Lysophosphatidic acid (LPA) is known to have diverse cellular effects, but although LPA is present in many biological fluids, including blood, its effects on glucose metabolism have not been elucidated. In this study, we investigated whether LPA stimulation is related to glucose regulation. LPA was found to enhance glucose uptake in a dose-dependent manner both in L6 GLUT4myc myotubes and 3T3-L1 adipocytes by triggering GLUT4 translocation to the plasma membrane. Moreover, the effect of LPA on glucose uptake was completely inhibited by pretreating both cells with LPA receptor antagonist Ki16425 and Gi inhibitor pertussis toxin. In addition, LPA increased the phosphorylation of AKT-1 with no effects on IRS-1, and LPA-induced glucose uptake was abrogated by pretreatment with the PI 3-kinase inhibitor LY294002. When low concentration of insulin and LPA were treated simultaneously, an additive effect on glucose uptake was observed in both cell types. In line with its cellular functions, LPA significantly lowered blood glucose levels in normal mice but did not affect insulin secretion. LPA also had a glucose-lowering effect in streptozotocin-treated type 1 diabetic mice. In combination, these results suggest that LPA is involved in the regulation of glucose homeostasis in muscle and adipose tissues.