Ligand interactions of a zebrafish bradykinin (BK) receptor expressed in vitro were characterized by measuring inositol phosphate accumulation. The ligands were analogues of zebrafish BK. Substitutions of Arg1, Gly4, Ser6, Pro7, Leu8, and Arg9 caused greatly reduced potency and maximum response. The Pro3 to Ala analogue had higher potency, but lower maximum response. These and other differences show that the zebrafish BK receptor has a ligand-interaction profile that is distinct from mammalian B1 and B2 receptors and from the previously characterized BK receptor in trout stomach. The results increase our understanding of the evolution of BK receptors and their ligands.