The origin of the vasopressin-containing fibers in the rat lateral septum was studied by means of lesioning specific areas, in which vasopressin-containing cells are found, or by surgically separating the septum from the underlying structures. Following these procedures sections of the brain were stained immunocytochemically for the presence of vasopressin. In addition, retrograde labeling tracers were injected in the lateral septum. Lesioning of the paraventricular nucleus did not result in the disappearance of vasopressin fibers from the lateral septum, nor from the various other areas studied. It did, however, cause the disappearance of fibers from the nucleus of the solitary tract and the nucleus ambiguus. By contrast, after the same lesion practically the whole oxytocinergic innervation of the brain disappeared. Injection of tracers into the lateral septum revealed retrograde labeled cells, e.g. in the bed nucleus of the stria terminalis, but not in the paraventricular and supraoptic nucleus. Horizontal cuts under the lateral septum, intersecting the diagonal band of Broca, resulted in a dramatic decrease of the vasopressin fibers in the lateral septum, suggesting that the fibers enter the septum via this structure. Moreover, since the vasopressin fiber density was found to decrease drastically in the lateral septum after lesioning the bed nucleus of the stria terminalis, the vasopressin cells found in this area are probably the source of these fibers. Other areas where fibers were seen to decrease after lesions of the bed nucleus are the diagonal band of Broca, the area of the anterior amygdala, the lateral habenular nucleus, the periventricular gray, and the locus coeruleus.