Kv7.2 subunits encoded by the KCNQ2 gene provide a major contribution to the M-current (I_KM), a voltage-gated K⁺ current crucially involved in the regulation of neuronal excitability. Heterozygous missense variants in Kv7.2 are responsible for epileptic diseases characterized by highly heterogeneous genetic transmission and clinical severity, ranging from autosomal-dominant Benign Familial Neonatal Seizures (BFNS) to sporadic cases of severe epileptic and developmental encephalopathy (DEE). Here, we describe a patient with neonatal onset DEE, carrying a previously undescribed heterozygous KCNQ2 c.418G > C, p.Glu140Gln (E140Q) variant. Patch-clamp recordings in CHO cells expressing the E140Q mutation reveal dramatic loss of function (LoF) effects. Multistate structural modelling suggested that the E140Q substitution impeded an intrasubunit electrostatic interaction occurring between the E140 side chain in S₂ and the arginine at position 210 in S₄ (R210); this interaction is critically involved in stabilizing the activated configuration of the voltage-sensing domain (VSD) of Kv7.2. Functional results from coupled charge reversal or disulfide trapping experiments supported such a hypothesis. Finally, retigabine restored mutation-induced functional changes, reinforcing the rationale for the clinical use of Kv7 activators as personalized therapy for DEE-affected patients carrying Kv7.2 LoF mutations.

Kv7.2-Kv7.5 channels mediate the M-current (I_KM), a K⁺-selective current regulating neuronal excitability and representing an attractive target for pharmacological therapy against hyperexcitability diseases such as pain. Kv7 channels interact functionally with transient receptor potential vanilloid 1 (TRPV1) channels activated by endogenous and/or exogenous pain-inducing substances, such as bradykinin (BK) or capsaicin (CAP), respectively; however, whether Kv7 channels of specific molecular composition provide a dominant contribution in BK- or CAP-evoked responses is yet unknown. To this aim, Kv7 transcripts expression and function were assessed in F11 immortalized sensorial neurons, a cellular model widely used to assess nociceptive molecular mechanisms. In these cells, the effects of the pan-Kv7 activator retigabine were investigated, as well as the effects of ICA-27243 and (S)-1, two Kv7 activators acting preferentially on Kv7.2/Kv7.3 and Kv7.4/Kv7.5 channels, respectively, on BK- and CAP-induced changes in intracellular Ca²⁺ concentrations ([Ca²⁺]_i). The results obtained revealed the expression of transcripts of all Kv7 genes, leading to an I_KM-like current. Moreover, all tested Kv7 openers inhibited BK- and CAP-induced responses by a similar extent (~60%); at least for BK-induced Ca²⁺ responses, the potency of retigabine (IC₅₀~1 µM) was higher than that of ICA-27243 (IC₅₀~5 µM) and (S)-1 (IC₅₀~7 µM). Altogether, these results suggest that I_KM activation effectively counteracts the cellular processes triggered by TRPV1-mediated pain-inducing stimuli, and highlight a possible critical contribution of Kv7.4 subunits.