scholarly journals In silico Identification of Disrupted Myocardial Calcium Homeostasis as Proarrhythmic Trigger in Arrhythmogenic Cardiomyopathy

2021 ◽  
Vol 12 ◽  
Author(s):  
Aurore Lyon ◽  
Chantal J. M. van Opbergen ◽  
Mario Delmar ◽  
Jordi Heijman ◽  
Toon A. B. van Veen
Keyword(s):  
Calcium Entry ◽  
Calcium Handling ◽  
Human Model ◽  
Loss Of Function ◽  
Adrenergic Stimulation ◽  
Arrhythmogenic Cardiomyopathy ◽  
Beta Adrenergic ◽  
Plakophilin 2 ◽  
Calcium Affinity ◽  
Delayed Afterdepolarizations

Background: Patients with arrhythmogenic cardiomyopathy may suffer from lethal ventricular arrhythmias. Arrhythmogenic cardiomyopathy is predominantly triggered by mutations in plakophilin-2, a key component of cell-to-cell adhesion and calcium cycling regulation in cardiomyocytes. Calcium dysregulation due to plakophilin-2 mutations may lead to arrhythmias but the underlying pro-arrhythmic mechanisms remain unclear.Aim: To unravel the mechanisms by which calcium-handling abnormalities in plakophilin-2 loss-of-function may contribute to proarrhythmic events in arrhythmogenic cardiomyopathy.Methods: We adapted a computer model of mouse ventricular electrophysiology using recent experimental calcium-handling data from plakophilin-2 conditional knock-out (PKP2-cKO) mice. We simulated individual effects of beta-adrenergic stimulation, modifications in connexin43-mediated calcium entry, sodium-calcium exchanger (NCX) activity and ryanodine-receptor 2 (RyR2) calcium affinity on cellular electrophysiology and occurrence of arrhythmogenic events (delayed-afterdepolarizations). A population-of-models approach was used to investigate the generalizability of our findings. Finally, we assessed the potential translation of proposed mechanisms to humans, using a human ventricular cardiomyocyte computational model.Results: The model robustly reproduced the experimental calcium-handling changes in PKP2-cKO cardiomyocytes: an increased calcium transient amplitude (562 vs. 383 nM), increased diastolic calcium (120 vs. 91 nM), reduced L-type calcium current (15.0 vs. 21.4 pA/pF) and an increased free SR calcium (0.69 vs. 0.50 mM). Under beta-adrenergic stimulation, PKP2-cKO models from the population of models (n = 61) showed a higher susceptibility to delayed-afterdepolarizations compared to control (41 vs. 3.3%). Increased connexin43-mediated calcium entry further elevated the number of delayed-afterdepolarizations (78.7%, 2.5-fold increase in background calcium influx). Elevated diastolic cleft calcium appeared responsible for the increased RyR2-mediated calcium leak, promoting delayed-afterdepolarizations occurrence. A reduction in RyR2 calcium affinity prevented delayed-afterdepolarizations in PKP2-cKO models (24.6 vs. 41%). An additional increase in INCX strongly reduced delayed-afterdepolarizations occurrence, by lowering diastolic cleft calcium levels. The human model showed similar outcomes, suggesting a potential translational value of these findings.Conclusion: Beta-adrenergic stimulation and connexin43-mediated calcium entry upon loss of plakophilin-2 function contribute to generation of delayed-afterdepolarizations. RyR2 and NCX dysregulation play a key role in modulating these proarrhythmic events. This work provides insights into potential future antiarrhythmic strategies in arrhythmogenic cardiomyopathy due to plakophilin-2 loss-of-function.

Abstract 279: Loss Of Rad GTPase Produces A Sympathomimetic Cardiac Phenotype Leading To Calcium Overload And Arrhythmia.

2013 ◽  
Vol 113 (suppl_1) ◽  
Author(s):  
Janet R Manning ◽  
Guo Yin ◽  
John Penn ◽  
Catherine Kaminski ◽  
Gail Sievert ◽  
...  
Keyword(s):  
Calcium Current ◽  
Ventricular Myocytes ◽  
Isolated Heart ◽  
Small Gtpase ◽  
Calcium Transients ◽  
Calcium Handling ◽  
Adrenergic Stimulation ◽  
Biological Organization ◽  
Beta Adrenergic ◽  
Cardiac Phenotype

Background: Rad is a small GTPase that regulates inward calcium current in excitable cells. In human heart failure, calcium dysregulation is accompanied by lowered levels of Rad expression, and dampened responsiveness to beta-adrenergic stimulation. However, the effects of Rad ablation on calcium cycling in the heart have yet to be examined. Hypothesis: Loss of Rad emulates beta-adrenergic stimulation in cardiac myocytes, elevating calcium levels, and promoting after depolarizations. Methods: The role of Rad was evaluated using knockout mice (RadKO)at several levels of biological organization. Inward calcium current, action potentials, fractional shortening, and calcium transients were measured in isolated ventricular myocytes. Functional parameters were recorded in the isolated working heart, and telemetry was used to monitor ECGs in intact freely-roaming mice. Results: Isolated ventricular myoctyes from Rad null mice display increased ICaL density, greater inward current at low voltages, and after depolarizations at low frequencies. Isolated cells exhibit increased diastolic calcium levels and increased dynamic calcium changes in transient amplitude during pacing. Further, RadKO cardiomyocytes develop a significantly greater number of spontaneous calcium transients than wildtype cohorts. Ventricular myocytes and intact working hearts from RadKO mice fail to respond to beta-adrenergic stimulation at the level of channel activation, calcium transient kinetics, and exhibit a significantly dampened change in +dP/dt in the isolated heart. Consistent with these effects, PKA substrates are phosphorylated at baseline in RadKO cardiomyocytes, suggesting tonic PKA activation. Conclusion: These data suggest the new provocative hypothesis that beta-adrenergic-mediated changes in excitation, calcium handling, and heart contraction involve relief of Rad-dependent negative regulation of channel function.

scholarly journals NOS1AP polymorphisms reduce NOS1 activity and interact with prolonged repolarization in arrhythmogenesis

2020 ◽  
Author(s):  
Carlotta Ronchi ◽  
Joyce Bernardi ◽  
Manuela Mura ◽  
Manuela Stefanello ◽  
Beatrice Badone ◽  
...  
Keyword(s):  
Action Potential ◽  
Nucleotide Polymorphisms ◽  
Loss Of Function ◽  
Adrenergic Stimulation ◽  
Single Nucleotide ◽  
Current Events ◽  
Delayed Afterdepolarizations ◽  
Cardiac Sudden Death ◽  
Prolonged Action Potential Duration

Abstract Aims  NOS1AP single-nucleotide polymorphisms (SNPs) correlate with QT prolongation and cardiac sudden death in patients affected by long QT syndrome type 1 (LQT1). NOS1AP targets NOS1 to intracellular effectors. We hypothesize that NOS1AP SNPs cause NOS1 dysfunction and this may converge with prolonged action-potential duration (APD) to facilitate arrhythmias. Here we test (i) the effects of NOS1 inhibition and their interaction with prolonged APD in a guinea pig cardiomyocyte (GP-CMs) LQT1 model; (ii) whether pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) from LQT1 patients differing for NOS1AP variants and mutation penetrance display a phenotype compatible with NOS1 deficiency. Methods and results  In GP-CMs, NOS1 was inhibited by S-Methyl-L-thiocitrulline acetate (SMTC) or Vinyl-L-NIO hydrochloride (L-VNIO); LQT1 was mimicked by IKs blockade (JNJ303) and β-adrenergic stimulation (isoproterenol). hiPSC-CMs were obtained from symptomatic (S) and asymptomatic (AS) KCNQ1-A341V carriers, harbouring the minor and major alleles of NOS1AP SNPs (rs16847548 and rs4657139), respectively. In GP-CMs, NOS1 inhibition prolonged APD, enhanced ICaL and INaL, slowed Ca2+ decay, and induced delayed afterdepolarizations. Under action-potential clamp, switching to shorter APD suppressed ‘transient inward current’ events induced by NOS1 inhibition and reduced cytosolic Ca2+. In S (vs. AS) hiPSC-CMs, APD was longer and ICaL larger; NOS1AP and NOS1 expression and co-localization were decreased. Conclusion  The minor NOS1AP alleles are associated with NOS1 loss of function. The latter likely contributes to APD prolongation in LQT1 and converges with it to perturb Ca2+ handling. This establishes a mechanistic link between NOS1AP SNPs and aggravation of the arrhythmia phenotype in prolonged repolarization syndromes.

Electrophysiological effects of alpha 2-adrenergic stimulation in canine cardiac Purkinje fibers

1995 ◽  
Vol 268 (5) ◽  
pp. H2024-H2035 ◽  
Author(s):  
R. A. Samson ◽  
J. J. Cai ◽  
E. F. Shibata ◽  
J. B. Martins ◽  
H. C. Lee
Keyword(s):  
Action Potential ◽  
Pertussis Toxin ◽  
Adrenergic Stimulation ◽  
Beta Adrenergic ◽  
Purkinje Fibers ◽  
Cardiac Purkinje Fibers ◽  
Action Potential Prolongation ◽  
Delayed Afterdepolarizations ◽  
Baseline Values ◽  
Electrophysiological Effects

The effects of alpha 2-adrenergic stimulation on action potentials were measured in isolated canine Purkinje fibers. Action potential durations at 50 and 90% of repolarization (APD50 and APD90) were significantly prolonged by 0.25 microM l-norepinephrine + 0.5 microM dl-propranolol (NE+P) from baseline values of 166 +/- 7 and 249 +/- 9 (SE) ms (n = 7) to 174 +/- 7 and 265 +/- 9 ms, respectively (P < 0.05 for both). Selective alpha 2-blockade with 0.01 microM yohimbine (YO) reduced this prolongation by NE+P in APD50 and APD90 to 169 +/- 7 and 256 +/- 8 ms, respectively (P < 0.05 compared with NE+P). Additional selective alpha 1-blockade with 0.01 microM prazosin (PZ) completely blocked the effects of NE+P, returning APD50 and APD90 to 163 +/- 7 and 250 +/- 9 ms (not different from baseline). After incubation of isolated Purkinje fibers with pertussis toxin (1 microgram/ml), which reduced the availability of a 41-kDa membrane protein for ADP ribosylation by 70 +/- 7% (n = 4, P < 0.05), YO failed to reverse the prolongation in action potential durations brought on by NE+P, but the effects of PZ were intact. The effects of alpha 2-stimulation on beta-adrenergic-induced delayed afterdepolarizations (DADs) were studied by burst pacing of Purkinje fibers in Tyrode solution containing 7.5 mM Ca2+. The DADs induced in the presence of NE+PZ (beta- + alpha 2-stimulation) were significantly smaller in amplitude and required a shorter pacing cycle length to reach threshold than those induced in the presence of NE+PZ+YO (unopposed beta-adrenergic stimulation). Furthermore sustained triggered activity, seen in five of eight preparations under beta-stimulation, could no longer be elicited in the presence of beta- + alpha 2-stimulation. These results suggest that the postjunctional alpha 2-adrenergic receptors in canine Purkinje fibers are coupled to a pertussis toxin-sensitive G protein and that stimulation of these receptors leads to action potential prolongation and suppression of DADs induced by beta-adrenergic stimulation.

scholarly journals Calcium as a Key Player in Arrhythmogenic Cardiomyopathy: Adhesion Disorder or Intracellular Alteration?

2019 ◽  
Vol 20 (16) ◽  
pp. 3986 ◽  
Author(s):  
Francesco Moccia ◽  
Francesco Lodola ◽  
Ilaria Stadiotti ◽  
Chiara Assunta Pilato ◽  
Milena Bellin ◽  
...  
Keyword(s):  
Cardiac Contractility ◽  
Induced Pluripotent Stem Cell ◽  
Arrhythmogenic Cardiomyopathy ◽  
Mechanical Coupling ◽  
Plakophilin 2 ◽  
Genes Encoding ◽  
Life Threatening ◽  
Delayed Afterdepolarizations ◽  
Induced Pluripotent

Arrhythmogenic cardiomyopathy (ACM) is an inherited heart disease characterized by sudden death in young people and featured by fibro-adipose myocardium replacement, malignant arrhythmias, and heart failure. To date, no etiological therapies are available. Mutations in desmosomal genes cause abnormal mechanical coupling, trigger pro-apoptotic signaling pathways, and induce fibro-adipose replacement. Here, we discuss the hypothesis that the ACM causative mechanism involves a defect in the expression and/or activity of the cardiac Ca2+ handling machinery, focusing on the available data supporting this hypothesis. The Ca2+ toolkit is heavily remodeled in cardiomyocytes derived from a mouse model of ACM defective of the desmosomal protein plakophilin-2. Furthermore, ACM-related mutations were found in genes encoding for proteins involved in excitation‒contraction coupling, e.g., type 2 ryanodine receptor and phospholamban. As a consequence, the sarcoplasmic reticulum becomes more eager to release Ca2+, thereby inducing delayed afterdepolarizations and impairing cardiac contractility. These data are supported by preliminary observations from patient induced pluripotent stem-cell-derived cardiomyocytes. Assessing the involvement of Ca2+ signaling in the pathogenesis of ACM could be beneficial in the treatment of this life-threatening disease.

scholarly journals Hemi- and Homozygous Loss-of-Function Mutations in DSG2 (Desmoglein-2) Cause Recessive Arrhythmogenic Cardiomyopathy with an Early Onset

2021 ◽  
Vol 22 (7) ◽  
pp. 3786
Author(s):  
Andreas Brodehl ◽  
Alexey Meshkov ◽  
Roman Myasnikov ◽  
Anna Kiseleva ◽  
Olga Kulikova ◽  
...  
Keyword(s):  
Medical History ◽  
Splice Site Mutation ◽  
Pathogenic Mutation ◽  
Large Deletion ◽  
Loss Of Function ◽  
Arrhythmogenic Cardiomyopathy ◽  
Site Mutation ◽  
Snp Arrays ◽  
Number Of Patients ◽  
History Of

About 50% of patients with arrhythmogenic cardiomyopathy (ACM) carry a pathogenic or likely pathogenic mutation in the desmosomal genes. However, there is a significant number of patients without positive familial anamnesis. Therefore, the molecular reasons for ACM in these patients are frequently unknown and a genetic contribution might be underestimated. Here, we used a next-generation sequencing (NGS) approach and in addition single nucleotide polymor-phism (SNP) arrays for the genetic analysis of two independent index patients without familial medical history. Of note, this genetic strategy revealed a homozygous splice site mutation (DSG2–c.378+1G>T) in the first patient and a nonsense mutation (DSG2–p.L772X) in combination with a large deletion in DSG2 in the second one. In conclusion, a recessive inheritance pattern is likely for both cases, which might contribute to the hidden medical history in both families. This is the first report about these novel loss-of-function mutations in DSG2 that have not been previously identi-fied. Therefore, we suggest performing deep genetic analyses using NGS in combination with SNP arrays also for ACM index patients without obvious familial medical history. In the future, this finding might has relevance for the genetic counseling of similar cases.

scholarly journals Long-QT founder variant T309I-Kv7.1 with dominant negative pattern may predispose delayed afterdepolarizations under β-adrenergic stimulation

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Iva Synková ◽  
Markéta Bébarová ◽  
Irena Andršová ◽  
Larisa Chmelikova ◽  
Olga Švecová ◽  
...  
Keyword(s):  
Calcium Current ◽  
Cell Model ◽  
Dominant Negative ◽  
Qtc Interval ◽  
Adrenergic Stimulation ◽  
Patch Clamp Technique ◽  
Long Qt ◽  
Channel Trafficking ◽  
Whole Cell Patch Clamp ◽  
Delayed Afterdepolarizations

AbstractThe variant c.926C > T (p.T309I) in KCNQ1 gene was identified in 10 putatively unrelated Czech families with long QT syndrome (LQTS). Mutation carriers (24 heterozygous individuals) were more symptomatic compared to their non-affected relatives (17 individuals). The carriers showed a mild LQTS phenotype including a longer QTc interval at rest (466 ± 24 ms vs. 418 ± 20 ms) and after exercise (508 ± 32 ms vs. 417 ± 24 ms), 4 syncopes and 2 aborted cardiac arrests. The same haplotype associated with the c.926C > T variant was identified in all probands. Using the whole cell patch clamp technique and confocal microscopy, a complete loss of channel function was revealed in the homozygous setting, caused by an impaired channel trafficking. Dominant negativity with preserved reactivity to β-adrenergic stimulation was apparent in the heterozygous setting. In simulations on a human ventricular cell model, the dysfunction resulted in delayed afterdepolarizations (DADs) and premature action potentials under β-adrenergic stimulation that could be prevented by a slight inhibition of calcium current. We conclude that the KCNQ1 variant c.926C > T is the first identified LQTS-related founder mutation in Central Europe. The dominant negative channel dysfunction may lead to DADs under β-adrenergic stimulation. Inhibition of calcium current could be possible therapeutic strategy in LQTS1 patients refractory to β-blocker therapy.

scholarly journals CRISPR/Cas9-edited PKP2 knock-out (JMUi001-A-2) and DSG2 knock-out (JMUi001-A-3) iPSC lines as an isogenic human model system for arrhythmogenic cardiomyopathy (ACM)

2021 ◽  
Vol 53 ◽  
pp. 102256
Author(s):  
Anna Janz ◽  
Miriam Zink ◽  
Alexandra Cirnu ◽  
Annika Hartleb ◽  
Christina Albrecht ◽  
...  
Keyword(s):  
Model System ◽  
Human Model ◽  
Arrhythmogenic Cardiomyopathy ◽  
Knock Out

Compounds that increase cAMP prevent ischemia-reperfusion pulmonary capillary injury

1992 ◽  
Vol 72 (2) ◽  
pp. 492-497 ◽  
Author(s):  
W. K. Adkins ◽  
J. W. Barnard ◽  
S. May ◽  
A. F. Seibert ◽  
J. Haynes ◽  
...  
Keyword(s):  
Adenylate Cyclase ◽  
Vascular Permeability ◽  
Vascular Resistance ◽  
Capillary Permeability ◽  
Ischemia Reperfusion ◽  
Receptor Activation ◽  
Adrenergic Stimulation ◽  
Beta Adrenergic ◽  
Beta Receptor ◽  
Pulmonary Capillary

This study evaluated the physiological effects of compounds that increase adenosine 3′,5′-cyclic monophosphate (cAMP) on changes in pulmonary capillary permeability and vascular resistance induced by ischemia-reperfusion (I-R) in isolated blood-perfused rabbit lungs. cAMP was elevated by 1) beta-adrenergic stimulation with isoproterenol (ISO, 10(-5) M), 2) post-beta-receptor stimulation of adenylate cyclase with forskolin (FSK, 10(-5) M), 3) and dibutyryl cAMP (DBcAMP, 1 mM), a cAMP analogue. Vascular permeability was assessed by determining the capillary filtration coefficient (Kf,c), and capillary pressure was measured using the double occlusion technique. The total, arterial, and venous vascular resistances were calculated from measured pulmonary arterial, venous, and capillary pressures and blood flow. Reperfusion after 2 h of ischemia significantly (P less than 0.05) increased Kf,c (from 0.115 +/- 0.028 to 0.224 +/- 0.040 ml.min-1.cmH2O-1.100 g-1). These I-R-induced changes in capillary permeability were prevented when ISO, FSK, or DBcAMP was added to the perfusate at reperfusion (0.110 +/- 0.022 and 0.103 +/- 0.021, 0.123 +/- 0.029 and 0.164 +/- 0.024, and 0.153 +/- 0.030 and 0.170 +/- 0.027 ml.min-1.cmH2O-1.100 g-1, respectively). I-R significantly increased total, arterial, and venous vascular resistances. These increases in vascular resistance were also blocked by ISO, FSK, and DBcAMP. These data suggest that beta-adrenergic stimulation, post-beta-receptor activation of adenylate cyclase, and DBcAMP prevent the changes in pulmonary vascular permeability and vascular resistances caused by I-R in isolated rabbit lungs through a mechanism involving an increase in intracellular levels of cAMP.(ABSTRACT TRUNCATED AT 250 WORDS)

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