Differential effects of the peroxynitrite donor, SIN-1, on atrial and ventricular myocyte electrophysiology

Abstract

Oxidative stress has been implicated in the pathogenesis of heart failure and atrial fibrillation and can result in increased peroxynitrite production in the myocardium. Atrial and ventricular canine cardiac myocytes were superfused with 3-morpholinosydnonimine-N-ethylcarbamide (SIN-1), a peroxynitrite donor, to evaluate the acute electrophysiologic effects of peroxynitrite. Perforated whole-cell patch clamp techniques were used to record action potentials. SIN-1 (200 µM) increased the action potential duration (APD) in atrial and ventricular myocytes; however, in the atria, APD prolongation was rate independent, whereas in the ventricle APD, prolongation was rate dependent. In addition to prolongation of the action potential, beat-to-beat variability of repolarization was significantly increased in ventricular but not in atrial myocytes. We examined the contribution of intracellular calcium cycling to the effects of SIN-1 by treating myocytes with the SERCA blocker, thapsigargin (5-10 µM). Inhibition of calcium cycling prevented APD prolongation in the atrial and ventricular myocytes, and prevented the SIN-1-induced increase in ventricular beat-to-beat APD variability. Collectively, these data demonstrate that peroxynitrite affects atrial and ventricular electrophysiology differentially. A detailed understanding of oxidative modulation of electrophysiology in specific chambers is critical to optimize therapeutic approaches for cardiac diseases.

Figure 1

SIN-1 prolongs the atrial and ventricular action potential. (A) The peroxynitrite scavenger uric acid was used to evaluate specificity of SIN-1. Atrial action potentials were recorded at baseline, with uric acid and with uric acid + SIN-1. No statistical change in atrial APD₉₀ was observed after co-superfusion. (B) SIN-1 prolongs the atrial action potential and (C) SIN-1 prolongs the action potential and decreases the phase 1 notch in ventricular myocytes.

Figure 2

SIN-1 affects atrial and ventricular repolarization. (A) SIN-1 causes a significant (p<0.05 vs baseline) prolongation in the atrial APD₅₀ only at 2 Hz while the APD₉₀ is significantly increased (p<0.05 vs baseline) after SIN-1 in a rate independent manner. (B) Contrary to the atria, no change was observed in the ventricular APD₅₀ after SIN-1. At baseline ventricular APD₉₀ was significantly shorter at 2 Hz than 0.5 Hz. A significant increase (p<0.05 vs baseline) was observed in the ventricular APD₉₀ at all rates during SIN-1 superfusion. While SIN-1 prolonged the ventricular APD₉₀, in contrast to the atria, APD₉₀ retains rate-dependence, with APD₉₀ being significantly shorter at 2 Hz, compared to 0.5 or 1 Hz (P<0.05). (C) SIN-1 does not affect atrial beat to beat variability of repolarization (SD APD₉₀). (D) A significant (p<0.05 vs baseline) increase in beat to beat variability of repolarization (SD APD₉₀) occurs during SIN-1 superperfusion in ventricular myocytes at 0.5 and 1 Hz, but not at 2 Hz. (n=5-8 cells in each group, N= 3-4 animals per group).

Figure 3

Peroxynitrite-induced changes in atrial and ventricular repolarization and variability of repolarization are prevented with thapsigargin pre-treatment. (A) and (B) Thapsigargin pre-treatment (to deplete SR calcium stores and prevent calcium cycling) prevented both atrial and ventricular SIN-1 dependent APD₉₀ prolongation (p>0.05 from baseline). (C) Atrial beat to beat variability in repolarization is unchanged by peroxynitrite (p=NS vs baseline). (D) Peroxynitrite-induced increase in ventricular beat to beat variability in repolarization is prevented by thapsigargin (p=NS vs baseline). (n=5-8 cells in each group and N= 3-5 animals per group)