Introduction: Peripheral neuropathic pain, typified by the development of spontaneous pain or pain hypersensitivity following injury to the peripheral nervous system, is common, greatly impairs quality of life, and is inadequately treated with available drugs. Maladaptive changes in Cl- homeostasis due to a decrease in the functional expression of the K+-Cl- cotransporter KCC2 in spinal cord dorsal horn neurons are a major contributor to the central disinhibition of gamma-aminobutyric acid type A (GABA-A) receptor characterizes neuropathic pain. A compelling novel analgesic strategy is to restore spinal GABA-mediated ionotropic inhibition by enhancing KCC2-mediated Cl- extrusion.

Methods: We approached this problem by performing a high-content functional genetic siRNA screen looking for molecules required for a potent inhibitory phosphorylation switch in the KCCs, including KCC2, to find targets that when inhibited, activate KCC2. We utilized biochemistry with phospho-specific antibodies targeting this inhibitory regulatory motif to confirm hits with other shRNAs, or in knock-out ES cell lines. We then tested top hits using Rb86 flux and/or electrophysiological assays (gramicidin perforated patch clamp) for KCC activation capacity in cells and neurons. We tested genetic and pharmacological inhibition of one of our top-scoring hits (WNK-SPAK kinase complex) in the spared nerve injury (SNI) model of neuropathic pain.

Results: We have identified a highly-druggable signaling node comprised of the WNK-SPAK kinases that inhibit the KCCs via phosphorylation, and when genetically or pharmacologically inhibited, activate K-Cl cotransport to facilitate Cl- extrusion, normalize EGABA, and/or normalize behavior in the SNI model of pain in rats.

Conclusions: Exploiting the Cl--dependent functional plasticity of GABAARs by activating KCC2 via WNK-SPAK kinase inhibition may be a tenable method of restoring ionotropic inhibition in neuropathic pain, and potentially other "hyperexcitable" diseases of the nervous system, such as seizures and spasticity.

Patient Care: We have identified a novel strategy to restore ionotropic inhibition downstream of GABA signaling to combat the hyperexcitability of injured pain circuits; since inhibitors of the WNK-SPAK kinase cascade have recently been developed, this strategy has potential pharmacotherapeutic value

Learning Objectives: 1. To appreciate that neuropathic pain is a large area of unmet clinical need 2. To understand the role of GABA and Cl- homeostasis via KCC2 in neuropathic pain pathophysiology 3. To appreciate a novel molecular pharmacotherapeutic strategy aimed at restoring iontropic inhibition in neuropathic pain

References: Note: this research was funded by the 2013 CNS Christopher C. Getch Award, given to K. Kahle.