The recent development of in vitro autoradiography techniques has enabled investigators to determine the distribution and relative levels of multiple ligand binding sites in discrete anatomical areas. In this study we used semi-quantitative in vitro autoradiography to compare the levels of binding to central mu1, mu2, and delta opioid sites in two strains of mice, C57BL/6BY and CXBK. The CXBK strain is known to be deficient in whole brain opioid binding sites and to be less sensitive than the C57 strain to the analgesic and locomotor stimulatory effects of opiates and opioids. Delta sites were visualized using [3H](D-Ala2-D-Leu5]-enkephalin (DADL) plus a low concentration of morphine, total mu sites (mu1 and mu2) were visualized using [3H] dihydromorphine (DHM), and mu2 sites were visualized using [3H]DHM plus a low concentration of DADL. Binding to mu1 sites was determined by subtracting mu2 binding from total mu binding. We found that the two strains did not consistently differ in the levels of delta site; in some areas the CXBKs had lower levels but in many areas they had levels equal to or greater than those for the C57s. The CXBK strain, however, either had less or the same amount of mu binding as the C57 strain in all areas studied. The CXBK strain was especially deficient in mu1 binding, particularly in areas involved in pain processing.