Glutamate receptor composition of the post-synaptic density is altered in genetic mouse models of NMDA receptor hypo- and hyperfunction

Abstract

The N-methyl-d-aspartate receptor (NMDAR) and α-amino-3-hydroxyl-5-methyl-4-isoxazole-propionate receptor (AMPAR) are ionotropic glutamate receptors responsible for excitatory neurotransmission in the brain. These excitatory synapses are found on dendritic spines, with the abundance of receptors concentrated at the postsynaptic density (PSD). We utilized two genetic mouse models, the serine racemase knockout (SR-/-) and the glycine transporter subtype 1 heterozygote mutant (GlyT1+/-), to determine how constitutive NMDAR hypo- and hyperfunction, respectively, affect the glutamate receptor composition of the PSD in the hippocampus and prefrontal cortex (PFC). Using cellular fractionation, we found that SR-/- mice had elevated protein levels of NR1 and NR2A NMDAR subunits specifically in the PSD-enriched fraction from the hippocampus, but not from the PFC. There were no changes in the amounts of AMPAR subunits (GluR1, GluR2), or PSD protein of 95 kDa (PSD95) in either brain region. GlyT1+/- mice also had elevated protein expression of NR1 and NR2A subunits in the PSD, as well as an increase in total protein. Moreover, GlyT1+/- mice had elevated amounts of GluR1 and GluR2 in the PSD, and higher total amounts of GluR1. Similar to SR-/- mice, there were no protein changes observed in the PFC. These findings illustrate the complexity of synaptic adaptation to altered NMDAR function.

Fig. 1

Altered hippocampal NMDAR expression selectively in the PSD of SR−/− mice. (A,B) Postsynaptic density enriched (PSD) and (C,D) whole cell lysate (WCL) hippocampal fractions were collected from adult wild type (WT; black bars; n = 6–12) and mutant (SR−/−; white bars; n = 6–12) mice. The amount of NMDAR subunits (NR1, NR2A, NR2B), postsynaptic density protein of 95kDa (PSD95), and α-amino-3-hydroxyl-5-methyl-4-isoxazole-propionate receptor (AMPAR) subunits (GluR1, GluR2) were quantified by Western blot analysis. Values are expressed as the optical density (OD) normalized to WT values (% control). Bars represent mean values ± s.e.m. Asterisk (*) indicates that SR−/− differed significantly from WT(p< 0.05).

Fig. 2

Altered hippocampal NMDAR and AMPAR expression in GlyT1 +/− mice. (A,B) Postsynaptic density enriched (PSD) and (C,D) whole cell lysate (WCL) hippocampal fractions were collected from adult wild type (WT; black bars; n = 6–12) and mutant (GlyT1 +/−; hatched bars; n = 6–12) mice. The amount of NMDAR subunits (NR1, NR2A, NR2B, postsynaptic density protein of 95kDa (PSD95), and α-amino-3-hydroxyl-5-methyl-4-isoxazole-propionate receptor (AMPAR) subunits (GluR1, GluR2) were quantified by Western blot analysis. Values are expressed as the optical density (OD) normalized to WT values (% control). Bars represent mean values ± s.e.m. Asterisk (*) indicates that GlyT1+/− differed significantly from WT(p< 0.05).