Effects of γ-Ketoaldehydes on Ca2+ Cuttrent Induced SR Ca2+ Release in Ventricular Myocytes

Abstract

• Oxidation increases RyR2 channel activity, enhances cardiac SR Ca2+ release and causes spontaneous SR Ca2+ waves. Isoprostanes have become a recognized marker of oxidative stress in rodents and humans. γ-ketoaldehydes(γ-KAs) are the most reactive product of the isoprostane pathway. Recently, we found that lipophilic pyridoxamine analogues, salicylamine(SA) scavenge γ-KAs and thereby prevent formation of γ-KA protein adducts in response to oxidative stress. We hypothesized that γ-KAs are potential mediators of oxidant-induced RyR2 channel dysfunction and spontaneous SR Ca2+ waves, and that SA would prevent oxidant-induced spontaneous SR Ca2+ waves(SCW) in the ventricular myocytes. • We compared the effect of γ-KAs(1uM) or H2O2(10uM) and the effect of SA on Ca-current induced Ca release(CICR) in murine ventricular myocytes loaded with Fura-2AM or Fluo-4. All data are expressed relative to vehicle(Mean±SEM, n=15-50 per group). • Acute exposure(3 min) to γ-KAs(1 uM) or H2O2(10 uM) increased the amplitude of Ca2+ transients, and the fraction of Ca2+ released from the SR(γ-KAs130±10%∗, H2O2120±10%, ∗p<0.05) during each beat. Furthermore, the rate of SCW was significantly increased(γ-KAs 42%∗, H2O233%∗, ∗p<0.05) and SR Ca2+ content was reduced. In voltage-clamped myocytes, dialysis with γ-KAs enhanced Ca2+ release without changing L-type Ca2+ current, demonstrating that the effect of γ-KAs is the result of RyR2 modification. However, after chronic exposure(30 min) to γ-KAs(1 uM) or H2O2(10 uM), Ca2+ transients(γ-KAs 0.53±0.1∗, H2O2 0.7±0.1∗, ∗p<0.05) and SR Ca2+ contents decreased, and SCW remained elevated. Pre-treatment(3 days) of salicylamine reduced H2O2-induced spontaneous Ca2+ waves(SCWs/sec, H2O21.2±0.3∗, SA-H2O20.4±0.2∗, ∗p<0.05) preserved with SR Ca2+ content in ventricular myocytes. • We found that H2O2 and γ-KAs have analogous biphasic effects on SR Ca2+ release in ventricular myocytes. The protective effect of γ-KA scavengers suggests that γ-KAs are possible mediators of oxidant-induced RyR2 channel dysfunction.