Modern pain-control theory predicts that a loss of inhibition (disinhibition) in the dorsal horn of the spinal cord is a crucial substrate for chronic pain syndromes. However, the nature of the mechanisms that underlie such disinhibition has remained controversial. Here we present evidence for a novel mechanism of disinhibition following peripheral nerve injury. It involves a trans-synaptic reduction in the expression of the potassium-chloride exporter KCC2, and the consequent disruption of anion homeostasis in neurons of lamina I of the superficial dorsal horn, one of the main spinal nociceptive output pathways. In our experiments, the resulting shift in the transmembrane anion gradient caused normally inhibitory anionic synaptic currents to be excitatory, substantially driving up the net excitability of lamina I neurons. Local blockade or knock-down of the spinal KCC2 exporter in intact rats markedly reduced the nociceptive threshold, confirming that the reported disruption of anion homeostasis in lamina I neurons was sufficient to cause neuropathic pain.

The K+-Cl− cotransporter KCC2 is responsible for maintaining low Cl− concentration in neurons of the central nervous system (CNS), essential for postsynaptic inhibition through GABAA and glycine receptors. While no CNS disorders have been associated with KCC2 mutations, loss of activity of this transporter has emerged as a key mechanism underlying several neurological and psychiatric disorders including epilepsy, motor spasticity, stress, anxiety, schizophrenia, morphine-induced hyperalgesia and chronic pain1–9. Recent reports indicate that enhancing KCC2 activity may be the favoured therapeutic strategy to restore inhibition and normal function in pathological condition involving impaired Cl− transport10–12. We designed an assay for high-throughput screening which led to the identification of KCC2 activators that reduce [Cl−]i. Optimization of a first-in-class arylmethylidine family of compounds resulted in a KCC2-selective analog (CLP257) that lowers [Cl−]i. CLP257 restored impaired Cl− transport in neurons with diminished KCC2 activity. The compound rescued KCC2 plasma membrane expression, renormalised stimulus-evoked responses in spinal nociceptive pathways sensitized after nerve injury and alleviated hypersensitivity in a rat model of neuropathic pain. Oral efficacy for analgesia equivalent to that of Pregabalin but without motor impairment was achievable with a CLP257 prodrug. These results validate KCC2 as a druggable target for CNS diseases.

Spinal disinhibition has been hypothesized to underlie pain hypersensitivity in neuropathic pain. Apparently contradictory mechanisms have been reported, raising questions on the best target to produce analgesia. Here, we show that nerve injury is associated with a reduction in the number of inhibitory synapses in the spinal dorsal horn. Paradoxically, this is accompanied by a BDNF-TrkB-mediated upregulation of synaptic GABA A Rs and by an α1-to-α2GABA A R subunit switch, providing a mechanistic rationale for the analgesic action of the α2,3GABA A R benzodiazepine-site ligand L838,417 after nerve injury. Yet, we demonstrate that impaired Clextrusion underlies the failure of L838,417 to induce analgesia at high doses due to a resulting collapse in Clgradient, dramatically limiting the benzodiazepine therapeutic window. In turn, enhancing KCC2 activity not only potentiated L838,417-induced analgesia, it rescued its analgesic potential at high doses, revealing a novel strategy for analgesia in pathological pain, by combined targeting of the appropriate GABA A Rsubtypes and restoring Clhomeostasis.

Summary Trigeminal neuralgia (TN) is a common, debilitating neuropathic face pain syndrome often resistant to therapy. The familial clustering of TN cases suggests that genetic factors play a role in disease pathogenesis. However, no unbiased, large-scale genomic study of TN has been performed to date. Analysis of 290 whole exome-sequenced TN probands, including 20 multiplex kindreds and 70 parent-offspring trios, revealed enrichment of rare, damaging variants in GABA receptor-binding genes in cases. Mice engineered with a TN-associated de novo mutation (p.Cys188Trp) in the GABA A receptor Cl − channel γ-1 subunit ( GABRG1 ) exhibited trigeminal mechanical allodynia and face pain behavior. Other TN probands harbored rare damaging variants in Na + and Ca + channels, including a significant variant burden in the α-1H subunit of the voltage-gated Ca 2+ channel Ca v 3.2 ( CACNA1H ). These results provide exome-level insight into TN and implicate genetically encoded impairment of GABA signaling and neuronal ion transport in TN pathogenesis.