Reduction of intracellular Mg²⁺ caused by reactive oxygen species in rat ventricular myocytes

The concentration of intracellular free Mg²⁺ ([Mg²⁺]_i) should be maintained strictly for the regulation of cellular functions. Since reactive oxygen species (ROS) are liable to increase in various pathological conditions and induce cellular damage, we investigated whether ROS affect intracellular Mg²⁺ homeostasis. We measured [Mg²⁺]_i in ventricular myocytes from Wistar rats using the fluorescent indicator, mag-fura-2. The administration of hydrogen peroxide (H₂O₂) decreased [Mg²⁺]_i in Ca²⁺-free Tyrode's solution. Intracellular free Mg²⁺ was also reduced by endogenous ROS as generated by pyocyanin, which was inhibited by pretreatment with n-acetyl cysteine (NAC). The rate of change in [Mg²⁺]_i by 500 μM H₂O₂ in 5 min (on average, -0.61 μM/s) was independent of extracellular Na⁺, and intra- and extracellular Mg²⁺ concentrations. When extracellular Ca²⁺ was present, the rate of Mg²⁺ decrease was significantly reduced, on average, by ∼60%. The half-maximal effective concentration of H₂O₂ on the Mg²⁺ decrease was estimated to be between 400 and 425 μM. The Mg²⁺ decrease by H₂O₂ in the absence of Na⁺ was inhibited by 200 μM imipramine, a known inhibitor of Na⁺/Mg²⁺ exchange. We perfused rat hearts with the Ca²⁺-free Tyrode's solution containing H₂O₂ (500 μM, 5 min) on the Langendorff apparatus,_. H₂O₂ stimulation increased Mg²⁺ concentration in the perfusate, suggesting the H₂O₂-induced decrease in [Mg²⁺]_i was caused by Mg²⁺ extrusion. Collectively, these results suggest the existence of a Na⁺-independent Mg²⁺ efflux system activated by ROS in cardiomyocytes. The lower [Mg²⁺]_i may in part be attributed to ROS-mediated cardiac dysfunction.