Subunit-specific roles of NR2A and NR2B in the trigeminal ganglion contribute to orofacial neuropathic pain via distinct molecular cascades.

Orofacial neuropathic pain (ONP) is a common complication following nerve injury driven by peripheral mechanisms. N-methyl-D-aspartate receptors (NMDARs), particularly NR2A and NR2B, are involved in pain processing in the trigeminal ganglion (TG), but their downstream pathways in ONP remain unclear. Here, chronic constriction injury of the infraorbital nerve (CCI-ION) was established in wild-type mice and in mice with TG-targeted adeno-associated virus-mediated conditional deletion of NR2A or NR2B. CCI-ION upregulated NR2A and NR2B in the TG, as verified by quantitative reverse transcription-polymerase chain reaction, Western blot, and immunofluorescence, with increased immunoreactivity in both TG neurons and satellite glial cells (SGCs). Conditional deletion of either subunit attenuated CCI-ION-induced mechanical hypersensitivity. Molecular analyses showed that these subunits were preferentially associated with distinct transcriptional modules: following CCI-ION, NR2A deletion counteracted the suppression of Cyp1a1, Cyp2e1, and Zfp593, whereas NR2B deletion counteracted the upregulation of Slc6a1 and Tlr2 and the downregulation of Cacna1g. N-methyl-D-aspartate (NMDA) stimulation and subsequent NR2A or NR2B knockdown by siRNA produced directionally consistent changes in Neuro-2a cells and primary TG-derived SGCs. Kinase-inhibitor profiling in Neuro-2a cells revealed distinct response patterns. Local pharmacological induction of CYP2E1 or inhibition of TLR2 or SLC6A1 in the TG attenuated CCI-ION-induced mechanical hypersensitivity. Overall, peripheral NMDAR signaling following CCI-ION is associated with subunit-specific transcriptional modules involving both neuronal and glial compartments. These findings identify candidate molecular targets for ONP.