The specific binding of [3H]ethylketocyclazocine to frog brain membrane preparation was enhanced in the presence of sodium ions administered as NaCl, both at 0 degree C and at room temperature. The optimal NaCl concentration was 25 mM at 0 degree C and 50 mM at 24 degrees C. MgCl2 inhibited the [3H]ethylketocyclazocine binding. Two binding sites (high and low affinity) were established with [3H]ethylketocyclazocine as ligand by equilibrium binding studies. Addition of NaCl increased the Bmax of the low-affinity site more than that of the high-affinity site at both temperatures. Affinities were higher at 0 degree C than at 24 degrees C. The KD values were not significantly influenced by sodium ions. The dissimilarities between the rat and frog brain opioid receptors in [3H]ethylketocyclazocine binding are attributed to the different lipid composition of the two membranes.