Pharmacological characterization of recombinant NR1/NR2A NMDA receptors with truncated and deleted carboxy termini expressed in Xenopus laevis oocytes

C. A. Puddifoot, P. E. Chen, R. Schoepfer, D. J. A. Wyllie

Research output: Contribution to journalArticlepeer-review

Abstract

The carboxy terminal domain (CTD) of NR2 N-methyl-d-aspartate receptor (NMDAR) subunits interacts with numerous scaffolding and signal transduction proteins. Mutations of this region affect trafficking and downstream signalling of NMDARs. This study determines to what extent characteristic pharmacological properties of NR2A-containing NMDARs are influenced by this key functional domain.

Using recombinant receptor expression in Xenopus laevis oocytes and two electrode voltage clamp recordings we characterized pharmacological properties of rat NR1/NR2A NMDARs with altered CTDs. We assessed the effects of truncating [at residue Iso1098; NR2A(trunC)] and deleting [from residue Phe822; NR2A(delC)] the CTD of NR2A NMDAR subunits on agonist potencies, channel block by Mg2+ and memantine and potentiation of NMDAR-mediated responses by chelating contaminating divalent cations.

Truncation or deletion of the CTD of NR2A NMDAR subunits did not affect glutamate potency [EC50 = 2.2 mu mol.L-1, NR2A(trunC); 2.7 mu mol.L-1, NR2A(delC) compared with 3.3 mu mol.L-1, NR2A(WT)] but did significantly increase glycine potency [EC50 = 500 nmol.L-1, NR2A(trunC); 900 nmol.L-1, NR2A(delC) compared with 1.3 mu mol.L-1, NR2A(WT)]. Voltage-dependent Mg2+ block of NR2A(WT)- and NR2A(trunC)-containing NMDARs was similar but low concentrations of Mg2+ (1 mu mol.L-1) potentiated NR1/NR2A(delC) NMDARs. Memantine block was not affected by changes to the structure of the NR2A CTD. EDTA-induced potentiation was similar at each of the three NMDAR constructs.

Of the parameters studied only minor influences of the CTD were observed; these are unlikely to compromise interpretation of studies that make use of CTD-mutated recombinant receptors or transgenic mice in investigations of the role of the CTD in NMDAR signalling.

Original languageEnglish
Pages (from-to)509-518
Number of pages10
JournalBritish Journal of Pharmacology
Volume156
Issue number3
DOIs
Publication statusPublished - Feb 2009

Keywords

  • NMDA
  • glutamate
  • glycine
  • memantine
  • oocyte
  • two electrode voltage clamp
  • electrophysiology
  • ion channel

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