The α2δ-1-NMDA Receptor Complex Is Critically Involved in Neuropathic Pain Development and Gabapentin Therapeutic Actions.

α2δ-1, commonly known as a voltage-activated Ca2+ channel subunit, is a binding site of gabapentinoids used to treat neuropathic pain and epilepsy. However, it is unclear how α2δ-1 contributes to neuropathic pain and gabapentinoid actions. Here, we show that Cacna2d1 overexpression potentiates presynaptic and postsynaptic NMDAR activity of spinal dorsal horn neurons to cause pain hypersensitivity. Conversely, Cacna2d1 knockdown or ablation normalizes synaptic NMDAR activity increased by nerve injury. α2δ-1 forms a heteromeric complex with NMDARs in rodent and human spinal cords. The α2δ-1-NMDAR interaction predominantly occurs through the C terminus of α2δ-1 and promotes surface trafficking and synaptic targeting of NMDARs. Gabapentin or an α2δ-1 C terminus-interfering peptide normalizes NMDAR synaptic targeting and activity increased by nerve injury. Thus, α2δ-1 is an NMDAR-interacting protein that increases NMDAR synaptic delivery in neuropathic pain. Gabapentinoids reduce neuropathic pain by inhibiting forward trafficking of α2δ-1-NMDAR complexes.

A conserved structural mechanism of NMDA receptor inhibition: A comparison of ifenprodil and zinc.

N-methyl-D-aspartate (NMDA) receptors, one of the three main types of ionotropic glutamate receptors (iGluRs), are involved in excitatory synaptic transmission, and their dysfunction is implicated in various neurological disorders. NMDA receptors, heterotetramers typically composed of GluN1 and GluN2 subunits, are the only members of the iGluR family that bind allosteric modulators at their amino-terminal domains (ATDs). We used luminescence resonance energy transfer to characterize the conformational changes the receptor undergoes upon binding ifenprodil, a synthetic compound that specifically inhibits activation of NMDA receptors containing GluN2B. We found that ifenprodil induced an overall closure of the GluN2B ATD without affecting conformation of the GluN1 ATD or the upper lobes of the ATDs, the same mechanism whereby zinc inhibits GluN2A. These data demonstrate that the conformational changes induced by zinc and ifenprodil represent a conserved mechanism of NMDA receptor inhibition. Additionally, we compared the structural mechanism of zinc inhibition of GluN1-GluN2A receptors to that of ifenprodil inhibition of GluN1-GluN2B. The similarities in the conformational changes induced by inhibitor binding suggest a conserved structural mechanism of inhibition independent of the binding site of the modulator.

Subtype-dependent N-methyl-D-aspartate receptor amino-terminal domain conformations and modulation by spermine.

The N-methyl-d-aspartate (NMDA) subtype of the ionotropic glutamate receptors is the primary mediator of calcium-permeable excitatory neurotransmission in the central nervous system. Subunit composition and binding of allosteric modulators to the amino-terminal domain determine the open probability of the channel. By using luminescence resonance energy transfer with functional receptors expressed in CHO cells, we show that the cleft of the amino-terminal domain of the GluN2B subunit, which has a lower channel open probability, is on average more closed than the GluN2A subunit, which has a higher open probability. Furthermore, the GluN1 amino-terminal domain adopts a more open conformation when coassembled with GluN2A than with GluN2B. Binding of spermine, an allosteric potentiator, opens the amino-terminal domain cleft of both the GluN2B subunit and the adjacent GluN1 subunit. These studies provide direct structural evidence that the inherent conformations of the amino-terminal domains vary based on the subunit and match the reported open probabilities for the receptor.

Amino-terminal domain tetramer organization and structural effects of zinc binding in the N-methyl-D-aspartate (NMDA) receptor.

N-Methyl-D-aspartate (NMDA) receptors mediate excitatory neurotransmission in the mammalian central nervous system. An important feature of these receptors is their capacity for allosteric regulation by small molecules, such as zinc, which bind to their amino-terminal domain (ATD). Zinc inhibition through high affinity binding to the ATD has been examined through functional studies; however, there is no direct measurement of associated conformational changes. We used luminescence resonance energy transfer to show that the ATDs undergo a cleft closure-like conformational change upon binding zinc, but no changes are observed in intersubunit distances. Furthermore, we find that the ATDs are more closely packed than the related AMPA receptors. These results suggest that the stability of the upper lobe contacts between ATDs allow for the efficient propagation of the cleft closure conformational change toward the ligand-binding domain and transmembrane segments, ultimately inhibiting the channel.