Modulation of Mg²⁺ influx and cytoplasmic free Mg²⁺ concentration in rat ventricular myocytes

To examine whether TRPM7, a member of the melastatin family of transient receptor potential channels, is a physiological pathway for Mg²⁺ entry in mammalian cells, we studied the effect of TRPM7 regulators on cytoplasmic free Mg²⁺ concentration ([Mg²⁺]_i) of rat ventricular myocytes. Acutely isolated single cells were AM-loaded with the fluorescent indicator furaptra, and [Mg²⁺]_i was estimated at 25 °C. After [Mg²⁺]_i was lowered by soaking the cells with a high-K⁺ and Mg²⁺-Ca²⁺-free solution, [Mg²⁺]_i was recovered by extracellular perfusion of Ca²⁺-free Tyrode's solution that contained 1 mM Mg²⁺. The initial rate of increase in [Mg²⁺]_i was analyzed as the Mg²⁺ influx rate. The Mg²⁺ influx rate was increased by the TRPM7 activator, naltriben (2-50 μM), in a concentration-dependent manner with a half maximal effective concentration (EC₅₀) of 24 μM. This EC₅₀ value is similar to that reported for the activation of recombinant TRPM7 overexpressed in HEK293 cells. Naltriben (50 μM) caused little change in basal [Mg²⁺]_i (~ 0.9 mM) in Ca²⁺-free Tyrode's solution, but significantly raised [Mg²⁺]_i to 1.31 ± 0.03 mM in 94 min after the removal of extracellular Na⁺. Re-introduction of extracellular Na⁺ lowered [Mg²⁺]_i back to the basal level even in the presence of naltriben. Application of 10 μM NS8593, an inhibitor of TRPM7, significantly lowered [Mg²⁺]_i to 0.72 ± 0.03 mM in 50-60 min independent of extracellular Na⁺. The results suggest that Mg²⁺ entry through TRPM7 significantly contributes to physiological Mg²⁺ homeostasis in mammalian heart cells.