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Key points • We investigated how the F1486 deletion LQT3 mutation impairs the functional properties of the human cardiac voltage-gated sodium channel (hNav1.5) and alters action potential firing. • Voltage-clamp recordings from HEK 293 cells and cardiomyocytes expressing recombinant channels demonstrated that the F1486del mutation reduces peak current density but also impairs inactivation and increases late current density. • Current-clamp recordings from cardiomyocytes indicated that the increase in late current density would result in prolonged action potential duration and this was confirmed using computer simulations. • The deletion of F1486 abolished the ability of lidocaine to stabilize the inactivated state and eliminated the high-affinity binding of lidocaine to inactivated channels. • Our data show that the hNav1.5-F1486del mutation has complex functional consequences and indicate that knowledge of the specific molecular defect is critical when developing potential treatments for individuals with prolonged QT intervals. Abstract The deletion of phenylalanine 1486 (F1486del) in the human cardiac voltage-gated sodium channel (hNav1.5) is associated with fatal long QT (LQT) syndrome. In this study we determined how F1486del impairs the functional properties of hNav1.5 and alters action potential firing in heterologous expression systems (human embryonic kidney (HEK) 293 cells) and their native cardiomyocyte background. Cells expressing hNav1.5-F1486del exhibited a loss-of-function alteration, reflected by an 80% reduction of peak current density, and several gain-of-function alterations, including reduced channel inactivation, enlarged window current, substantial augmentation of persistent late sodium current and an increase in ramp current. We also observed substantial action potential duration (APD) prolongation and prominent early afterdepolarizations (EADs) in neonatal cardiomyocytes expressing the F1486del channels, as well as in computer simulations of myocyte activity. In addition, lidocaine sensitivity was dramatically reduced, which probably contributed to the poor therapeutic outcome observed in the patient carrying the hNav1.5-F1486del mutation. Therefore, despite the significant reduction in peak current density, the F1486del mutation also leads to substantial gain-of-function alterations that are sufficient to cause APD prolongation and EADs, the predominant characteristic of LQTs. These data demonstrate that hNav1.5 mutations can have complex functional consequences and highlight the importance of identifying the specific molecular defect when evaluating potential treatments for individuals with prolonged QT intervals.

Hainantoxin-I is a novel peptide toxin, purified from the venom of the Chinese bird spider Selenocosmia hainana (=Ornithoctonus hainana). It includes 33 amino acid residues with a disulfide linkage of I–IV, II–V and III–VI, assigned by partial reduction and sequence analysis. Under two-electrode voltage-clamp conditions, hainantoxin-I can block rNav1.2/β1 and the insect sodium channel para/tipE expressed in Xenopus laevis oocytes with IC50 values of 68±6 μM and 4.3±0.3 μM respectively. The three-dimensional solution structure of hainantoxin-I belongs to the inhibitor cystine knot structural family determined by two-dimensional 1H nuclear magnetic resonance techniques. Structural comparison of hainantoxin-I with those of other toxins suggests that the combination of the charged residues and a vicinal hydrophobic patch should be responsible for ligand binding. This is the first report of an insect sodium channel blocker from spider venom and it provides useful information for the structure–function relationship studies of insect sodium channels.