Ancient and modern anticonvulsants act synergistically in a KCNQ potassium channel binding pocket

Rían W. Manville,Geoffrey W. Abbott

doi:10.1038/s41467-018-06339-2

Abstract

Epilepsy has been treated for centuries with herbal remedies, including leaves of the African shrub Mallotus oppositifolius, yet the underlying molecular mechanisms have remained unclear. Voltage-gated potassium channel isoforms KCNQ2–5, predominantly KCNQ2/3 heteromers, underlie the neuronal M-current, which suppresses neuronal excitability, protecting against seizures. Here, in silico docking, mutagenesis and cellular electrophysiology reveal that two components of M. oppositifolius leaf extract, mallotoxin (MTX) and isovaleric acid (IVA), act synergistically to open neuronal KCNQs, including KCNQ2/3 channels. Correspondingly, MTX and IVA combine to suppress pentylene tetrazole-induced tonic seizures in mice, whereas individually they are ineffective. Co-administering MTX and IVA with the modern, synthetic anticonvulsant retigabine creates a further synergy that voltage independently locks KCNQ2/3 open. Leveraging this synergy, which harnesses ancient and modern medicines to exploit differential KCNQ isoform preferences, presents an approach to developing safe yet effective anticonvulsants.

Highlights

Epilepsy has been treated for centuries with herbal remedies, including leaves of the African shrub Mallotus oppositifolius, yet the underlying molecular mechanisms have remained unclear
Extract taken from leaves of the shrub Mallotus oppositifolius has been used traditionally in folk medicine to treat disorders such as seizures in African countries[4,5] including Ghana, where it is known as nyanyaforowa, and Nigeria, where it is referred to as okpo-biriba
Using two-electrode voltage-clamp electrophysiology, we screened all ten compounds for their ability to activate heterologously expressed KCNQ2/3 channels in Xenopus laevis oocytes, with the exception of valeric acid, which we had previously found to be inactive in this respect[28] (Fig. 1d)

Summary

Results

Multiple M. oppositifolius leaf compounds activate KCNQ2/3. Igwe et al.[26] recently identified by mass spectrometry nine primary components of an ethanolic extract of the M. oppositifolius leaf, in addition to MTX, which was previously identified in M. oppositifolius bark and leaves[27] (Fig. 1c). Four components—glutaconic acid, isovaleric acid (IVA), MTX, and palmitic acid, negativeshifted the voltage dependence of KCNQ2/3 activation, as quantified using KCNQ2/3 tail currents at −30 mV immediately following channel activation at voltages between −80 mV and +40 mV (Fig. 1e, f). Three of these compounds (MTX excepted) possessed strong negative electrostatic surface potential close to a carbonyl oxygen, a property previously shown important for KCNQ2/3 activation by RTG and related synthetic anticonvulsants[29]. MTX was an effective KCNQ5 opener a b

Ctrl 100 μM

Discussion

Methods