The cellular localization of uterine oxytocin binding sites in the rat was studied by means of in vitro receptor autoradiography. Using [tyrosyl-3,5-3H]oxytocin as ligand, binding sites were localized in tissue sections from uteri of estrous, mated, and artificially cervically stimulated rats (n = 4 per group), and specificity of binding was investigated by means of simultaneous incubations with oxytocin, [Gly4,Thr7]oxytocin and [Arg]vasopressin. A previously unidentified type of cell was densely labelled by tritiated oxytocin. The labelled cells were preferentially localized near the endomyometrial border and at the interface of the circular and longitudinal muscle layers. In addition, these cells were found in the muscle layers. The dense labelling of these cells, which did not constitute part of the endometrial epithelium or blood vessels, was abolished when oxytocin or [Arg]vasopressin, but not [Gly4,Thr7]oxytocin, was added to the incubation medium. Binding of the radioligand was also found on muscle cells of the circular and longitudinal layers of the myometrium and cells of the endometrial luminal and glandular epithelium. Whereas incubation with oxytocin and [Gly4,Thr7]oxytocin diminished the labelling in both myometrium and endometrium, incubation with [Arg]vasopressin reduced labelling only in the myometrium. Similar results were obtained in tissues from rats in different reproductive states. This study demonstrates the presence of oxytocin binding sites in three different types of cell in the uterus of the rat. While the sites in the myometrium may be associated with the contractile response of this type of tissue to oxytocin, the functional significance of oxytocin binding sites on the endometrial epithelium and in the densely labelled, scattered cells remains to be elucidated.