Na+-dependent SR Ca2+ overload induces arrhythmogenic events in mouse cardiomyocytes with a human CPVT mutation.
OBJECTIVE
Mutations in the cardiac ryanodine receptor Ca(2+) release channel, RyR2, underlie catecholaminergic polymorphic ventricular tachycardia (CPVT), an inherited life-threatening arrhythmia. CPVT is triggered by spontaneous RyR2-mediated sarcoplasmic reticulum (SR) Ca(2+) release in response to SR Ca(2+) overload during beta-adrenergic stimulation. However, whether elevated SR Ca(2+) content--in the absence of protein kinase A activation--affects RyR2 function and arrhythmogenesis in CPVT remains elusive.


RESULTS
Isolated murine ventricular myocytes harbouring a human RyR2 mutation (RyR2(R4496C+/-)) associated with CPVT were investigated in the absence and presence of 1 micromol/L JTV-519 (RyR2 stabilizer) followed by 100 micromol/L ouabain intervention to increase cytosolic [Na(+)] and SR Ca(2+) load. Changes in membrane potential and intracellular [Ca(2+)] were monitored with whole-cell patch-clamping and confocal Ca(2+) imaging, respectively. At baseline, action potentials (APs), Ca(2+) transients, fractional SR Ca(2+) release, and SR Ca(2+) load were comparable in wild-type (WT) and RyR2(R4496C+/-) myocytes. Ouabain evoked significant increases in diastolic [Ca(2+)], peak systolic [Ca(2+)], fractional SR Ca(2+) release, and SR Ca(2+) content that were quantitatively similar in WT and RyR2(R4496C+/-) myocytes. Ouabain also induced arrhythmogenic events, i.e. spontaneous Ca(2+) waves, delayed afterdepolarizations and spontaneous APs, in both groups. However, the ouabain-induced increase in the frequency of arrhythmogenic events was dramatically larger in RyR2(R4496C+/-) when compared with WT myocytes. JTV-519 greatly reduced the frequency of ouabain-induced arrhythmogenic events.


CONCLUSIONS
The elevation of SR Ca(2+) load--in the absence of beta-adrenergic stimulation--is sufficient to increase the propensity for triggered arrhythmias in RyR2(R4496C+/-) cardiomyocytes. Stabilization of RyR2 by JTV-519 effectively reduces these triggered arrhythmias.
Sedej
Simon
S
Division of Cardiology, Medical University of Graz, Auenbruggerplatz 15, Graz A-8036, Austria.
Heinzel
Frank R
FR
Walther
Stefanie
S
Dybkova
Nataliya
N
Wakula
Paulina
P
Groborz
Jan
J
Gronau
Phillip
P
Maier
Lars S
LS
Vos
Marc A
MA
Lai
F Anthony
FA
Napolitano
Carlo
C
Priori
Silvia G
SG
Kockskämper
Jens
J
Pieske
Burkert
B
D000200:Action Potentials; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D020013:Calcium Signaling; D002395:Catecholamines / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005260:Female; D055879:Gene Knock-In Techniques; D006801:Humans; D008297:Male; D051379:Mice; D008822:Mice, Transgenic; D018613:Microscopy, Confocal; D009154:Mutation; D032383:Myocytes, Cardiac / Q000187:drug effects / Q000378:metabolism*; D010042:Ouabain / Q000494:pharmacology; D018408:Patch-Clamp Techniques; D010766:Phosphorylation; D019837:Ryanodine Receptor Calcium Release Channel / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*; D012519:Sarcoplasmic Reticulum / Q000187:drug effects / Q000378:metabolism*; D012964:Sodium / Q000378:metabolism*; D000254:Sodium-Potassium-Exchanging ATPase / Q000037:antagonists & inhibitors / Q000378:metabolism; D017180:Tachycardia, Ventricular / Q000235:genetics / Q000378:metabolism* / Q000517:prevention & control; D013841:Thiazepines / Q000494:pharmacology; D013997:Time Factors
D002395:Catecholamines; D004791:Enzyme Inhibitors; D019837:Ryanodine Receptor Calcium Release Channel; D013841:Thiazepines; C109183:K201 compound; D010042:Ouabain; D012964:Sodium; D000254:Sodium-Potassium-Exchanging ATPase; D002118:Calcium