A rare case of Brugada syndrome induced by hyperglycemia

Brugada syndrome is a rare genetic disorder of the cardiac sodium channels associated with an increased risk of sudden cardiac death. It is characterized by an electrocardiogram (EKG) showing a right bundle branch block with an elevation in the ST segment. This condition is associated with mutations in several pathologic genes including the most notable mutation in the SCN5A gene, which encodes for a voltage-gated cardiac sodium channel. The Brugada pattern on EKG can be spontaneous but can also be induced by a variety of etiologies including fever, electrolyte abnormalities, increased vagal tone and drugs such as sodium channel blockers, calcium channel blockers, tricyclic antidepressants and alcohol. One uncommon cause of Brugada syndrome is hyperglycemia. Of particular importance in diabetic patients, hyperglycemia can induce chronic cardiovascular complications as well as acute cardiac events via the induction of the Brugada pattern on EKG. We present a case of a 21-year-old non-insulin compliant diabetic man presenting to the Emergency Department with diabetic ketoacidosis (DKA) who exhibits the Brugada pattern EKG prior to developing ventricular tachycardia followed by cardiac arrest. The patient’s condition was induced by prolonged hyperglycemia in the setting of DKA with relatively mild electrolyte and pH abnormalities. Herein, this case is presented to highlight the Brugada pattern leading to cardiac arrest as a potential consequence of hyperglycemia and inform physicians on its incidence.

NMR solution structure and analysis of isolated S3b-S4a motif of repeat IV of the human cardiac sodium channel

Voltage-gated sodium channels are membrane proteins that play an important role in the propagation of electrical signals by mediating the rising phase of an action potential. Numerous diseases, including epilepsy, extreme pain, and certain cardiac arrhythmias have been linked to defects in these channels. The S3b-S4a helix-turn-helix motif (paddle motif) is a region of the channel that is involved in voltage sensing and undergoes significant structural changes during gating. It is also the binding site for many gating-modifier toxins. We determined the solution structure of the paddle motif from the fourth repeat of NaV1.5 in dodecylphosphocholine micelles by NMR spectroscopy and investigated its dynamics and micelle interactions. The structure displays a helix hairpin with a short connecting loop, and likely represents the activated conformation with three of the first four gating charges facing away from S3. Furthermore, paramagnetic relaxation measurements showed that the paddle motif is mainly interacting with the interface region of the micelle. NMR relaxation studies revealed that the paddle motif is mostly rigid, with some residues around the loop region and the last 4 residues on the C-terminus displaying heightened mobility. The structural findings reported here allowed the interpretation of three disease-causing mutations in this region of the human cardiac sodium channel, S1609W, F1617del and T1620M. The establishment of this model system for NMR studies of the paddle region offers a promising platform for future toxin interaction studies in the cardiac sodium channels, and similar approaches may be applied to other sodium channel isoforms.

Desmosomal vulnerability renders left atria more susceptible to detrimental effects of androgenic anabolic steroids

Funding Acknowledgements Type of funding sources: Public grant(s) – EU funding. Main funding source(s): CATCH ME Foundation Leducq BACKGROUND In cardiac myocytes, desmosomal proteins and ion channels form macromolecular complexes important for maintaining cell adhesion and electrical integrity. High serum levels of androgenic anabolic steroids (AAS) promote cardiac muscle growth, but any detrimental impact on atrial gene transcription and/or electrophysiological function is unknown. PURPOSE To investigate the effects of chronic AAS exposure on atria in a mouse model with desmosomal impairment. METHODS Young (8-10 week) male wild-type (WT) and heterozygous plakoglobin-deficient (plako+/-) mice were challenged with the AAS dihydrotestosterone (DHT) or placebo for 6 weeks by osmotic mini pumps. RNA sequencing (n = 3-6 atria/group) revealed effects of genotype and DHT on left atrial (LA) transcription. Membrane-localised cardiac sodium channels (Nav1.5) were visualised using direct STochastic Optical Reconstruction Microscopy (dSTORM, n = 5-11 LA/group, 122 cells in total) and clustering of individual molecules was quantified using persistence-based clustering. Patch clamping of LA cardiac myocytes was used to record whole cell sodium currents (n = 4-5 LA/group, 77 cells in total). LA action potentials and conduction velocity were evaluated using microelectrode and optical mapping techniques (n = 5-9 LA/group). RESULTS DHT increased expression of pro-hypertrophic transcripts, e.g. Igf1, Mtpn, fibrosis-associated transcripts, e.g. Col1a1, Col3a1, Lox and pro-inflammatory transcripts, e.g. Ccl6, C7, in both WT and plako+/- LA. Despite Scn5a transcript levels being maintained, dSTORM identified a 29% reduction (p = 0.042) in the number of Nav1.5 localisations at the membrane of plako+/- DHT LA cardiomyocytes, and 25% fewer localisations (p = 0.005) were found within Nav1.5 clusters, compared to WT DHT. Electrophysiological methods revealed a significant reduction in peak sodium current density, decreased action potential amplitude and conduction slowing in plako+/- LA after exposure to DHT. CONCLUSION This data suggests that a reduction in plakoglobin expression predisposes atrial cardiomyocytes to detrimental electrophysiological effects of high testosterone levels. This is characterised by a perturbed spatial organisation of Nav1.5, decreased sodium current density and conduction slowing. Abstract Figure. Abstract Picture

Protein Kinases Mediate Anti-Inflammatory Effects of Cannabidiol and Estradiol Against High Glucose in Cardiac Sodium Channels

Background: Cardiovascular anomalies are predisposing factors for diabetes-induced morbidity and mortality. Recently, we showed that high glucose induces changes in the biophysical properties of the cardiac voltage-gated sodium channel (Nav1.5) that could be strongly correlated to diabetes-induced arrhythmia. However, the mechanisms underlying hyperglycemia-induced inflammation, and how inflammation provokes cardiac arrhythmia, are not well understood. We hypothesized that inflammation could mediate the high glucose-induced biophyscial changes on Nav1.5 through protein phosphorylation by protein kinases A and C. We also hypothesized that this signaling pathway is, at least partly, involved in the cardiprotective effects of cannabidiol (CBD) and 17β-estradiol (E2).Methods and Results: To test these ideas, we used Chinese hamster ovarian (CHO) cells transiently co-transfected with cDNA encoding human Nav1.5 α-subunit under control, a cocktail of inflammatory mediators or 100 mM glucose conditions (for 24 h). We used electrophysiological experiments and action potential modeling. Inflammatory mediators, similar to 100 mM glucose, right shifted the voltage dependence of conductance and steady-state fast inactivation and increased persistent current leading to computational prolongation of action potential (hyperexcitability) which could result in long QT3 arrhythmia. We also used human iCell cardiomyocytes derived from inducible pluripotent stem cells (iPSC-CMs) as a physiologically relevant system, and they replicated the effects produced by inflammatory mediators observed in CHO cells. In addition, activators of PK-A or PK-C replicated the inflammation-induced gating changes of Nav1.5. Inhibitors of PK-A or PK-C, CBD or E2 mitigated all the potentially deleterious effects provoked by high glucose/inflammation.Conclusion: These findings suggest that PK-A and PK-C may mediate the anti-inflammatory effects of CBD and E2 against high glucose-induced arrhythmia. CBD, via Nav1.5, may be a cardioprotective therapeutic approach in diabetic postmenopausal population.

RYR2 Channel Inhibition Is the Principal Mechanism 0f Flecainide Action in CPVT

Rationale: The class Ic antiarrhythmic drug flecainide prevents ventricular tachyarrhythmia in patients with catecholaminergic polymorphic ventricular tachycardia (CPVT), a disease caused by hyperactive cardiac ryanodine receptor (RyR2) calcium (Ca) release. Although flecainide inhibits single RyR2 channels in vitro, reports have claimed that RyR2 inhibition by flecainide is not relevant for its mechanism of antiarrhythmic action and concluded that sodium channel block alone is responsible for flecainide's efficacy in CPVT. Objective: To determine whether RyR2 block independently contributes to flecainide's efficacy for suppressing spontaneous sarcoplasmic reticulum (SR) Ca release and for preventing ventricular tachycardia in vivo. Methods and Results: We synthesized N-methylated flecainide analogues (QX-FL and NM-FL) and showed that N-methylation reduces flecainide's inhibitory potency on RyR2 channels incorporated into artificial lipid bilayers. N-Methylation did not alter flecainide's inhibitory activity on human cardiac sodium channels expressed in HEK293T cells. Antiarrhythmic efficacy was tested utilizing a calsequestrin knockout (Casq2-/-) CPVT mouse model. In membrane-permeabilized Casq2-/- cardiomyocytes — lacking intact sarcolemma and devoid of sodium channel contribution — flecainide, but not its analogues, suppressed RyR2-mediated Ca release at clinically relevant concentrations. In voltage-clamped, intact Casq2-/- cardiomyocytes pretreated with tetrodotoxin (TTX) to inhibit sodium channels and isolate the effect of flecainide on RyR2, flecainide significantly reduced the frequency of spontaneous SR Ca release, while QX-FL and NM-FL did not. In vivo, flecainide effectively suppressed catecholamine-induced ventricular tachyarrhythmias in Casq2-/- mice, whereas NM-FL had no significant effect on arrhythmia burden, despite comparable sodium channel block. Conclusions: Flecainide remains an effective inhibitor of RyR2-mediated arrhythmogenic Ca release even when cardiac sodium channels are blocked. In mice with CPVT, sodium channel block alone did not prevent ventricular tachycardia. Hence, RyR2 channel inhibition likely constitutes the principal mechanism of antiarrhythmic action of flecainide in CPVT.

Protein kinases mediate anti-inflammatory effects of cannabidiol and estradiol against high glucose in cardiac sodium channels

Background and purpose. Cardiovascular anomalies are predisposing factors for diabetes-induced morbidity and mortality. Recently, we showed that high glucose induces changes in the biophysical properties of Nav1.5 that could be strongly correlated to diabetes-induced arrhythmia. However, the mechanisms underlying hyperglycemia-induced inflammation, and how inflammation provokes cardiac arrhythmia, are not well understood. We hypothesized that inflammation could mediate the high glucose-induced biophyscial changes on Nav1.5 through protein phosphorylation by protein kinases A and C. We also hypothesized that this signaling pathway is, at least partly, involved in the cardiprotective effects of CBD and E2. Experimental approach. To test these ideas, we used Chinese hamster ovarian (CHO) cells transiently co-transfected with cDNA encoding human Nav1.5 α-subunit under control, a cocktail of inflammatory mediators or 100 mM glucose conditions (for 24 hours). We used electrophysiological experiments and action potential modelling. Key Results. Inflammatory mediators, similar to 100 mM glucose, right shifted the voltage dependence of conductance and steady state fast inactivation and increased persistent current leading to computational prolongation of action potential (hyperexcitability) which could result in long QT3 arrhythmia. In addition, activators of PK-A or PK-C replicated the inflammation-induced gating changes of Nav1.5. Inhibitors of PK-A or PK-C, CBD or E2 mitigated all the potentially deleterious effects provoked by high glucose/inflammation. Conclusions and implications. These findings suggest that PK-A and PK-C may mediate the anti-inflammatory effects of CBD and E2 against high glucose-induced arrhythmia. CBD, via Nav1.5, may be a cardioprotective therapeutic approach in diabetic postmenopausal population.

Vascular endothelial growth factor promotes atrial arrhythmias by inducing acute intercalated disk remodeling

Atrial fibrillation (AF) is the most common arrhythmia and is associated with inflammation. AF patients have elevated levels of inflammatory cytokines known to promote vascular leak, such as vascular endothelial growth factor A (VEGF). However, the contribution of vascular leak and consequent cardiac edema to the genesis of atrial arrhythmias remains unknown. Previous work suggests that interstitial edema in the heart can acutely promote ventricular arrhythmias by disrupting ventricular myocyte intercalated disk (ID) nanodomains rich in cardiac sodium channels (NaV1.5) and slowing cardiac conduction. Interestingly, similar disruption of ID nanodomains has been identified in atrial samples from AF patients. Therefore, we tested the hypothesis that VEGF-induced vascular leak can acutely increase atrial arrhythmia susceptibility by disrupting ID nanodomains and slowing atrial conduction. Treatment of murine hearts with VEGF (30–60 min, at clinically relevant levels) prolonged the electrocardiographic P wave and increased susceptibility to burst pacing-induced atrial arrhythmias. Optical voltage mapping revealed slower atrial conduction following VEGF treatment (10 ± 0.4 cm/s vs. 21 ± 1 cm/s at baseline, p < 0.05). Transmission electron microscopy revealed increased intermembrane spacing at ID sites adjacent to gap junctions (GJs; 64 ± 9 nm versus 17 ± 1 nm in controls, p < 0.05), as well as sites next to mechanical junctions (MJs; 63 ± 4 nm versus 27 ± 2 nm in controls, p < 0.05) in VEGF–treated hearts relative to controls. Importantly, super-resolution microscopy and quantitative image analysis revealed reorganization of NaV1.5 away from dense clusters localized near GJs and MJs to a more diffuse distribution throughout the ID. Taken together, these data suggest that VEGF can acutely predispose otherwise normal hearts to atrial arrhythmias by dynamically disrupting NaV1.5-rich ID nanodomains and slowing atrial conduction. These data highlight inflammation-induced vascular leak as a potential factor in the development and progression of AF.

Abstract 15420: Resolving a Controversy: Inhibition of Cardiac Ryanodine Receptors is the Principal Mechanism of Antiarrhythmic Action of Flecainide in Catecholaminergic Polymorphic Ventricular Tachycardia

Rationale: The class Ic antiarrhythmic drug flecainide prevents ventricular tachyarrhythmia in patients with catecholaminergic polymorphic ventricular tachycardia (CPVT), a disease caused by hyperactive cardiac ryanodine receptor (RyR2) calcium (Ca) release. Although flecainide inhibits single RyR2 channels in vitro , reports have claimed that RyR2 inhibition by flecainide is not relevant for its mechanism of antiarrhythmic action and concluded that sodium channel block alone is responsible for flecainide’s efficacy in CPVT. Objective: To determine whether RyR2 block independently contributes to flecainide’s efficacy for suppressing spontaneous sarcoplasmic reticulum (SR) Ca release and for preventing ventricular tachycardia in vivo . Methods and Results: We synthesized N -methyl flecainide analogues (QX-FL and NM-FL) and showed that N -methylation reduces flecainide’s inhibitory potency on RyR2 channels but not on cardiac sodium channels. Antiarrhythmic efficacy was tested utilizing a calsequestrin knockout (Casq2-/-) CPVT mouse model. In membrane-permeabilized Casq2-/- cardiomyocytes — lacking intact sarcolemma and devoid of sodium channel contribution — flecainide, but not its analogues, suppressed RyR2-mediated Ca release at clinically relevant concentrations. In voltage-clamped, intact Casq2-/- cardiomyocytes pretreated with tetrodotoxin (TTX) to inhibit sodium channels and isolate the effect of flecainide on RyR2, flecainide significantly reduced the frequency of spontaneous SR Ca release, while QX-FL and NM-FL did not. In vivo , flecainide effectively suppressed catecholamine-induced ventricular tachyarrhythmias in Casq2-/- mice, whereas NM-FL did not, despite comparable sodium channel block. Conclusions: Flecainide remains an effective inhibitor of RyR2-mediated arrhythmogenic Ca release even when cardiac sodium channels are blocked. In mice with CPVT, sodium channel block alone was not enough to prevent arrhythmias. Hence, RyR2 inhibition by flecainide is critical for its mechanism of antiarrhythmic action.

Abstract 16722: Perinexal Width Modulates Sodium Channel Recruitment During Extracellular Stimulation

Excitability in cardiomyocytes is dependent on the subthreshold current required to raise transmembrane potential to the activation threshold of voltage gated sodium channels and sodium channel recruitment to trigger an action potential. Cardiac sodium channels are densely expressed in the intercalated disc within the perinexal nanodomain, which is 2 orders of magnitude narrower than bulk extracellular interstitium. We hypothesized that perinexal narrowing reduces extracellular induced excitability because the perinexus functions as a voltage divider. Methods: Excitability with an extracellular stimulus was quantified in isolated Langendorff perfused male retired breeder guinea pig hearts by strength duration curves using the Lapicque method. Interventions included changing extracellular potassium (K+: 3, 4.5, and 10 mM), inhibiting sodium channels (90-uM Flecainide), and narrowing the perinexus by increasing extracellular calcium (Ca2+: 1.25 to 2.5 mM). Results: Consistent with previous studies, decreasing K+ from 4.56 to 3 mM depressed excitability with 2.5 mM Ca2+ but not 1.25 mM Ca2+, and conduction velocity (CV) decreased by 10.5 % with both 1.25 and 2.5 mM Ca2+. When K+ was raised from 4.56 to 10 mM, no change was seen in excitability with both Ca2+ concentrations. However, CV decreased by 16% with both Ca2+ concentrations. Flecainide depressed excitability only with 2.5 but not 1.25 mM Ca2+. Meanwhile CV decreased by 13% with 1.25 but CV did not change with 2.5 mM Ca2+. Finally, raising Ca2+ alone at baseline decreased excitability, without substantially changing conduction. Conclusions: Elevating extracellular calcium to narrow perinexi reduces excitability measured by extracellular stimulation consistent with a hypothesis that sodium channels in the intercalated disc are electrically isolated from the bulk interstitium. Furthermore, excitability and conduction do not correlate in response to similar K+ changes when Ca2+ also varies, suggesting cardiac excitability and propagation are independent mechanisms when the excitatory current occurs through regenerative propagation as occurs through gap junctions or arrives via an extracellular field as occurs with pacing and ephaptic coupling.

Enantioselective Analysis of Mexiletine Using Chromatographic Techniques: A Review

Background: Mexiletine belongs to the β-amino-aryl-ether group of pharmaceutical and applied in the diagnosis of antiarrhythmics, allodynia, and myotonic disorders. In its chemical structure, it possesses a chiral center and practiced in the form of the racemic mixture. The production and accessibility of mexiletine have accompanied with a meaningful development in awareness of its pharmacologic actions. But in clinical arrhythmias and binding experiments on cardiac sodium channels, the (R)-enantiomer of mexiletine is more potent than the (S)-enantiomer. Also, (S)-enantiomer is further active in the diagnosis of allodynia than the (R)-enantiomer. Methods: During the last two decades, chromatographic techniques such as HPLC, and GC coupled with mass spectrometry or field ionization detector was used for the stereoselective analysis of MEX enantiomers. Results: The direct enantioresolution deal with the use of chiral stationary phases (CSPs) with or no pre-derivatization which depend on a chromophoric entity in the racemates whereas indirect HPLC process involved the use of chiral derivatization reagents (CDR) for the synthesis of diastereomeric derivatives of racemates. Different techniques have their strengths and weaknesses. Conclusion: Regulation of enantiomeric purity and estimation of particular enantiomers of drug molecules stays an essential topic for therapeutic, diagnostic, and regulatory uses and to promote a precise assessment of the hazards to human health by false enantiomers. This review aims to offer a systematic survey of the analytical methods (chromatography based) used in the enantioselective analysis of MEX developed in the last two decades (the year 2000 onwards).