Several types of opioid binding sites have been differentiated using biochemical and pharmacological criteria. We have used quantitative in vitro autoradiography to compare the levels of mu1 and mu2 opioid binding in the mouse central nervous system. Mu1 sites have a high affinity for all labeled opioids studied to date and have been associated with their analgesic effects, whereas mu2 sites have a high affinity only for opiate alkaloids and have been associated with their respiratory depressant effects. We used [3H]dihydromorphine (DHM) to visualize total mu sites (mu1 and mu2) and [3H]DHM plus a low concentration of [D-Ala2-D-Leu5]enkephalin (DADL) to visualize mu2 sites. Levels of mu1 binding were determined by subtracting mu2 binding from total mu binding. This mu1 distribution was confirmed in selected regions by an alternate method using [3H]DADL. High ratios of mu1 to mu2 binding were noted in frontal cortex, nucleus accumbens, rostral striatum, ventral pallidum, ventral periaqueductal gray matter, and laminae I and II of the spinal cord. The observation of high densities of mu1 binding in certain pain processing areas correlates with behavioral and pharmacological studies suggesting that analgesia from opiates and opioids is mediated primarily by mu1 sites. In other areas, such as the limbic system, dorsal nucleus of the vagus nerve, and nucleus of the solitary tract, either a low ratio of mu1 to mu2 binding or no mu1 binding was observed. This differential regional localization of mu1 and mu2 binding provides further evidence for the distinctness of these sites.