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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.

Lipids in different forms can directly affect the electric activity of excitable tissues, in some cases via direct interactions with ion channels. For instance is the activity of Kv7 (KCNQ) channels dependent on the abundance of the phospholipid PIP2 in the inner leaflet of the plasma membrane. In this work we instead studied the effect of free extracellular polyunsaturated fatty acids (PUFAs) on Kv7 channels expressed in Xenopus oocytes. We show that ω-3 and ω-6 PUFAs affected the voltage dependence of Kv7.1 and Kv7.2/3 by shifting the conductance versus voltage (G(V)) curves in negative direction along the voltage axis. The effect was pH dependent. In contrast, uncharged methyl esters of the PUFAs did not affect the voltage dependences. Fatty acid requirements and PUFA-induced effects were similar to those previously reported for the Shaker K channel suggesting a similar modulatory mechanism of action, i.e. our previously proposed lipoelectric mechanism in which PUFAs electrostatically affect channel opening. The putative PUFA interaction site is close to the positioning of auxiliary KCNE subunits. We therefore also investigated the impact of KCNE subunits and found that PUFA potency was influenced by KCNE association. Kv7.1 and Kv7.2/3 channels are important for cardiac and neuronal excitability, respectively, and mutated channels resulting in loss-of-function cause heart arrhythmias and epilepsy. The PUFA-induced opening of Kv7 channels found in this study may therefore help explain the antiexcitable properties of PUFAs and the fat-rich ketogenic diet. Lipoelectric modification of the channels voltage dependence could be a future new approach for pharmacological treatment.

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.