BACKGROUND AND PURPOSEFlupirtine is a non-opioid analgesic that has been in clinical use for more than 20 years. It is characterized as a selective neuronal potassium channel opener (SNEPCO). Nevertheless, its mechanisms of action remain controversial and are the purpose of this study. EXPERIMENTAL APPROACHEffects of flupirtine on native and recombinant voltage-and ligand-gated ion channels were explored in patch-clamp experiments using the following experimental systems: recombinant KIR3 and KV7 channels and a3b4 nicotinic acetylcholine receptors expressed in tsA 201 cells; native voltage-gated Na, and TRPV1 channels, as well as GABAA, glycine, and ionotropic glutamate receptors expressed in rat dorsal root ganglion, dorsal horn and hippocampal neurons. KEY RESULTSTherapeutic flupirtine concentrations (Յ10 mM) did not affect voltage-gated Na + or Ca 2+ channels, inward rectifier K + channels, nicotinic acetylcholine receptors, glycine or ionotropic glutamate receptors. Flupirtine shifted the gating of KV7 K + channels to more negative potentials and the gating of GABAA receptors to lower GABA concentrations. These latter effects were more pronounced in dorsal root ganglion and dorsal horn neurons than in hippocampal neurons. In dorsal root ganglion and dorsal horn neurons, the facilitatory effect of therapeutic flupirtine concentrations on KV7 channels and GABAA receptors was comparable, whereas in hippocampal neurons the effects on KV7 channels were more pronounced. CONCLUSIONS AND IMPLICATIONSThese results indicate that flupirtine exerts its analgesic action by acting on both GABAA receptors and KV7 channels. AbbreviationsBMI, bicuculline methiodide; CNQX, cyano-2,3-dihydroxy-7-nitroquinoxaline; DRG, dorsal root ganglion; NBQX, 2,3-dioxo-6-nitro-1,2,3,4-tetrahydrobenzo[f]quinoxaline-7-sulfonamide; SCG, superior cervical ganglion; SNEPCO, selective neuronal potassium channel opener

Neuronal M-channels are low threshold, slowly activating and non-inactivating, voltage dependent K+ channels that play a crucial role in controlling neuronal excitability. The native M-channel is composed of heteromeric or homomeric assemblies of subunits belonging to the Kv7/KCNQ family, with KCNQ2/3 heteromers being the most abundant form. KCNQ2 and KCNQ3 subunits have been found to be expressed in various neurons in the central and peripheral nervous system of rodents and humans. Previous evidence shows preferential localization of both subunits to axon initial segments, somata and nodes of Ranvier. In this work, we show the distribution and co-localization of KCNQ2 and KCNQ3 subunits throughout the hippocampal formation, via immunostaining experiments on unfixed rat brain slices and confocal microscopy. We find intense localization and colocalization to the axonal initial segment in several regions of the hippocampus, as well as staining for non-neuronal cells in the area of the lateral ventricle. We did not observe colocalization of KCNQ2 or KCNQ3 with the presynaptic protein, synaptophysin.

SummaryAdenosine triphosphate and its degradation product adenosine diphosphate excite sensory neurons via 2 different G protein-coupled receptors, P2Y1 and P2Y2, which mediate inhibition KV7 and sensitization of TRPV1 channels.

Background and PurposeThe Kv7 channel activator flupirtine is a clinical analgesic characterized as ‘selective neuronal potassium channel opener’. Flupirtine was found to exert comparable actions at GABAA receptors and Kv7 channels in neurons of pain pathways, but not in hippocampus.Experimental ApproachExpression patterns of GABAA receptors were explored in immunoblots of rat dorsal root ganglia, dorsal horns and hippocampi using antibodies for 10 different subunits. Effects of flupirtine on recombinant and native GABAA receptors were investigated in patch clamp experiments and compared with the actions on Kv7 channels.Key ResultsImmunoblots pointed towards α2, α3, β3 and γ2 subunits as targets, but in all γ2-containing receptors the effects of flupirtine were alike: leftward shift of GABA concentration-response curves and diminished maximal amplitudes. After replacement of γ2S by δ, flupirtine increased maximal amplitudes. Currents through α1β2δ receptors were more enhanced than those through Kv7 channels. In hippocampal neurons, flupirtine prolonged inhibitory postsynaptic currents, left miniature inhibitory postsynaptic currents (mIPSCs) unaltered and increased bicuculline-sensitive tonic currents; penicillin abolished mIPSCs, but not tonic currents; concentration-response curves for GABA-induced currents were shifted to the left by flupirtine without changes in maximal amplitudes; in the presence of penicillin, maximal amplitudes were increased; GABA-induced currents in the presence of penicillin were more sensitive towards flupirtine than K+ currents. In dorsal horn neurons, currents evoked by the δ-preferring agonist THIP (gaboxadol) were more sensitive towards flupirtine than K+ currents.Conclusions and ImplicationsFlupirtine prefers δ-containing GABAA receptors over γ-containing ones and over Kv7 channels.

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