Insulin Modulates the Na+/Mg2+ Exchanger SLC41A1 and Influences Mg2+ Efflux from Intracellular Stores in Transgenic HEK293 Cells

Background: Magnesium deficiency is a common complication of diabetes with an unclear molecular background. Objective: We investigated the effect of the insulin (INS)-signaling pathway (ISP) on the regulation of Mg2+ efflux (Mg2+E) conducted by solute carrier family 41, member A1 (SLC41A1; activated by protein kinase A) in transgenic human embryonic kidney (HEK) 293 cells. Methods: HEK293 cells overexpressing SLC41A1 were loaded with the Mg2+ fluorescent indicator mag-fura-2 and Mg2+. Measurements of Mg2+E were conducted in Mg2+-free buffer by using fast-filter fluorescence spectrometry. We examined the effects of INS, inhibitors of ISP or p38 mitogen-activated protein kinase (p38 MAPK), an activator of adenylate cyclase (ADC), and their combinations on SLC41A1-attributed Mg2+E. Results: The application of 400 mu U/mL INS inhibited SLC41A1-mediated Mg2+E by up to 50.6% compared with INS-untreated cells (P < 0.001). Moreover, INS evoked the early onset of Mg2+ release from intracellular stores. The application of 0.1 mu M wortmannin or 10 mu M zardaverine (both ISP inhibitors) restored SLC41A1 Mg2+E capacity in the presence of INS to the same levels in INS-untreated cells. The simultaneous application of 10 mu M forskolin, an ADC activator, and INS resulted in a reduction of Mg2+E of up to 59% compared with untreated cells (P < 0.001), which was comparable to that in cells treated with INS alone. Inhibition of p38 MAPK with 10 mu M SB 202190 (SB) in the absence of INS resulted in a decrease (P < 0.001) of SLC41A1-dependent Mg2+E (by up to 49%) compared with Mg2+E measured in untreated cells. Simultaneous exposure of cells to SB and INS had a stronger inhibitory effect on SLC41A1 activity than INS alone (P < 0.05). Conclusions: INS affects intracellular Mg2+ concentration in transgenic HEK293 cells by regulating SLC41A1 activity (via ISP) and by influencing the compartmentalization and cellular distribution of Mg2+. In addition, p38 MAPK activates SLC41A1 independently of INS action.