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The anticonvulsant retigabine targets the neuronal M-channel but has adverse clinical effects due to its poor KV7 subtype specificity. Larsson et al. reveal that the selectivity of retigabine is improved by coapplication with the endocannabinoid arachidonoyl-L-serine., Retigabine is unique among anticonvulsant drugs by targeting the neuronal M-channel, which is composed of KV7.2/KV7.3 and contributes to the negative neuronal resting membrane potential. Unfortunately, retigabine causes adverse effects, which limits its clinical use. Adverse effects may be reduced by developing M-channel activators with improved KV7 subtype selectivity. The aim of this study was to evaluate the prospect of endocannabinoids as M-channel activators, either in isolation or combined with retigabine. Human KV7 channels were expressed in Xenopus laevis oocytes. The effect of extracellular application of compounds with different properties was studied using two-electrode voltage clamp electrophysiology. Site-directed mutagenesis was used to construct channels with mutated residues to aid in the mechanistic understanding of these effects. We find that arachidonoyl-L-serine (ARA-S), a weak endocannabinoid, potently activates the human M-channel expressed in Xenopus oocytes. Importantly, we show that ARA-S activates the M-channel via a different mechanism and displays a different KV7 subtype selectivity compared with retigabine. We demonstrate that coapplication of ARA-S and retigabine at low concentrations retains the effect on the M-channel while limiting effects on other KV7 subtypes. Our findings suggest that improved KV7 subtype selectivity of M-channel activators can be achieved through strategically combining compounds with different subtype selectivity.

There exist many different ion channel modulators that inhibit channel function, but few that increase it. Liin et al. tested 18,000 compounds by high-throughput electrophysiology and found several channel openers with a biaryl-sulfonamide motif that acts on the voltage-sensing machinery., Voltage-gated ion channels are key molecules for the generation of cellular electrical excitability. Many pharmaceutical drugs target these channels by blocking their ion-conducting pore, but in many cases, channel-opening compounds would be more beneficial. Here, to search for new channel-opening compounds, we screen 18,000 compounds with high-throughput patch-clamp technology and find several potassium-channel openers that share a distinct biaryl-sulfonamide motif. Our data suggest that the negatively charged variants of these compounds bind to the top of the voltage-sensor domain, between transmembrane segments 3 and 4, to open the channel. Although we show here that biaryl-sulfonamide compounds open a potassium channel, they have also been reported to block sodium and calcium channels. However, because they inactivate voltage-gated sodium channels by promoting activation of one voltage sensor, we suggest that, despite different effects on the channel gates, the biaryl-sulfonamide motif is a general ion-channel activator motif. Because these compounds block action potential–generating sodium and calcium channels and open an action potential–dampening potassium channel, they should have a high propensity to reduce excitability. This opens up the possibility to build new excitability-reducing pharmaceutical drugs from the biaryl-sulfonamide scaffold.

Polyunsaturated fatty acids (PUFAs) affect cardiac excitability. Kv7.1 and the beta-subunit KCNE1 form the cardiac I-Ks channel that is central for cardiac repolarization. In this study, we explore the prospects of PUFAs as I-Ks channel modulators. We report that PUFAs open Kv7.1 via an electrostatic mechanism. Both the polyunsaturated acyl tail and the negatively charged carboxyl head group are required for PUFAs to open Kv7.1. We further show that KCNE1 coexpression abolishes the PUFA effect on Kv7.1 by promoting PUFA protonation. PUFA analogs with a decreased pK(a) value, to preserve their negative charge at neutral pH, restore the sensitivity to open I-Ks channels. PUFA analogs with a positively charged head group inhibit I-Ks channels. These different PUFA analogs could be developed into drugs to treat cardiac arrhythmias. In support of this possibility, we show that PUFA analogs act antiarrhythmically in embryonic rat cardiomyocytes and in isolated perfused hearts from guinea pig. Funding Agencies|National Institutes of Health [R01GM109762]; American Heart Association [14GRNT20380041]; Swedish Research Council; Swedish Heart-Lung Foundation; County Council of Ostergotland; Queen Silvias Anniversary Foundation; Academy of Finland

About 300 loss-of-function mutations in the IKs channel have been identified in patients with Long QT syndrome and cardiac arrhythmia. How specific mutations cause arrhythmia is largely unknown and there are no approved IKs channel activators for treatment of these arrhythmias. We find that several Long QT syndrome-associated IKs channel mutations shift channel voltage dependence and accelerate channel closing. Voltage-clamp fluorometry experiments and kinetic modeling suggest that similar mutation-induced alterations in IKs channel currents may be caused by different molecular mechanisms. Finally, we find that the fatty acid analogue N-arachidonoyl taurine restores channel gating of many different mutant channels, even though the mutations are in different domains of the IKs channel and affect the channel by different molecular mechanisms. N-arachidonoyl taurine is therefore an interesting prototype compound that may inspire development of future IKs channel activators to treat Long QT syndrome caused by diverse IKs channel mutations. DOI: http://dx.doi.org/10.7554/eLife.20272.001, eLife digest Every heartbeat relies on an electric wave that travels through the heart. This wave must reach different parts of the heart in a specific sequence to ensure that the heart muscle cells contract in a coordinated manner. Such coordinated contractions enable the heart to pump enough blood around the body. By allowing specific ions to flow into or out of the heart muscle cell, proteins called ion channels in the cell membrane generate the electric wave, keep it going and stop it. One such protein called the IKs channel controls the flow of potassium ions, and in doing so stops the electric wave in heart muscle cells. About 300 different mutations in the IKs channel have been shown to cause abnormal heart rhythms in individuals with a disorder called long QT syndrome. People with this condition may suddenly black out because their heart develops prolonged electric waves that prevent blood from being pumped properly. To investigate how mutations in the IKs channel produce heart rhythm abnormalities, Liin et al. genetically engineered the egg cells of African clawed frogs to have one of eight mutant forms of the human IKs channel. Studying these channels revealed that the mutations reduce how well the channels work in a wide variety of ways. However, treating the cells with a particular fatty acid helped to normalize how each of the mutant channels worked. Therefore, variants of the fatty acid could potentially form a useful treatment for people with heart rhythm problems caused by mutations in the IKs channel. More studies are needed to confirm whether the fatty acid is as effective at combating the effects of the mutations in whole hearts and animals. As ion channels related to the IKs channel are found in many types of cells, it is also important to investigate whether treatment with the fatty acid could cause any side effects that affect other organs. DOI: http://dx.doi.org/10.7554/eLife.20272.002

We propose that a two-dimensional electric network may be used for fundamental studies of wave function properties, transport, and related statistics. Using Kirchhoff 's current law and the jw-method we find that the network is analogous to a discretized Schrodinger equation for quantum billiards and clots. Thus the complex electric potentials play the role of quantum rnechanical wave functions.

The cardiac I←Ks channel is important for cardiomyocyte repolarization. More than 300 loss-of-function mutations in the genes encoding I←Ks have been identified in patients with Long QT syndrome. These mutations cause cardiac arrhythmias, such as torsades de pointes and ventricular fibrillation. How specific mutations cause arrhythmia is, however, not known in most cases and there is no approved I←Ks channel activator for treatment of arrhythmia. In this work, we study the biophysical properties and mechanism of loss of function of Long QT syndrome-associated I←Ks channel mutations expressed in Xenopus laevis oocytes. We next test the ability of the fatty acid analogue N-arachidonoyl taurine to restore the function of these mutants. Using two-electrode voltage clamp, we find that a number of Long QT syndrome-associated, loss-of-function I←Ks channel mutations shift the voltage dependence of opening towards positive voltages and speed up channel closing. Using voltage clamp fluorometry, we show that these alterations are caused by either disrupted voltage sensor movement or gate opening. We also find that the fatty acid analogue N-arachidonoyl taurine activates all tested Long QT syndrome-associated I←Ks channel mutations by shifting their voltage dependence of activation towards more negative voltages and altering the kinetics of channel opening and closing. Thus, N-arachidonoyl taurine restores the function of these mutants and compensates for the loss of function induced by these mutations, whether the mutations disrupted voltage sensor movement or gate opening. N-AT also shortens a drug-induced prolonged QT interval in isolated perfused guinea pig hearts in Langendorff preparation back to a normal range. N-AT is therefore a promising future I←Ks channel activator that may restore a physiological QT interval by enhancing the function of diverse Long QT syndrome I←Ks mutants, independent of the mutational defect.

Kv7.1 and the β-subunit KCNE1 together form the cardiac IKs channel. IKs has an important role in the cardiac repolarization. In a previous study, we showed that polyunsaturated fatty acids (PUFAs) activate the Kv7.1 channel expressed in Xenopus oocytes via an electrostatic mechanism. However, the pharmacological effect was abolished when Kv7.1 was co-expressed with KCNE1. We suggested that KCNE1 promotes protonation of the PUFAs which thereby lose their charge. PUFAs regained their pharmacological effect if pH of the extracellular solution was increased. How KCNE1 promotes protonation of the PUFAs is however unknown. We hypothesise that charged amino acids in the IKs channel change the local pH in the binding site of PUFA. Plausible amino acids to mediate this effect could be in the extracellular part of KCNE1 or that KCNE1 induces a conformational change in Kv7.1 which moves charged amino acids in Kv7.1 close to the PUFA. To determine the mechanism for KCNE1-induced protonation we use a mutagenesis approach where we primarily focus on acidic and basic residues. We also study how different KCNE subunits affect the sensitivity of Kv7.1 to PUFA. We report that deletion of the extracellular part of KCNE1 does not restore the PUFA sensitivity of IKs channels, suggesting that the extracellular part of KCNE1 is not responsible for the protonation of PUFAs. In contrast, specific point mutations in Kv7.1 alter the pharmacological sensitivity of IKs to PUFA. This suggests that residues in Kv7.1 are important for the PUFA sensitivity of IKs. An understanding of how KCNE1 changes the pharmacological sensitivity of IKs is important for the development of drugs that target IKs.

We propose that a two-dimensional electric network may be used for fundamental studies of wave function properties, transport, and related statistics. Using Kirchhoff's current law and the $j\ensuremath{\omega}$ method we find that the network is analogous to a discretized Schr\"odinger equation for quantum billiards and dots. Thus complex electric potentials play the role of quantum mechanical wave functions. Ways of realizing the electric network are discussed briefly. The role of symmetries is outlined, and a direct way of selecting states with a given symmetry is presented.

Polyunsaturated fatty acids (PUFAs) affect cardiac excitability. Kv7.1 and the β-subunit KCNE1 form the cardiac IKs channel that is central for cardiac repolarization. In this study, we explore the prospects of PUFAs as IKs channel modulators. We report that PUFAs open Kv7.1 via an electrostatic mechanism. Charged n-3 and n-6 PUFAs affect the voltage dependence of Kv7.1 by shifting the conductance versus voltage curve towards more negative voltages. In contrast, uncharged methyl esters of the PUFAs do not affect the voltage dependence of Kv7.1. Both the polyunsaturated acyl tail and the negatively charged carboxyl head group are required for PUFAs to open Kv7.1. The PUFA effect is pH dependent. This is likely because high pH deprotonates the PUFA, making a larger fraction of PUFA molecules negatively charged and thereby able to affect Kv7.1 channel voltage dependence. We further show that KCNE1 co-expression abolishes the PUFA effect on Kv7.1 by promoting PUFA protonation. PUFA analogues with a decreased pKa value, to preserve their negative charge at neutral pH, restore the sensitivity to open IKs channels. PUFA analogues with a positively charged head group inhibit IKs channels. These different PUFA analogues could be developed into drugs to treat cardiac arrhythmias. In support of this possibility, we show that a PUFA analogue with a permanently negatively charged head group acts anti-arrhythmic in cardiomyocytes. This permanently negatively charged PUFA analogue induces a shortening of action potential duration in embryonic rat cardiomyocytes and restores rhythmic beating in an arrhythmia model.

Polyunsaturated fatty acids (PUFAs) affect cardiac excitability by interacting with ion channels. In this work we studied the effect of PUFAs on cardiac Kv7.1 channels. We show that charged n-3 and n-6 PUFAs affect the voltage dependence of Kv7.1 by shifting the conductance versus voltage (G(V)) curve towards hyperpolarized voltages. In contrast, uncharged methyl esters of the PUFAs do not affect the voltage dependence of Kv7.1. The PUFA effect is pH dependent. This is likely because high pH deprotonates the PUFA, making a larger fraction of PUFA molecules negatively charged and able to affect Kv7.1 channel voltage dependence. The structural requirement of PUFAs to induce Kv7.1 channel opening and the PUFA-induced effect on channel voltage dependence were similar to those previously reported for the Shaker K channel. This suggests a general modulatory mechanism in which PUFAs electrostatically interact with the voltage-sensor domain of the channel to induce channel opening. The putative PUFA interaction site on Kv7.1, homologous to the previously reported PUFA interaction site on the Shaker K channel, is close to the position of the auxiliary KCNE1 subunit in the cardiac IKs channel. We therefore also investigated the impact of KCNE1 on PUFA potency and found that the PUFA effect on Kv7.1 was decreased by KCNE1 coexpression. By investigating the mechanism for KCNE1-induced reduction of channel sensitivity to PUFAs, we found that KCNE1 protonates the PUFA molecule making PUFAs uncharged and ineffective. We furthermore explored the possibility to circumvent KCNE1-effects on PUFA potency by testing PUFA analogues with different properties. This study provides mechanistic information on how KCNE1 affects pharmacological sensitivity of the Kv7.1 channel. This study may also form the basis for the development of future Kv7.1 channel openers to be used in the treatment of cardiac arrhythmia.