High affinity [3H]dextrorphan binding in rat brain is localized to a noncompetitive antagonist site of the activated N-methyl-D-aspartate receptor-cation channel

P. H. Franklin, T. F. Murray

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[3H]Dextrorphan recognition sites were characterized in rat brain membranes. The pharmacological profile and regional distribution of [3H]dextrorphan binding sites appear to distinguish these sites from those labeled either by [3H]dextromethorphan or by putative σ receptor radioligands. Data from thoroughly washed forebrain membranes suggest that [3H]dextrorphan predominantly labels a high affinity site defined by the activated state of the N-methyl-D-aspartate (NMDA) receptor-channel complex. Regulation of [3H]dextrorphan binding by specific modulators of NMDA receptor function suggests that [3H]dextrorphan binding is predominantly localized to a domain of the receptor-channel complex also recognized by the prototypical noncompetitive antagonist radioligands (+)-[3H]5-methyl-10, 11-dihydro-5H-dibenzo[a,d]cyclohepten-5, 10-imine (MK-801) and [3H]1-[1-(2-thienyl)cyclohexyl]piperidine (TCP). The critical relationship between [3H]dextrorphan binding and activation of the NMDA receptor-complex is suggested by the profound dependence of [3H]dextrorphan binding on glutamate in well washed membranes. Basal specific [3H]dextrorphan binding is nearly totally suppressed by the specific competitive NMDA antagonist D(-)-2-amino-5-phosphonopentanoic acid (D-AP5), in a glutamate- but not glycine-surmountable manner. Glutamate and glycine each stimulate [3H]dextrorphan binding in a concentration-dependent manner, effecting maximal increases from control of up to 30-and 14-fold, respectively. The NMDA receptor specificity of the modulation of [3H]dextrorphan binding by glutamate and glycine is indicated by the sensitivity of their effects to competitive antagonism by D-AP5 and 3-amino-1-hydroxy-2-pyrrolidone (HA-966), respectively, and by the accordant rank orders of potency of glycine analogs as modulators of [3H]dextrorphan binding and as ligands at the strychnine-insensitive glycine site. The divalent cations Mg2+ and Zn2+ and the polyamines spermine and spermidine regulate [3H]dextrorphan binding in a manner consistent with radioligand interaction at the noncompetitive NMDA antagonist domain. Mg2+ and spermidine regulate [3H]dextrorphan binding biphasically in well washed forebrain membranes, whereas Zn2+ monotonically inhibits [3H]dextrorphan binding. Mg2+ and spermidine regulate [3H]dextrorphan binding with qualitative similarity and in a contrasting fashion to their regulation of [3H]MK-801 and [3H]TCP binding. First, spermidine and Mg2+ are significantly more potent modulators of [3H]dextrorphan binding than of [3H]MK-801 and [3H]TCP binding in well washed membranes; second, whereas the potencies of spermidine and Mg2+ as modulators of [3H]MK-801 and [3H]TCP binding are significantly increased by glutamate and glycine in well washed membranes, their potencies as regulators of [3H]dextrorphan binding appear to be unaffected by glutamate and glycine. Furthermore, putrescine, which does not influence [3H]MK-801 or [3H]TCP binding, inhibits basal [3H]dextrorphan binding in a manner dissimilar from that of spermidine- and spermine-mediated inhibition of binding. The kinetics of [3H]dextrorphan binding in the presence of saturating concentrations of glutamate and glycine are complex and inadequately described by monoexponential association and dissociation processes. The differential distribution of [3H]dextrorphan recognition sites in rat brain regions and the pharmacological profile specified by the rank order of potency of an extensive set of compounds as competitors for high affinity [3H]dextrorphan binding unambiguously suggest that [3H]dextrorphan binding in rat brain membranes corresponds to the site of the NMDA antagonist activity of dextrorphan in vivo. In contrast, the pharmacological signature and distribution of high affinity [3H]dextrorphan binding sites in rat brain are incongruous with those of either the σ receptor or [3H]dextromethorphan binding sites. Accordingly, the interaction of dextrorphan and dextromethorphan at sites labeled by [3H]dextrorphan, but not at sites labeled by [3H]dextromethorphan or by σ ligands, adequately accounts for the anticonvulsant and neuroprotective efficacies of these compounds in vivo.

Original languageEnglish (US)
Pages (from-to)134-146
Number of pages13
JournalMolecular Pharmacology
Issue number1
StatePublished - Feb 21 1992
Externally publishedYes

All Science Journal Classification (ASJC) codes

  • Molecular Medicine
  • Pharmacology


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