Anniek Frederike Lubberding

Glucagon-like peptide-1 receptor agonists (GLP-1 RAs) are increasingly used to treat type 2 diabetes and obesity. Albeit cardiovascular outcomes generally improve, treatment with GLP-1 RAs is associated with increased heart rate, the mechanism of which is unclear. We employed a large animal model, the female landrace pig, and used multiple in vivo and ex vivo approaches including pharmacological challenges, electrophysiology, and high-resolution mass spectrometry to explore how GLP-1 elicits an increase in heart rate. In anaesthetized pigs, neither cervical vagotomy, adrenergic blockers (alpha, beta, or combined alpha-beta blockade), ganglionic blockade (hexamethonium), nor inhibition of hyperpolarization-activated cyclic nucleotide–gated (HCN) channels (ivabradine) abolished the marked chronotropic effect of GLP-1. GLP-1 administration to isolated perfused pig hearts also increased heart rate, which was abolished by GLP-1 receptor blockade. Electrophysiological characterization of GLP-1 effects in vivo and in isolated perfused hearts localized electrical modulation to the atria and conduction system. In isolated sinus nodes, GLP-1 administration shortened the action potential cycle length of pacemaker cells and shifted the site of earliest activation. The effect was independent of HCN blockade. Collectively, these data support a direct effect of GLP-1 on GLP-1 receptors within the heart. Consistently, single nucleus RNA sequencing showed GLP-1 receptor expression in porcine pacemaker cells. Quantitative phosphoproteomics analyses of sinus node samples revealed that GLP-1 administration leads to phosphorylation changes of calcium cycling proteins of the sarcoplasmic reticulum, known to regulate heart rate. GLP-1 has direct chronotropic effects on the heart mediated by GLP-1 receptors in pacemaker cells of the sinus node, inducing changes in action potential morphology and the leading pacemaker site through a calcium signalling response characterized by PKA-dependent phosphorylation of Ca²⁺ cycling proteins involved in pacemaking. Targeting the pacemaker calcium clock may be a strategy to lower heart rate in people treated with GLP-1 RAs.

Aim: The voltage‐gated potassium channel K _v 11.1 is important for repolarizing the membrane potential in excitable cells such as myocytes, pancreatic α‐ and β‐cells. Moxifloxacin blocks the K _v 11.1 channel and increases the risk of hypoglycaemia in patients with diabetes. We investigated glucose regulation and secretion of glucoregulatory hormones in young people with and without moxifloxacin, a drug known to block the K _v 11.1 channel. Materials and Methods: The effect of moxifloxacin (800 mg/day for 4 days) or placebo on glucose regulation was assessed in a randomized, double‐blind, crossover study of young men and women (age 20‐40 years and body mass index 18.5‐27.5 kg/m ² ) without chronic disease, using 6‐h oral glucose tolerance tests and continuous glucose monitoring. Results: Thirty‐eight participants completed the study. Moxifloxacin prolonged the QTcF interval and increased heart rate. Hypoglycaemia was more frequently observed with moxifloxacin, both during the 8 days of continuous glucose monitoring and during the oral glucose tolerance tests. Hypoglycaemia questionnaire scores were higher after intake of moxifloxacin. Moxifloxacin reduced the early plasma‐glucose response (AUC _0‐30 min ) by 7% (95% CI: −9% to −4%, p < .01), and overall insulin response (AUC _{0‐360 min} ) decreased by 18% (95% CI: −24% to −11%, p < .01) and plasma glucagon increased by 17% (95% CI: 4%‐33%, p = .03). Insulin sensitivity calculated as the Matsuda index increased by 11%, and MISI, an index of muscle insulin sensitivity, increased by 34%. Conclusions: In young men and women, moxifloxacin, a drug known to block the K _v 11.1 channel, increased QT interval, decreased glucose levels and was associated with increased muscle insulin sensitivity and more frequent episodes of hypoglycaemia.

Voltage‐gated potassium (K_v) channels play an important role in the repolarization of a variety of excitable tissues, including in the cardiomyocyte and the pancreatic beta cell. Recently, individuals carrying loss‐of‐function (LoF) mutations in KCNQ1, encoding K_v7.1, and KCNH2 (hERG), encoding K_v11.1, were found to exhibit post‐prandial hyperinsulinaemia and episodes of hypoglycaemia. These LoF mutations also cause the cardiac disorder long QT syndrome (LQTS), which can be aggravated by hypoglycaemia. Interestingly, patients with LQTS also have a higher burden of diabetes compared to the background population, an apparent paradox in relation to the hyperinsulinaemic phenotype, and KCNQ1 has been identified as a type 2 diabetes risk gene. This review article summarizes the involvement of delayed rectifier K⁺ channels in pancreatic beta cell function, with emphasis on K_v7.1 and K_v11.1, using the cardiomyocyte for context. The functional and clinical consequences of LoF mutations and polymorphisms in these channels on blood glucose homeostasis are explored using evidence from pre‐clinical, clinical and genome‐wide association studies, thereby evaluating the link between LQTS, hyperinsulinaemia and type 2 diabetes.

Background: Adenosine leads to atrial action potential (AP) shortening through activation of adenosine 1 receptors (A₁-R) and subsequent opening of G-protein-coupled inwardly rectifying K⁺ channels. Extracellular production of adenosine is drastically increased during stress and ischemia. Objective: The aim of this study was to address whether the pharmacological blockade of endogenous production of adenosine and of its signaling prevents atrial fibrillation (AF). Methods: The role of A₁-R activation on atrial action potential duration, refractoriness, and AF vulnerability was investigated in rat isolated beating heart preparations (Langendorff) with an A₁-R agonist [2-chloro-N⁶-cyclopentyladenosine (CCPA), 50 nM] and antagonist [1-butyl-3-(3-hydroxypropyl)-8-(3-noradamantyl)xanthine (PSB36), 40 nM]. Furthermore, to interfere with the endogenous adenosine release, the ecto-5′-nucleotidase (CD73) inhibitor was applied [5′-(α,β-methylene) diphosphate sodium salt (AMPCP), 500 μM]. Isolated trabeculae from human right atrial appendages (hRAAs) were used for comparison. Results: As expected, CCPA shortened AP duration at 90% of repolarization (APD₉₀) and effective refractory period (ERP) in rat atria. PSB36 prolonged APD₉₀ and ERP in rat atria, and CD73 inhibition with AMPCP prolonged ERP in rats, confirming that endogenously produced amount of adenosine is sufficiently high to alter atrial electrophysiology. In human atrial appendages, CCPA shortened APD₉₀, while PSB36 prolonged it. Rat hearts treated with CCPA are prone to AF. In contrast, PSB36 and AMPCP prevented AF events and reduced AF duration (vehicle, 11.5 ± 2.6 s; CCPA, 40.6 ± 16.1 s; PSB36, 6.5 ± 3.7 s; AMPCP, 3.0 ± 1.4 s; P < 0.0001). Conclusion: A₁-R activation by intrinsic adenosine release alters atrial electrophysiology and promotes AF. Inhibition of adenosine pathway protects atria from arrhythmic events.

Ventricular fibrillation (VF) during acute myocardial infarction (AMI) is an important contributor to sudden cardiac death. Large animal models are widely used to study AMI-induced arrhythmia, but the mode of AMI induction ranges from thoracotomy and surgical ligation of a coronary vessel (open chest) to minimally invasive techniques, including balloon occlusion (closed chest). How the choice of induction affects arrhythmia development is unclear. The aim of this study was to compare an open-chest and a closed-chest model with regard to hemodynamics, electrophysiology, and arrhythmia development. Forty-two female Danish Landrace pigs (20 open chest, 22 closed chest) were anesthetized, and occlusion of the mid-left anterior descending coronary artery was performed for 60 min. Opening the chest reduced blood pressure and cardiac output (Δ −22 mmHg, Δ −1.5 L/min from baseline, both P < 0.001 intragroup). Heart rate decreased with opening of the chest but increased with balloon placement ( P < 0.001). AMI-induced ST elevation was lower in the open-chest group ( P < 0.001). Premature ventricular contractions occurred in two distinct phases (0–15 and 15–40 min), the latter of which was delayed in the open-chest group ( P = 0.005). VF occurred in 7 out of 20 and 12 out of 22 pigs in the open-chest and closed-chest groups, respectively ( P = 0.337), with longer time-to-VF in the open-chest group (23.4 ± 1.2 min in open chest and 17.8 ± 1.4 min in closed chest; P = 0.007). In summary, opening the chest altered hemodynamic parameters and delayed the onset of ventricular arrhythmias. Hence, in the search for mechanisms and novel treatments of AMI-induced arrhythmia, caution should be taken when choosing between or comparing the results from these two models. NEW & NOTEWORTHY We demonstrated pronounced differences in hemodynamic parameters and time course of ventricular arrhythmias in regard to mode of infarct induction. Inducing myocardial infarction by thoracotomy and subsequent ligation decreased blood pressure and cardiac output and delayed the onset of ventricular arrhythmia, whereas balloon occlusion resulted in higher heart rates during infarct.

Aim: Cardiovascular complications, including cardiac arrhythmias, result in high morbidity and mortality in patients with type‐2 diabetes mellitus (T2DM). Clinical and experimental data suggest electrophysiological impairment of the natural pacemaker of the diabetic heart. The present study examined sinoatrial node (SAN) arrhythmias in a mouse model of T2DM and physiologically probed their underlying cause.Methods: Electrocardiograms were obtained from conscious diabetic db/db and lean control db/+ mice. In vivo SAN function was probed through pharmacological autonomic modulation with isoprenaline, atropine and carbachol. Blood pressure stability and heart rate variability (HRV) were evaluated. Intrinsic SAN function was evaluated through ex vivo imaging of spontaneous Ca²⁺ transients in isolated SAN preparations.Results: While lean control mice showed constant RR intervals during isoprenaline challenge, the diabetic mice experienced SAN arrhythmias with large RR fluctuations in a dose‐dependent manner. These arrhythmias were completely abolished by atropine pre‐treatment, while carbachol pretreatment significantly increased SAN arrhythmia frequency in the diabetic mice. Blood pressure and HRV were comparable in db/db and db/+ mice, suggesting that neither augmented baroreceptor feedback nor autonomic neuropathy is a likely arrhythmia mechanism. Cycle length response to isoprenaline was comparable in isolated SAN preparations from db/db and db/+ mice; however, Ca²⁺ spark frequency was significantly increased in db/db mice compared to db/+ at baseline and after isoprenaline.Conclusion: Our results demonstrate a dysfunction of cardiac pacemaking in an animal model of T2DM upon challenge with a β‐adrenergic agonist. Ex vivo, higher Ca²⁺ spark frequency is present in diabetic mice, which may be directly linked to in vivo arrhythmias.

Ventricular arrhythmia and subsequent sudden cardiac death (SCD) due to acute myocardial infarction (AMI) is one of the most frequent causes of death in humans. Lethal ventricular arrhythmias like ventricular fibrillation (VF) prior to hospitalization have been reported to occur in more than 10% of all AMI cases and survival in these patients is poor. Identification of risk factors and mechanisms for VF following AMI as well as implementing new risk stratification models and therapeutic approaches is therefore an important step to reduce mortality in people with high cardiovascular risk. Studying spontaneous VF following AMI in humans is challenging as it often occurs unexpectedly in a low risk subgroup. Large animal models of AMI can help to bridge this knowledge gap and are utilized to investigate occurrence of arrhythmias, involved mechanisms and therapeutic options. Comparable anatomy and physiology allow for this translational approach. Through experimental focus, using state-of-the-art technologies, including refined electrical mapping equipment and novel pharmacological investigations, valuable insights into arrhythmia mechanisms and possible interventions for arrhythmia-induced SCD during the early phase of AMI are now beginning to emerge. This review describes large experimental animal models of AMI with focus on first AMI-associated ventricular arrhythmias. In this context, epidemiology of first AMI, arrhythmogenic mechanisms and various potential therapeutic pharmacological targets will be discussed.

Aims : Acute myocardial infarction (AMI) is associated with intracellular Ca2+ build-up. In healthy ventricles, small conductance Ca2+-activated K+ (SK) channels are present but do not participate in repolarization. However, SK current is increased in chronic myocardial infarction and heart failure, and recently, SK channel inhibition was demonstrated to reduce arrhythmias in AMI rats. Hence, we hypothesized that SK channel inhibitors (NS8593 and AP14145) could reduce arrhythmia development during AMI in a porcine model.Methods and results : Twenty-seven pigs were randomized 1:1:1 to control, NS8593, or AP14145. Haemodynamic and electrophysiological parameters [electrocardiogram (ECG) and monophasic action potentials (MAP)] were continuously recorded. A balloon was placed in the mid-left anterior descending artery, blinded to treatment. Infusion lasted from 10 min before occlusion until 30 min after. Occlusion was maintained for 1 h, followed by 2 h of reperfusion. Upon occlusion, cardiac output dropped similarly in all groups, while blood pressure remained stable. Heart rate decreased in the NS8593 and AP14145 groups. QRS duration increased upon occlusion in all groups but more prominently in AP14145-treated pigs. Inhibition of SK channels did not affect QT interval. Infarct MAP duration shortened comparably in all groups. Ventricular fibrillation developed in 4/9 control-, 4/9 AP14145-, and 2/9 NS8593-treated pigs. Ventricular tachycardia was rarely observed in either group, whereas ventricular extrasystoles occurred comparably in all groups.Conclusion : Inhibition of SK channels was neither beneficial nor detrimental to ventricular arrhythmia development in the setting of AMI in this porcine model.

Adrenaline (epinephrine) is one of the prime messengers of the fight-or-flight response, favoring the activation of β-adrenergic receptors. Although general vasoconstriction to nonessential tissues is imperative, the vasodilatory effect of β-adrenergic receptor activation contends with this. We aimed to determine the dose-dependent effects of adrenaline on hemodynamics and to test whether adrenaline could lower blood pressure (BP) through a β₂-adrenergic pathway. Nineteen Danish landrace pigs were used to pharmacologically probe the hemodynamic effect of adrenaline. Pigs were anesthetized, intubated, and electrocardiogram, systolic BP (SBP), diastolic BP (DBP), and left ventricular pressure (LVP) were monitored continuously. First, we tested the dose-dependent effects of adrenaline (0.01–10 µg/kg). Second, we determined the response to adrenaline (0.3 µg/kg) after atropine, prazosin, and propranolol pretreatment. Finally, we tested the hemodynamic effect of salbutamol in a subset of pigs. All doses of adrenaline increased heart rate, while BP showed a biphasic response: At low doses, adrenaline decreased SBP from 118 ± 3 to 106 ± 4 mm Hg (n = 15; P < 0.05) and DBP from 86 ± 3 to 71 ± 3 (n = 15; P < 0.05), while at high doses, SBP and DBP increased. LVP showed a similar pattern, with a tendency of decreased pressure at low doses, and an increased pressure at high doses (P < 0.05). Pretreatment with autonomic blockers revealed that the increase in BP was due to α-adrenergic activity, while the decrease was due to β-adrenergic activity. In confirmation, β-adrenergic activation through salbutamol showed a similar decrease in SBP, DBP, and LVP. We conclude that adrenaline dose-dependently increases heart rate, while producing a biphasic response in BP with a decrease at low doses and an increase at high doses in an anesthetized, large-animal model.