Quinoline-related antimalarial drugs have been associated with cardiotoxicity risk, in particular QT prolongation and QRS complex widening. In collaboration with Medicines for Malaria Venture (MMV), we discovered novel plasmepsin X (PMX) inhibitors for malaria treatment. The first lead compounds tested in anesthetized guinea pigs (GP) induced profound QRS widening, although exhibiting weak inhibition of NaV1.5-mediated currents in standard patch clamp assays. To understand the mechanism(s) underlying QRS widening to identify further compounds devoid of such liability, we established a set of in vitro models including CaV1.2, NaV1.5 rate-dependence and NaV1.8 patch clamp assays, human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CM), and Langendorff-perfused isolated GP hearts. Six compounds were tested in all models including anesthetized GP, and 8 additional compounds were tested in vitro only. All compounds tested in anesthetized GP and isolated hearts showed a similar cardiovascular profile, consisting of QRS widening, bradycardia, negative inotropy, hypotension, and for some, QT prolongation. However, a left shift of the concentration-response curves was noted from in vitro to in vivo GP data. When comparing in vitro models, there was a good consistency between decrease in sodium spike amplitude in hiPSC-CM and QRS widening in isolated hearts. Patch clamp assay results showed that the QRS widening observed with PMX inhibitors is likely multifactorial, primarily due to NaV1.8 and NaV1.5 rate-dependent sodium blockade and/or calcium channel-mediated mechanisms. In conclusion, early de-risking of QRS widening using a set of different in vitro assays allowed to identify novel PMX inhibitors with improved cardiac safety profile.

In different areas of the heart, action potential waveforms differ due to differences in the expressions of sodium, calcium, and potassium channels. One of the characteristics of myocardial infarction (MI) is an imbalance in oxygen supply and demand, leading to ion imbalance. After MI, the regulation and expression levels of K+, Ca2+, and Na+ ion channels in cardiomyocytes are altered, which affects the regularity of cardiac rhythm and leads to myocardial injury. Myocardial fibroblasts are the main effector cells in the process of MI repair. The ion channels of myocardial fibroblasts play an important role in the process of MI. At the same time, a large number of ion channels are expressed in immune cells, which play an important role by regulating the in- and outflow of ions to complete intracellular signal transduction. Ion channels are widely distributed in a variety of cells and are attractive targets for drug development. This article reviews the changes in different ion channels after MI and the therapeutic drugs for these channels. We analyze the complex molecular mechanisms behind myocardial ion channel regulation and the challenges in ion channel drug therapy.

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BACKGROUND: Variants of the SCN5A gene, which encodes the NaV1.5 cardiac sodium channel, have been linked to arrhythmic disorders associated with dilated cardiomyopathy (DCM). However, the precise pathological mechanisms remain elusive. The present study aimed to elucidate the pathophysiological consequences of the DCM-linked Nav1.5/R219H variant, which is known to generate a gating pore current, using patient-specific human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) cultured in monolayers.
METHODS: Ventricular- and atrial-like hiPSC-CM monolayers were generated from DCM patients carrying the R219H SCN5A variant as well as from healthy control individuals. CRISPR-corrected hiPSC-CMs served as isogenic controls. Simultaneous optical mapping of action potentials (APs) and calcium transients (CaTs) was employed to measure conduction velocities (CVs) and AP durations (APDs) and served as markers of electrical excitability. Calcium handling was evaluated by assessing CaT uptake (half-time to peak), recapture (tau of decay), and durations (TD50 and TD80). A multi-electrode array (MEA) analysis was conducted on hiPSC-CM monolayers to measure field potential (FP) parameters, including corrected Fridericia FP durations (FPDc).
RESULTS: Our results revealed that CVs were significantly reduced by more than 50 % in both ventricular- and atrial-like hiPSC-CM monolayers carrying the R219H variant compared to the control group. APDs were also prolonged in the R219H group compared to the control and CRISPR-corrected groups. CaT uptake, reuptake, and duration were also markedly delayed in the R219H group compared to the control and CRISPR-corrected groups in both the ventricular- and the atrial-like hiPSC-CM monolayers. Lastly, the MEA data revealed a notably prolonged FPDc in the ventricular- and atrial-like hiPSC-CMs carrying the R219H variant compared to the control and isogenic control groups.
CONCLUSIONS: These findings highlight the impact of the gating pore current on AP propagation and calcium homeostasis within a functional syncytium environment and offer valuable insights into the potential mechanisms underlying DCM pathophysiology.

Pufferfish saxitoxin- and tetrodotoxin (TTX)-binding protein (PSTBP) is considered to transfer TTX between tissues. The immunohistochemical distribution of PSTBP-homolog (PSTBPh) and TTX in the brain and pituitary of hatchery-reared juvenile tiger puffer Takifugu rubripes was investigated. PSTBPh was observed mainly in the pars intermedia of the pituitary. TTX was only detected in a TTX-fed fish in the neurohypophysis of the pituitary and in several other brain regions. The relationship between PSTBPh and TTX is discussed.

Epilepsies have numerous specific mechanisms. The understanding of neural dynamics leading to seizures is important for disclosing pathological mechanisms and developing therapeutic approaches. We investigated electrographic activities and neural dynamics leading to convulsive seizures in patients and mouse models of Dravet syndrome (DS), a developmental and epileptic encephalopathy in which hypoexcitability of GABAergic neurons is considered to be the main dysfunction. We analyzed EEGs from DS patients carrying a SCN1A pathogenic variant, as well as epidural electrocorticograms, hippocampal local field potentials, and hippocampal single-unit neuronal activities in Scn1a+/- and Scn1aRH/+ DS mice. Strikingly, most seizures had low-voltage-fast onset in both patients and mice, which is thought to be generated by hyperactivity of GABAergic interneurons, the opposite of the main pathological mechanism of DS. Analyzing single-unit recordings, we observed that temporal disorganization of the firing of putative interneurons in the period immediately before the seizure (preictal) precedes the increase of their activity at seizure onset, together with the entire neuronal network. Moreover, we found early signatures of the preictal period in the spectral features of hippocampal and cortical field potential of Scn1a mice and of patients\' EEG, which are consistent with the dysfunctions that we observed in single neurons and that allowed seizure prediction. Therefore, the perturbed preictal activity of interneurons leads to their hyperactivity at the onset of generalized seizures, which have low-voltage-fast features that are similar to those observed in other epilepsies and are triggered by hyperactivity of GABAergic neurons. Preictal spectral features may be used as predictive seizure biomarkers.

FMRFamide, a member of the neuropeptide family, is involved in numerous physiological processes. FMRFamide-activated sodium channels (FaNaCs) are a family of non-voltage-gated, amiloride-sensitive, Na+-selective channels triggered by the neuropeptide FMRFamide. In the present study, the full-length cDNA of the FaNaC receptor of Sepiella japonica (SjFaNaC) was cloned. The cDNA of SjFaNaC was 3004 bp long with an open reading frame (ORF) of 1812 bp, encoding 603 amino acid residues with no signal peptide at the N-terminus. Sequence analysis indicated that SjFaNaC shared a high identity with other cephalopods FaNaCs and formed a sister clade with bivalves. The protein structure was predicted using SWISS-MODEL with AcFaNaC as the template. Quantitative real-time PCR (qRT-PCR) revealed that SjFaNaC transcripts were highly expressed in both female and male reproductive organs, as well as in the optic lobe and brain of the central nervous system (CNS). Results of in situ hybridisation (ISH) showed that SjFaNaC mRNA was mainly distributed in the medulla and deep retina of the optic lobe and in both the supraesophageal and subesophageal masses of the brain. Subcellular localisation indicated that the SjFaNaC protein was localised intracellularly and on the cell surface of HEK293T cells. In summary, these findings may lay the foundation for future exploration of the functions of SjFaNaC in cephalopods.

Pathogenic variants in SCN8A, which encodes the voltage-gated sodium (NaV) channel NaV1.6, associate with neurodevelopmental disorders, including developmental and epileptic encephalopathy. Previous approaches to determine SCN8A variant function may be confounded by use of a neonatally expressed, alternatively spliced isoform of NaV1.6 (NaV1.6N) and engineered mutations rendering the channel tetrodotoxin (TTX) resistant. We investigated the impact of SCN8A alternative splicing on variant function by comparing the functional attributes of 15 variants expressed in 2 developmentally regulated splice isoforms (NaV1.6N, NaV1.6A). We employed automated patch clamp recording to enhance throughput, and developed a neuronal cell line (ND7/LoNav) with low levels of endogenous NaV current to obviate the need for TTX-resistance mutations. Expression of NaV1.6N or NaV1.6A in ND7/LoNav cells generated NaV currents with small, but significant, differences in voltage dependence of activation and inactivation. TTX-resistant versions of both isoforms exhibited significant functional differences compared with the corresponding WT channels. We demonstrated that many of the 15 disease-associated variants studied exhibited isoform-dependent functional effects, and that many of the studied SCN8A variants exhibited functional properties that were not easily classified as either gain- or loss-of-function. Our work illustrates the value of considering molecular and cellular context when investigating SCN8A variants.

Small fiber neuropathy (SFN) is a common and debilitating disease in which the terminals of small diameter sensory axons degenerate, producing sensory loss, and in many patients neuropathic pain. While a substantial number of cases are attributable to diabetes, almost 50% are idiopathic. An underappreciated aspect of the disease is its late onset in most patients. Animal models of human genetic mutations that produce SFN also display age-dependent phenotypes suggesting that aging is an important contributor to the risk of development of the disease. In this review we define how particular sensory neurons are affected in SFN and discuss how aging may drive the disease. We also evaluate how animal models of SFN can define disease mechanisms that will provide insight into early risk detection and suggest novel therapeutic interventions.