We examined the structure–activity relationship of cloned lysophosphatidic acid (LPA) receptors (endothelial cell differentiation gene (EDG) 2, EDG4, and EDG7) by measuring [Ca²⁺]_i in Sf9 insect cells expressing each receptor using LPA with various acyl chains bound at either the sn‐1 or the sn‐2 position of the glycerol backbone. For EDG7 the highest reactivity was observed with LPA with Δ9‐unsaturated fatty acid (oleic (18:1), linoleic (18:2), and linolenic (18:3)) at sn‐2 followed by 2‐palmitoleoyl (16:1) and 2‐arachidonoyl (20:4) LPA. In contrast, EDG2 and EDG4 showed broad ligand specificities, although EDG2 and EDG4 discriminated between 14:0 (myristoyl) and 16:0 (palmitoyl), and 12:0 (lauroyl) and 14:0 LPAs, respectively. EDG7 recognizes the cis double bond at the Δ9 position of octadecanoyl residues, since 2‐elaidoyl (18:1, trans) and 2‐petroselinoyl (18:1, cis‐Δ12) LPA were poor ligands for EDG7. In conclusion, the present study demonstrates that each LPA receptor can be activated differentially by the LPA species.