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Dapagliflozin (dapa) and empagliflozin (empa) are sodium-glucose cotransporter-2 inhibitors (SGLT2is) that reduce morbidity and mortality in heart failure (HF) patients. Sodium and inward rectifier K+ currents (INa and IK1), carried by Nav1.5 and Kir2.1 channels, respectively, are responsible for cardiac excitability, conduction velocity, and refractoriness. In HF patients, Nav1.5 and Kir2.1 expression are reduced, enhancing risk of arrhythmia. Incubation with dapa or empa (24-h,1 µM) significantly increased INa and IK1 densities recorded in human-induced pluripotent stem cell-cardiomyocytes (hiPSC-CMs) using patch-clamp techniques. Dapa and empa, respectively, shifted to more hyperpolarized potentials the INa activation and inactivation curves. Identical effects were observed in Chinese hamster ovary (CHO) cells that were incubated with dapa or empa and transiently expressed human Nav1.5 channels. Conversely, empa but not dapa significantly increased human Kir2.1 currents in CHO cells. Dapa and empa effects on INa and IK1 were also apparent in Ca-calmodulin kinase II-silenced CHO cells. Cariporide, a Na+/H+ exchanger type 1 (NHE1) inhibitor, did not increase INa or IK1 in hiPSC-CMs. Dapa and empa at therapeutic concentrations increased INa and IK1 in healthy human cardiomyocytes. These SGLT2is could represent a new class of drugs with a novel and long-pursued antiarrhythmic mechanism of action.

Synapse-Associated Protein 97 (SAP97) is an anchoring protein that in cardiomyocytes targets to the membrane and regulates Na + and K + channels. Here we compared the electrophysiological effects of native (WT) and p.P888L SAP97, a common polymorphism. Currents were recorded in cardiomyocytes from mice trans-expressing human WT or p.P888L SAP97 and in Chinese hamster ovary (CHO)-transfected cells. The duration of the action potentials and the QT interval were significantly shorter in p.P888L-SAP97 than in WT-SAP97 mice. Compared to WT, p.P888L SAP97 significantly increased the charge of the Ca-independent transient outward (I to,f ) current in cardiomyocytes and the charge crossing Kv4.3 channels in CHO cells by slowing Kv4.3 inactivation kinetics. Silencing or inhibiting Ca/calmodulin kinase II (CaMKII) abolished the p.P888L-induced Kv4.3 charge increase, which was also precluded in channels (p.S550A Kv4.3) in which the CaMKII-phosphorylation is prevented. Computational protein-protein docking predicted that p.P888L SAP97 is more likely to form a complex with CaMKII than WT. The Na + current and the current generated by Kv1.5 channels increased similarly in WT-SAP97 and p.P888L-SAP97 cardiomyocytes, while the inward rectifier current increased in WT-SAP97 but not in p.P888L-SAP97 cardiomyocytes. The p.P888L SAP97 polymorphism increases the I to,f , a CaMKII-dependent effect that may increase the risk of arrhythmias.

The ZFHX3 and SCN5A genes encode the zinc finger homeobox 3 (Zfhx3) transcription factor (TF) and the human cardiac Na+ channel (Nav1.5), respectively. The effects of Zfhx3 on the expression of the Nav1.5 channel, and in cardiac excitability, are currently unknown. Additionally, we identified three Zfhx3 variants in probands diagnosed with familial atrial fibrillation (p.M1260T) and Brugada Syndrome (p.V949I and p.Q2564R). Here, we analyzed the effects of native (WT) and mutated Zfhx3 on Na+ current (INa) recorded in HL-1 cardiomyocytes. ZFHX3 mRNA can be detected in human atrial and ventricular samples. In HL-1 cardiomyocytes, transfection of Zfhx3 strongly reduced peak INa density, while the silencing of endogenous expression augmented it (from −65.9 ± 8.9 to −104.6 ± 10.8 pA/pF; n ≥ 8, p < 0.05). Zfhx3 significantly reduced the transcriptional activity of human SCN5A, PITX2, TBX5, and NKX25 minimal promoters. Consequently, the mRNA and/or protein expression levels of Nav1.5 and Tbx5 were diminished (n ≥ 6, p < 0.05). Zfhx3 also increased the expression of Nedd4-2 ubiquitin-protein ligase, enhancing Nav1.5 proteasomal degradation. p.V949I, p.M1260T, and p.Q2564R Zfhx3 produced similar effects on INa density and time- and voltage-dependent properties in WT. WT Zfhx3 inhibits INa as a result of a direct repressor effect on the SCN5A promoter, the modulation of Tbx5 increasing on the INa, and the increased expression of Nedd4-2. We propose that this TF participates in the control of cardiac excitability in human adult cardiac tissue.

A novel rare mutation in the pore region of Nav1.5 channels (p.L889V) has been found in three unrelated Spanish families that produces quite diverse phenotypic manifestations (Brugada syndrome, conduction disease, dilated cardiomyopathy, sinus node dysfunction, etc.) with variable penetrance among families. We clinically characterized the carriers and recorded the Na+ current (INa) generated by p.L889V and native (WT) Nav1.5 channels, alone or in combination, to obtain further insight into the genotypic–phenotypic relationships in patients carrying SCN5A mutations and in the molecular determinants of the Nav1.5 channel function. The variant produced a strong dominant negative effect (DNE) since the peak INa generated by p.L889V channels expressed in Chinese hamster ovary cells, either alone (−69.4 ± 9.0 pA/pF) or in combination with WT (−62.2 ± 14.6 pA/pF), was significantly (n ≥ 17, p < 0.05) reduced compared to that generated by WT channels alone (−199.1 ± 44.1 pA/pF). The mutation shifted the voltage dependence of channel activation and inactivation to depolarized potentials, did not modify the density of the late component of INa, slightly decreased the peak window current, accelerated the recovery from fast and slow inactivation, and slowed the induction kinetics of slow inactivation, decreasing the fraction of channels entering this inactivated state. The membrane expression of p.L889V channels was low, and in silico molecular experiments demonstrated profound alterations in the disposition of the pore region of the mutated channels. Despite the mutation producing a marked DNE and reduction in the INa and being located in a critical domain of the channel, its penetrance and expressivity are quite variable among the carriers. Our results reinforce the argument that the incomplete penetrance and phenotypic variability of SCN5A loss-of-function mutations are the result of a combination of multiple factors, making it difficult to predict their expressivity in the carriers despite the combination of clinical, genetic, and functional studies.