Inhibition of cardiac potassium currents by oxidation-activated protein kinase A contributes to early afterdepolarizations in the heart

Abstract

Reactive oxygen species (ROS) have been shown to prolong cardiac action potential duration resulting in afterdepolarizations, the cellular basis of triggered arrhythmias. As previously shown, protein kinase A type I is readily activated by oxidation of its regulatory subunits. However, the relevance of this mechanism of activation for cardiac pathophysiology is still elusive. In this study, we investigated the effects of oxidation-activated PKA I on cardiac electrophysiology. Ventricular cardiomyocytes were isolated from redox-dead PKA-RI Cys17Ser knock-in (KI) and wild-type (WT) mice and exposed to H2O2 (200 µmol/L) or vehicle (veh) solution. In WT myocytes, exposure to H2O2 significantly increased oxidation of the regulatory subunit I (RI) and thus its dimerization (3-fold increase in PKA RI dimer). Whole cell current-clamp and voltage clamp was used to measure cardiac action potentials (APs), transient outward potassium current (Ito) and inward rectifying potassium current (IK1), respectively. In WT myocytes, H2O2 exposure significantly prolonged AP duration due to significantly decreased Ito and IK1 resulting in frequent early afterdepolarizations (EADs). Pre-incubation with the PKA-specific inhibitor Rp-8-Br-cAMPS (10 µmol/L) completely abolished the H2O2-dependent decrease in Ito and IK1 in WT myocytes. Intriguingly, H2O2 exposure did not prolong AP duration, nor did it decrease Ito, and only slightly enhanced EAD frequency in KI myocytes. Treatment of WT and KI cardiomyocytes with the late INa inhibitor TTX (1µmol/L) completely abolished EAD formation. Our results suggest that redox-activated PKA may be important for H2O2-dependent arrhythmias and could be important for development of specific antiarrhythmic drugs.

Regulation of Cardiac PKA Signaling by cAMP and Oxidants

“…This assumption was indeed well founded, as the N-terminal D/D-domain of PKA-R that in the type I isoform harbors Cys17 and Cys38 does not only mediate regulatory subunit dimerization, but also the interaction with AKAPs ( Figure 2 ). In line with these results, the data of various studies showed subcellular translocation of PKA into the myofilament containing fraction of adult rat ventricular myocytes after exposure to NCA [ 44 ], to the lysosomal two pore channel during ischemia/reperfusion in the ventricle [ 48 ] or into cation channel vicinity to trigger arrhythmias upon oxidant-exposure [ 49 ]. A potential impact of PKA-RI disulfide formation on the interaction with AKAPs had previously been investigated for D-AKAP1, the first AKAP that was found to not exclusively bind to type II, but to be dual-specific and also interact with type I PKA regulatory subunits [ 50 ].…”

“…These data are in accordance with our study that rather than impacting directly on kinase activity, emphasized an important role of interdisulfide formation in PKA-RI for oxidant-mediated translocation into substrate vicinity. In the atria, PKA-RI dimer formation upon oxidant-exposure has been described to contribute to the prolongation of cardiac action potential duration and occurrence of after depolarizations due to inhibition of I to and I K1 [ 49 ]. Although the authors described increased oxidant-mediated disulfide formation leading to increased PKA activity, they did not assess kinase activity.…”

Cardioprotection by SGLT2 Inhibitors—Does It All Come Down to Na+?

“…Oxidative stress is a hallmark of cardiac disease including heart failure [ 108 , 109 ] and centrally involved in pathways promoting hypertrophy, fibrosis, cell death, as well as directly or indirectly (e.g., via oxidation of different kinases) involved in perturbation of cellular ion homeostasis [ 25 , 54 , 110 ]. Cytosolic Na + overload results in an extrusion of Ca 2+ from the mitochondria mediated by the NCLX.…”

“…Typically, EADs result from prolongation of the APD allowing for the reactivation of LTCC. APD prolongation in turn is caused either by a decrease in repolarizing currents (i.e., outward K + currents) and/or an increase in depolarizing currents (e.g., late I Na ) [ 23 , 40 , 110 , 138 ]. On the other hand, DADs result from spontaneous diastolic Ca 2+ release from the SR via RyR2, which activates a transient inward current that is mainly mediated by NCX.…”

Dynamic Regulation of Cysteine Oxidation and Phosphorylation in Myocardial Ischemia–Reperfusion Injury

“…While these indicate oxidant-activated PKA may be detrimental to the heart, in certain contexts, PKA oxidation may help to remedy arrhythmias by inhibiting potassium currents. Treating a “redox dead” PKA with H 2 O 2 and failing to prolong action potentials or alter potassium currents has been previously reported [ 150 ]. Genetic polymorphisms in PKA can alter PDE and AKAP binding, thus altering PKA activity and cardiovascular disease susceptibility [ 151 , 152 , 153 ].…”