Functional expression of the M7cd in HEK293 cells. (A) Time course of M7cd currents in the absence (open circles; n = 15) or presence of 2.8 µM (blue squares; n = 9), 20.9 µM (green triangles; n = 14), and 0.2 mM [Mg²⁺]_i (black circles; n = 16) in HEK293 cells that were treated with doxycycline. Each symbol represents the mean ± SEM (vertical bar). (B) Representative I-V relationship of M7cd current recorded in the presence of various [Mg²⁺]_i at 2 min after break-in. (C) [Mg²⁺]_i-dependent inhibition of M7cd current at 2 min. The current was inhibited by intracellular free Mg²⁺ with an IC₅₀ of 3.0 µM. Each symbol represents the mean ± SEM (vertical bar) of 15, 36, 19, 27, 14, and 20 recordings in the absence or presence of 0.8 µM, 2.8 µM, 7.4 µM, 20.9 µM, and 0.2 mM [Mg²⁺]_i, respectively. The dashed line is the [Mg²⁺]_i-dependent curve of the full-length TRPM7-wt current (from Fig. 1 D). (D) M7cd-S1107E current density was similar in the absence (open inversed triangles; n = 10) and presence of 0.2 mM [Mg²⁺]_i (closed inversed triangles; n = 7). Each symbol represents the mean ± SEM (vertical bar). (E) The effect of H₂O₂ (500 µM) on M7cd current in the absence of intracellular Mg²⁺ (open circles). Each symbol represents the mean ± SEM (vertical bar) of 19 recordings. (F) The time course of the M7cd current in the presence of 0.8 µM [Mg²⁺]_i with (red circles; n = 17) or without (black circles; n = 35) H₂O₂ application at 2 min. Each symbol represents the mean ± SEM (vertical bar). The current decreased over time even in the control conditions (black circles), and repeated-measures ANOVA revealed that H₂O₂ has no significant effect on the current decrease (degrees of freedom were corrected using the Greenhouse-Geisser estimate of epsilon; P = 0.247).