The distribution, possible origins and fine structure of neuropeptide Y (NPY)-containing nerve fibers in the rat liver were investigated by immunohistochemistry, nerve transection and immunoelectron microscopy. Light-microscopic immunohistochemistry showed NPY fibers forming a complex network in and around the walls (tunica adventitia and tunica media) of hepatic vessels. They were also closely associated with interlobular bile ducts. The NPY fibers in the liver were almost completely eliminated by transection of the greater splanchnic nerves just distal to the celiac ganglion. Transection of the greater splanchnic nerves just proximal to the celiac ganglion resulted in a marked decrease in NPY fibers, but a significant number remained intact. Under electron microscopy. NPY terminals without a covering of glial processes were seen not only in proximity to smooth muscle cells within the tunica media of hepatic vessels but also in the subendothelial areas of the tunica intima. Some NPY axon terminals devoid of glial ensheathment were located close to the basal lamina of interlobular bile ducts. Occasionally, single axon terminals with NPY were found in the vicinity of or in contact with hepatic cells. There was a small number of NPY fibers that had lost their glial sheaths while running toward lymphatic capillaries. These findings suggest that hepatic NPY arises from the celiac ganglion and paravertebral sympathetic ganglia, and that it is involved in more complex physiological processes than the previously described neuropeptides in the liver, which are localized exclusively to hepatic vessel walls.