Fig. 1.
Genetic deletion of Trpm7 causes loss of classical TRPM7 current. (A) Real-time RT-PCR of total RNA from Trpm7f/f (control) or Trpm7f/f;Gdf9-cre (Trpm7 cKO) eggs. Schematic shows partial genomic structure of Trpm7-floxed allele. Exons, blue boxes; loxP sites, red triangles; primer locations, gray arrows. Graph shows expression relative to that in control eggs, mean ± SEM. Eggs were collected from n = 5 individual control and n = 3 Trpm7 cKO mice. *P < 0.05 relative to control with same primer pair, t test. (B) Whole-cell patch-clamp recordings from eggs of control or Trpm7 cKO mice in response to a ramp (−100 to +100 mV). Graph shows current amplitude from individual eggs at +90 mV before (black symbols) and after (red symbols) addition of NS8593; bars indicate mean ± SEM. Control eggs, n = 8; Trpm7 cKO eggs, n = 5. *P < 0.05, paired t test. (C) Ca2+ response of control and Trpm7 cKO oocytes to the indicated concentrations of naltriben. Upper graphs show F340/F380 ratio in individual oocytes (gray) and the mean ratio (black) from both groups in one representative experiment. Lower graphs show mean area under the curve ± SEM for the indicated treatment. n = 3 biological replicates, a total of 18 control and 18 Trpm7 cKO oocytes were imaged. *P < 0.05, Mann–Whitney U test (40 μM naltriben); *P < 0.05, t test with Welch’s correction (80 μM naltriben). (D) Ca2+ response of control and Trpm7 cKO oocytes to the indicated concentrations of mibefradil (mib). Upper graphs in this panel and in all subsequent calcium traces show F340/F380 ratio in individual oocytes (gray) and the mean ratio (black) from both groups in one representative experiment. Lower graphs show mean area under the curve ± SEM for the indicated treatment. n = 2 biological replicates, a total of 23 control and 23 Trpm7 cKO oocytes were imaged. *P < 0.05, t test with Welch’s correction.