Retigabine and gabapentin restore channel function and neuronal firing of an epilepsy-associated dominant-negativeKCNQ5variant

Abstract

Objective:KCNQ5encodes the voltage-gated potassium channel KV7.5, a member of the KV7 channel family, which conducts the M-current. This current was shown to be a potent regulator of neuronal excitability by mediating the medium and slow afterhyperpolarization. Recently, we have identified five loss-of-function variants inKCNQ5in patients with genetic generalized epilepsy. Using the most severe dominant-negative variant p.(Arg359Cys) (R359C), we set out to investigate pharmacological therapeutic intervention by KV7 channel openers on channel function and neuronal firing. Methods: Whole-cell patch clamp recordings were conducted in human embryonic kidney cells to investigate the immediate effect of retigabine, gabapentin and intracellular application of zinc on the R359C variant in absence and presence of KV7.5-WT subunits. Transfected primary hippocampal cultures were used to examine the effect of R359C on neuronal firing and whether this effect could be reversed by drug application. Results: Retigabine and gabapentin both increased R359C-derived K+current density and M-current amplitudes in both homomeric and heteromeric mutant KV7.5 channels. Retigabine was most effective in restoring K+currents. Ten μM retigabine was sufficient to reach the level of WT currents without retigabine, whereas 100 μM of gabapentin showed less than half of this effect and application of 50 μM zinc only significantly increased M-current amplitude in heteromeric channels. Overexpression of KV7.5-WT potently inhibited neuronal firing by increasing the M-current, and medium afterhyperpolarization, whereas R359C overexpression had the opposite effect. All three aforementioned drugs reversed the effect of R359C reducing firing to nearly normal levels at high current injections. Significance: Our study shows that a dominant-negative complete loss-of-function variant in KV7.5 leads to largely increased neuronal firing indicating a neuronal hyperexcitability. KV7 channel openers, such as retigabine or gabapentin, could be treatment options for otherwise pharmacoresistant epilepsy patients carrying loss-of-function variants inKCNQ5.