Two endogenous brain peptides (Tyr-W-MIF-1 (Tyr-Pro-Trp-Gly-NH2) and Tyr-MIF-1 (Tyr-Pro-Leu-Gly-NH2)), a cyclized analog and two fragments of Tyr-W-MIF-1, and hemorphin (Tyr-Pro-Trp-Thr) were tested for binding to mu 1 and mu 2 opiate receptor. All these peptides bound to both mu 1 and mu 2 sites in assays optimized to discriminate these subtypes of the mu opiate receptor in membranes from bovine thalamus. The cyclized analog of Tyr-W-MIF-1, previously shown to have potency near that of Tyr-D-Ala-Gly-N-MePhe-Gly-ol (DAMGO) and morphine in producing analgesia after intracerebroventricular (i.c.v.) injection, bound to mu 1 and mu 2 sites with affinities similar to those of DAMGO. Tyr-W-MIF-1, previously shown to induce analgesia after i.c.v. injection but with much higher potency after intrathecal (i.t.) injection, also bound to both mu 1 and mu 2 sites with an affinity between that of morphiceptin and hemorphin. Although the highest ratios of Ki's for mu 2/mu 1 were shown by hemorphin, Tyr-W-MIF-1, and Tyr-W-MIF-1, none of the compounds were significantly different in selectivity. The results indicate that the relatively lower potency of Tyr-W-MIF-1 after i.c.v., compared with i.t. injection, is not due to a lack of binding to mu 1 sites. They suggest that it has relatively high efficacy at mu 2, but low efficacy at mu 1 sites, a possibility that might explain some of the novel properties of these peptides.