Abstract: Mg2+‐deficiency is linked to hypertension, Alzheimer's disease, stroke, migraine headaches, cardiovascular diseases, and diabetes, etc., but its exact role in these pathophysiological conditions remains elusive. Mg2+ can regulate vascular functions, yet the mechanistic insight remains ill‐defined. Data show that extracellular Mg2+ enters endothelium mainly through the TRPM7 channel and MagT1 transporter. Mg2+ can act as an antagonist to reduce Ca2+ signaling in endothelium. Mg2+ also reduces the intracellular reactive oxygen species (ROS) level and inflammation. In addition, Mg2+‐signaling increases endothelial survival and growth, adhesion, and migration. Endothelial barrier integrity is significantly enhanced with Mg2+‐treatment through S1P1‐Rac1 pathways and barrier‐stabilizing mediators including cAMP, FGF1/2, and eNOS. Mg2+ also promotes cytoskeletal reorganization and junction proteins to tighten up the barrier. Moreover, Mg2+‐deficiency enhances endothelial barrier permeability in mice, and Mg2+‐treatment rescues histamine‐induced transient vessel hyper‐permeability in vivo. In summary, Mg2+‐deficiency can cause deleterious effects in endothelium integrity, and Mg2+‐treatment may be effective in the prevention or treatment of vascular dysfunction.