Involvement of spinal phosphorylation cascade of Tyr1472-NR2B, Thr286-CaMKII, and Ser831-GluR1 in neuropathic pain

Previously we demonstrated that phosphorylation of NR2B subunits of the N-methyl-d-aspartate (NMDA) glutamate receptor at Tyr1472 is increased in a neuropathic-pain model and that this phosphorylation is required for the maintenance of neuropathic pain by L5-spinal nerve transection. We obtained these results by using a selective NR2B antagonist and mice deficient in Fyn, which is an Src-family tyrosine protein kinase. However, how Tyr1472 phosphorylation of NR2B is involved in the maintenance of neuropathic pain was unclear. Here, we demonstrated that neuropathic pain was markedly attenuated in the spared nerve injury model of mice with a knock-in mutation of the Tyr1472 site to phenylalanine of NR2B (Y1472F-KI). While phosphorylation of Ca2+/calmodulin-dependent protein kinase II (CaMKII) at its Thr286 and that of the GluR1 subunit of the AMPA receptor at its Ser831 was enhanced in the spinal dorsal horn after spared nerve injury in wild-type mice, such phosphorylation was markedly impaired in Y1472F-KI mice. Inhibition of CaMKII by intrathecal injection of KN93, an inhibitor of CaMKII, reduced mechanical allodynia and phosphorylation of CaMKII at its Thr286 and that of GluR1 at its Ser831 in the spinal cord 7 days after spared nerve injury. These results demonstrate that the phosphorylation of CaMKII and GluR1 occurs downstream of the Tyr1472 phosphorylation of NR2B subunits in the spinal cord and give the first suggestion that activation of CaMKII and GluR1-AMPA receptors may be involved in mechanical allodynia caused by peripheral nerve injury.

Research highlights
► The mice with a mutation of the Tyr1472 to Phe of NR2B (Y1472F-KI) were analyzed.
► We applied a neuropathic-pain model, spared nerve injury (SNI) to Y1472F-KI mice.
► Phosphorylation cascade of Y1472-NR2B, T286-CaMKII and S831-GluR1 occurs by SNI.
► Allodynia by SNI is reduced in Y1472F-KI mice and by i.t. KN93, CaMKII inhibitor.