NMDA receptor-mediated responses by competing with glycine for a modulatory site associated with the NMDA receptor complex. Furthermore, the results indicate that CNQX may not be as selective an antagonist for non-NMDA receptors as initially described, although its selectivity will depend on the concentration of the NMDA receptor ligand and may be enhanced by increasing the extracellular concentration of glycine.

Original languageEnglish (US)
Pages (from-to)565-570
Number of pages6
JournalMolecular Pharmacology
Volume35
Issue number5
StatePublished - 1989

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6-Cyano-7-nitroquinoxaline-2,3-dione
N-Methyl-D-Aspartate Receptors
Glycine
Binding Sites
N-Methylaspartate
D-Aspartic Acid
Kainic Acid Receptors
Strychnine
Kainic Acid
Glutamate Receptors
Frontal Lobe
Patch-Clamp Techniques

ASJC Scopus subject areas

  • Pharmacology

Cite this

Lester, R. A. J., Quarum, M. L., Parker, J. D., Weber, E., & Jahr, C. (1989). Interaction of 6-cyano-7-nitroquinoxaline-2,3-dione with the N-methyl-D-aspartate receptor-associated glycine binding site. Molecular Pharmacology, 35(5), 565-570.

Interaction of 6-cyano-7-nitroquinoxaline-2,3-dione with the N-methyl-D-aspartate receptor-associated glycine binding site. / Lester, R. A J; Quarum, M. L.; Parker, J. D.; Weber, E.; Jahr, Craig.

In: Molecular Pharmacology, Vol. 35, No. 5, 1989, p. 565-570.

Research output: Contribution to journalArticle

Lester, RAJ, Quarum, ML, Parker, JD, Weber, E & Jahr, C 1989, 'Interaction of 6-cyano-7-nitroquinoxaline-2,3-dione with the N-methyl-D-aspartate receptor-associated glycine binding site', Molecular Pharmacology, vol. 35, no. 5, pp. 565-570.
Lester, R. A J ; Quarum, M. L. ; Parker, J. D. ; Weber, E. ; Jahr, Craig. / Interaction of 6-cyano-7-nitroquinoxaline-2,3-dione with the N-methyl-D-aspartate receptor-associated glycine binding site. In: Molecular Pharmacology. 1989 ; Vol. 35, No. 5. pp. 565-570.
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abstract = "The interaction of newly described antagonist of the non-NMDA glutamate receptor 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX) with the glycine site of the NMDA receptor complex has been investigated. In whole-cell patch recordings from hippocampal neurons maintained in culture, currents induced by N-methyl-D-aspartate (NMDA) were dependent on extracellular glycine. Responses to both NMDA (30 μM) and kainate (20 μM) were reduced by CNQX (10-30 μM). The antagonism by CNQX of NMDA, but not kainate, receptor-mediated responses could be reversed by increasing the concentration of glycine in the external medium. Glycine concentration-response curves constructed in the presence of 30 μM NMDA were shifted to the right by CNQX, suggesting that CNQX was competing with glycine for the glycine binding site. However, even at high concentrations of glycine (300 μM) the maximal NMDA current obtained in the presence of CNQX (10-30 μM) was not restored to control levels. Because CNQX had no effect on responses produced by supramaximal concentrations of NMDA (500 μM) and glycine (300 μM), it is suggested that CNQX also interacts with the NMDA recognition site. The antagonism of currents induced by NMDA was not dependent on the membrane potential, and the rapid onset and offset of the block suggested that there was little or no use dependence. Radioligand binding experiments were performed using [3H]glycine to label the strychnine-insensitive glycine regulatory site of the NMDA receptor complex in guinea pig brain frontal cortex membranes. CNQX displaced [3H]glycine binding in a concentration-dependent manner (IC50 = 5.7 μM). Scatchard analysis of the inhibition showed a decrease in the affinity (increase in K(d)) of [3H]glycine binding, but no change in the number of binding sites (B(max)) in the presence of 5 μM CNQX, suggesting a competitive interaction. These data provide evidence that CNQX antagonizes NMDA receptor-mediated responses by competing with glycine for a modulatory site associated with the NMDA receptor complex. Furthermore, the results indicate that CNQX may not be as selective an antagonist for non-NMDA receptors as initially described, although its selectivity will depend on the concentration of the NMDA receptor ligand and may be enhanced by increasing the extracellular concentration of glycine.",
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N2 - The interaction of newly described antagonist of the non-NMDA glutamate receptor 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX) with the glycine site of the NMDA receptor complex has been investigated. In whole-cell patch recordings from hippocampal neurons maintained in culture, currents induced by N-methyl-D-aspartate (NMDA) were dependent on extracellular glycine. Responses to both NMDA (30 μM) and kainate (20 μM) were reduced by CNQX (10-30 μM). The antagonism by CNQX of NMDA, but not kainate, receptor-mediated responses could be reversed by increasing the concentration of glycine in the external medium. Glycine concentration-response curves constructed in the presence of 30 μM NMDA were shifted to the right by CNQX, suggesting that CNQX was competing with glycine for the glycine binding site. However, even at high concentrations of glycine (300 μM) the maximal NMDA current obtained in the presence of CNQX (10-30 μM) was not restored to control levels. Because CNQX had no effect on responses produced by supramaximal concentrations of NMDA (500 μM) and glycine (300 μM), it is suggested that CNQX also interacts with the NMDA recognition site. The antagonism of currents induced by NMDA was not dependent on the membrane potential, and the rapid onset and offset of the block suggested that there was little or no use dependence. Radioligand binding experiments were performed using [3H]glycine to label the strychnine-insensitive glycine regulatory site of the NMDA receptor complex in guinea pig brain frontal cortex membranes. CNQX displaced [3H]glycine binding in a concentration-dependent manner (IC50 = 5.7 μM). Scatchard analysis of the inhibition showed a decrease in the affinity (increase in K(d)) of [3H]glycine binding, but no change in the number of binding sites (B(max)) in the presence of 5 μM CNQX, suggesting a competitive interaction. These data provide evidence that CNQX antagonizes NMDA receptor-mediated responses by competing with glycine for a modulatory site associated with the NMDA receptor complex. Furthermore, the results indicate that CNQX may not be as selective an antagonist for non-NMDA receptors as initially described, although its selectivity will depend on the concentration of the NMDA receptor ligand and may be enhanced by increasing the extracellular concentration of glycine.

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