Shear stress enhances Ca2 + sparks through Nox2-dependent mitochondrial reactive oxygen species generation in rat ventricular myocytes

Abstract

Shear stress enhances diastolic and systolic Ca2+ concentration in ventricular myocytes. Here, using confocal Ca2+ imaging in rat ventricular myocytes, we assessed the effects of shear stress (~16dyn/cm2) on the frequency of spontaneous Ca2+ sparks and explored the mechanism underlying shear-mediated Ca2+ spark regulation. The frequency of Ca2+ sparks was immediately increased by shear stress (by ~80%), and increased further (by ~150%) during prolonged exposure (20s). The 2-D size and duration of individual sparks were increased by shear stimulation. Inhibition of nitric oxide synthase (NOS) only partially attenuated the prolonged shear-mediated enhancement in spark frequency. Pretreatment with antioxidants significantly attenuated the short- and long-term effects of shear on spark frequency. Microtubule or nicotinamide adenine dinucleotide phosphate oxidase 2 (Nox2) inhibition abolished the immediate shear-induced increase in spark frequency and suppressed the effects of prolonged exposure to shear stress by ~70%. Scavenging of mitochondrial reactive oxygen species (ROS) and mitochondrial uncoupling also abolished the effect of short-term shear on spark occurrence, and markedly reduced (by ~80%) the effects of prolonged shear. Mitochondrial ROS levels increased under shear; this was eliminated by blocking Nox2. Sarcoplasmic reticulum Ca2+ content was increased only by prolonged shear. Our data suggest that shear stress enhances ventricular spark frequency mainly via ROS generated from mitochondria through Nox2, and that NOS and higher sarcoplasmic reticulum Ca2+ concentrations may also contribute to the enhancement of Ca2+ sparks under shear stress. This article is part of a Special Issue entitled: ECS Meeting edited by Claus Heizmann, Joachim Krebs and Jacques Haiech.