Iron (II) Modulation of the Cardiac Ryanodine Receptor (RyR2)

Abstract

Cardiomyopathies and arrhythmias are major causes of mortality in chronic iron overload. There is evidence that iron overload impairs cardiomyocytes Ca2+ homeostasis (Baptista-Hon et al, 2005). However its molecular substrates remain unknown. Cardiac ryanodine receptors (RyR2) dysfunction is implicated in several diseases where Ca2+ homeostasis is lost. We therefore wanted to investigate RyR2 role as a potential target for iron-induced cardiomyopathies.We isolated heavy sarcoplasmic reticulum (HSR) vesicles containing RyR2 from sheep hearts. RyR2 were reconstituted into L-α-phosphatidylethanolamine (PE) bilayers. Unitary currents were measured under voltage-clamp with 250mM Cs+ as the charge carrier and 10μM activating Ca2+. High affinity [3H]ryanodine binding of the native vesicles was detected by liquid scintillation counting. Non-specific binding determined by incubations with 100x cold ryanodine.Fe2+ reduced RyR2 open probability and conductance in a dose dependent manner. Lifetime analysis revealed 5 shut times components and 3 open times components in control. Fe2+ caused an extra-shut component. Furthermore, there was a dose dependent shift in the open time constants towards the faster components. The binding assays revealed a [Fe2+] dependent, co-operative reduction in [3H]ryanodine binding to HSR vesicles. Preliminary data of [3H]ryanodine binding in increasing [Ca2+] showed a rightward shift in the presence of Fe2+.The results presented here show for the first time that Fe2+ is a potent inhibitor of RyR2. The mechanism of this inhibition may be due to competition with Ca2+ for RyR2 activation sites. Suppression of RyR2 activity by Fe2+ may therefore be one of the mechanisms involved in iron-induced cardiomyopathies.ReferencesBaptista-Hon, D, Diaz, M. E., and Elliot, A. C. Acute exposure to iron (II) alters calcium handling in isolated rat ventricular myocytes. Journal of Molecular and Cellular Cardiology 39, 179. 2005.