There is general agreement that the medial preoptic nucleus (MPN) receives projections from widespread regions of the brain, although there are significant discrepancies in the literature with regard to certain specific inputs. Therefore, we have reexamined the inputs to this nucleus with both retrograde and anterograde axonal transport techniques. First, injections of the retrograde tracers true blue, SITS, or wheat germ agglutinin were made into the region of the MPN and the distribution of retrogradely labeled cells was charted. Then, autoradiographic material was used to confirm the results of the retrograde studies, to identify the route taken by fibers projecting to the MPN, and to describe the distribution of projections with respect to the three cytoarchitectonic subdivisions of the nucleus. The results indicate that the MPN receives inputs from widely distributed areas in both the forebrain and brainstem, and that these inputs appear to be distributed topographically within the three cytoarchitectonic subdivisions of the nucleus. Direct inputs to the MPN arise from all major areas of the hypothalamus (except for the median and magnocellular preoptic nuclei, the supraoptic and suprachiasmatic nuclei, and the medial and lateral mammillary nuclei). Projections from nuclei within the periventricular zone of the hypothalamus end primarily in the medial part of the MPN, while inputs from the lateral zone are mainly confined to the lateral part of the nucleus, as are projections from the nuclei within the medial zone, except for those from the anterior and ventromedial nuclei, which appear to be more widespread. The MPN receives major inputs from limbic regions including the amygdala, ventral subiculum, and ventral lateral septal nucleus, all of which end preferentially in the lateral part of the MPN. In contrast, the projection from the encapsulated part of the bed nucleus of the stria terminalis appears to end preferentially in the central part of the MPN and in immediately adjacent regions of the medial subdivision. In addition, the MPN may receive relatively sparse inputs from infralimbic and insular cortical areas, the nucleus accumbens, and the substantia innominata. Finally, ascending serotoninergic projections from the raphe nuclei appear to terminate principally in the lateral part of the MPN, whereas inputs from regions containing noradrenergic cell groups are chiefly distributed to the central and medial parts of the nucleus. Other brainstem regions that appear to provide modest inputs include the ventral tegmental area, central tegmental field, periaqueductal gray, pedunculopontine nucleus, and the peripeduncular nucleus.(ABSTRACT TRUNCATED AT 400 WORDS)