Fig 1: Mg2+ promotes neurite outgrowth in a concentration-dependent manner by activating the PI3K/Akt signaling pathway and upregulating Sema5b expression. A–F) Primary culture of DRG neurons treated with varying Mg2+ concentrations (Ctrl, 10, 20, 30, 40, and 50 mM). G) Gene expression heatmap of axon guidance molecules and neurotrophic factors (n = 3). H) mRNA expression levels of NGF, Sema3b, Sema3e, Sema4c, Sema5b, NetrinG1, EFNA2, EFNB3 (mean ± standard deviation; one-way ANOVA and Tukey's post hoc tests, *P < 0.05; **P < 0.01 compared to the mRNA levels in the Ctrl group; n = 3 per group). I) Protein expression levels of NF200, GAP43, Sema5b, and GAPDH assessed by western blotting. J) Primary culture of DRG neurons treated with additional Mg2+ ions, LY294002, or DMSO. K) Protein expression levels of pPI3K, Akt, and p-Akt in four groups (Ctrl, Mg2+, Mg2+ with LY294002, and DMSO). L) Primary culture of DRG neurons with additional Mg2+, Lipo: Sema5b-siRNA or Lipo: Blank; M) Western blot results of GAP43, Sema5b expression in four groups. DRG, dorsal root ganglia; DMSO, dimethyl sulfoxide; Ctrl, control.