Opiate receptor expression in phylogenetically different species has played an important role in the study of opioid receptor pharmacology. Total opioid binding measured with the nonselective ligand 3H-diprenorphine reveals a Bmax of 21.7 +/- 1.37 fmol/mg tissue wet wt and a KD of 0.17 +/- 0.03 nM in Bufo marinus (giant toad), as well as a Bmax of 18.17 + 0.41 fmol/mg tissue wet wt and a KD of 0.47 +/- 0.18 nM in Carassius auratus (goldfish). Despite the similar levels of 3H-diprenorphine binding, the composition of binding subtypes in the two species differs. Approximately 30% of total binding corresponds to mu receptors in both species, whereas neither kappa 1 nor delta binding can be detected. However, the remaining 70% of binding differs between the toad and goldfish. In the toad, the non-mu binding corresponds to kappa 2 sites, whereas in the goldfish, the non-mu binding corresponds to kappa 3 sites. The sites can be distinguished biochemically, as well as pharmacologically. After affinity labeling the sites with 3H-NalBzoH, the retention times on an ion-exchange column differ for the peaks of kappa binding in the two species. Although Bufo marinus (giant toad) and Carassius auratus (goldfish) brains express kappa and mu opioid binding, the kappa subtypes in these two species differ.