Luminal Ca2+ regulation of RyR1 Ca2+ channel leak activation and inactivation in sarcoplasmic reticulum membrane vesicles.

In this study, we tested the hypothesis that the RyR1 Ca2+ channel closure is sensitive to outward trans-SR membrane Ca2+ gradients established by SERCA1 pumping. To perform these studies, we employed stopped-flow rapid-kinetic fluorescence methods to measure and assess how variation in trans-SR membrane Ca2+ distribution affects evolution of RyR1 Ca2+ leaks in RyR1/ CASQ1/SERCA1-rich membrane vesicles. Our studies showed that rapid filling of a Mag-Fura-2-sensitive free Ca2+ pool during SERCA1-mediated Ca2+ sequestration appears to be a crucial condition allowing RyR1 Ca2+ channels to close once reloading of luminal Ca2+ stores is complete. Disruption in the filling of this pool caused activation of Ruthenium Red inhibitable RyR1 Ca2+ leaks, suggesting that SERCA1 pump formation of outward Ca2+ gradients is an important aspect of Ca2+ flux control channel opening and closing. In addition, our observed ryanodine-induced shift in luminal Ca2+ from free to a CTC-Ca+-sensitive, CASQ1-associated bound compartment underscores the complex organization and regulation of SR luminal Ca2+. Our study provides strong evidence that RyR1 functional states directly and indirectly influence the compartmentation of luminal Ca2+. This, in turn, is influenced by the activity of SERCA1 pumps to fill luminal pools while synchronously reducing Ca2+ levels on the cytosolic face of RyR1 channels.

Effects of dietary flaxseed on vascular contractile function and atherosclerosis during prolonged hypercholesterolemia in rabbits.

Dietary flaxseed has significant anti-atherogenic effects. However, the limits of this action and its effects on vascular contractile function are not known. We evaluated the effects of flaxseed supplementation on atherosclerosis and vascular function under prolonged hypercholesterolemic conditions in New Zealand White rabbits assigned to one of four groups for 6, 8, or 16 wk of feeding: regular diet (RG), 10% flaxseed-supplemented diet (FX), 0.5% cholesterol-supplemented diet (CH), and 0.5% cholesterol- and 10% flaxseed-supplemented diet (CF). Cholesterol feeding resulted in elevated plasma cholesterol levels and the development of atherosclerosis. The CF group had significantly less atherosclerotic lesions in the aorta and carotid arteries after 6 and 8 wk than the CH animals. However, the anti-atherogenic effect of flaxseed supplementation was completely attenuated by 16 wk. Maximal tension induced in aortic rings either by KCl or norepinephrine was not impaired by dietary cholesterol until 16 wk. This functional impairment was not prevented by including flaxseed in the high-cholesterol diet. Aortic rings from the cholesterol-fed rabbits exhibited an impaired relaxation response to acetylcholine at all time points examined. Including flaxseed in the high-cholesterol diet completely normalized the relaxation response at 6 and 8 wk and partially restored it at 16 wk. No significant changes in the relaxation response induced by sodium nitroprusside were observed in any of the groups. In summary, dietary flaxseed is a valuable strategy to limit cholesterol-induced atherogenesis as well as abnormalities in endothelial-dependent vasorelaxation. However, these beneficial effects were attenuated during prolonged hypercholesterolemic conditions.