Late INa increases diastolic SR-Ca2+-leak in atrial myocardium by activating PKA and CaMKII

Abstract

AimsEnhanced cardiac late Na current (late INa) and increased sarcoplasmic reticulum (SR)-Ca2+-leak are both highly arrhythmogenic. This study seeks to identify signalling pathways interconnecting late INa and SR-Ca2+-leak in atrial cardiomyocytes (CMs).Methods and resultsIn murine atrial CMs, SR-Ca2+-leak was increased by the late INa enhancer Anemonia sulcata toxin II (ATX-II). An inhibition of Ca2+/calmodulin-dependent protein kinase II (Autocamide-2-related inhibitory peptide), protein kinase A (H89), or late INa (Ranolazine or Tetrodotoxin) all prevented ATX-II-dependent SR-Ca2+-leak. The SR-Ca2+-leak induction by ATX-II was not detected when either the Na+/Ca2+ exchanger was inhibited (KBR) or in CaMKIIδc-knockout mice. FRET measurements revealed increased cAMP levels upon ATX-II stimulation, which could be prevented by inhibition of adenylyl cyclases (ACs) 5 and 6 (NKY 80) but not by inhibition of phosphodiesterases (IBMX), suggesting PKA activation via an AC-dependent increase of cAMP levels. Western blots showed late INa-dependent hyperphosphorylation of CaMKII as well as PKA target sites at ryanodine receptor type-2 (-S2814 and -S2808) and phospholamban (-Thr17, -S16). Enhancement of late INa did not alter Ca2+-transient amplitude or SR-Ca2+-load. However, upon late INa activation and simultaneous CaMKII inhibition, Ca2+-transient amplitude and SR-Ca2+-load were increased, whereas PKA inhibition reduced Ca2+-transient amplitude and load and additionally slowed Ca2+ elimination. In atrial CMs from patients with atrial fibrillation, inhibition of late INa, CaMKII, or PKA reduced the SR-Ca2+-leak.ConclusionLate INa exerts distinct effects on Ca2+ homeostasis in atrial myocardium through activation of CaMKII and PKA. Inhibition of late INa represents a potential approach to attenuate CaMKII activation and decreases SR-Ca2+-leak in atrial rhythm disorders. The interconnection with the cAMP/PKA system further increases the antiarrhythmic potential of late INa inhibition.

The functional consequences of sodium channel NaV1.8 in human left ventricular hypertrophy

“…Inhibition of Na V 1.8 with specific blockers showed a significant abbreviation of APD. 37 An increase in diastolic Ca 2+ not only results in impaired relaxation but also plays a key role in activating pro-hypertrophic signalling pathways, leading to increased stiffness of the left ventricle. This prolonged APD can give rise to early after-depolarizations, thereby posing increased fatal risk for ventricular arrhythmias.…”

Triggered activity in atrial myocytes is influenced by Na+/Ca2+ exchanger activity in genetically altered mice

“…Spontaneous Ca 2+ activity leads to a sudden elevation of the cytosolic Ca 2+ concentration driving NCX forward mode, resulting in a DAD. Among other factors, increased SR Ca 2+ load[28], altered RyR expression or function[29, 30], altered Na + cycling[31, 32] and altered oxidation[33–35] increase spontaneous activity thus increasing the number of DADs which will result in proarrhythmia. In NCX KO and OE, SR Ca 2+ load, RyR receptor expression and phosphorylation (Ser2808) were unaltered.…”

The Significance of the Late Na+ Current for Arrhythmia Induction and the Therapeutic Antiarrhythmic Potential of Ranolazine

“…34 Sodium ion influx via I Na,late also leads to APD prolongation. 12,13,16,35 When sufficient APD prolongation occurs, inward currents are able to recover from inactivation, become reactivated, and allow charge to enter the cell. If current entering the cell exceeds repolarizing K þ currents, an EAD can occur.…”

“…17,29,[43][44][45] The resulting calcium overload is thought to trigger intracellular Ca 2þ release from the sarco-endoplasmic reticulum (SR), leading to cytosolic Ca 2þ oscillations, automaticity, and triggered activity. 16,35,[46][47][48] Calcium overload and oscillatory activity are able to activate forward mode NCX. 49 This carries an inward positive current, also known as the transient inward current due to NCX stoichiometry…”

“…91 Ryanodine receptor instability and increased SR Ca 2þ leak are also considered to play an important role in AF 50,87,100 with CaMKII, again implicated as a significant mediator. 35,87 It is becoming increasingly clear that upregulated I Na,late , intracellular Na þ overload, and increased activity of CaMKII are not acting alone, but rather in concert in cardiac disease. Acute overexpression of CaMKIId c has been shown to facilitate I Na,late by altering channel gating.…”