In mammals, photoperiodic information is conveyed from the retina to the pineal through a polysynaptic pathway, which includes the suprachiasmatic nucleus (SCN), the paraventricular nucleus of the hypothalamus (PVN), the spinal preganglionic neurons and, finally, the superior cervical ganglion. Precise data on the site in the PVN or which SCN transmitters are involved in the transmission of information in this pathway is lacking. In the present experiment we investigated whether SCN efferents containing vasoactive intestinal polypeptide (VIP) innervate PVN neurons that project to the spinal cord. A combination of retrograde tracing and immunocytochemistry with the aid of a confocal laser scanning microscope allowed us to assess possible interaction of SCN efferents and spinal cord projecting neurons in the PVN. Approximately 30% of identified autonomic projecting neurons in the dorsal PVN and 40% in the ventral PVN received VIP innervation mainly on their dendrites. These results provide further evidence for the involvement of SCN-derived VIP in the transmission of circadian information to the pineal.