Neurotensin-like immunoreactivity (NT-LI) was demonstrated in projection neurons of the striatum and nucleus accumbens in the cat by combining immunohistochemistry and the fluorescent retrograde neuronal labeling method. In colchicine-treated cats, many neurons with NT-LI were found in the caudate nucleus, nucleus accumbens, and putamen. Most of these neurons were medium-sized neurons with spiny dendrites. NT-LI of neuronal elements in the caudate nucleus and nucleus accumbens formed dense aggregates with irregular figures, which appeared to correspond to the striosomes of Graybiel et al. (Proc. Natl. Acad. Sci. USA 75:5723-5726, '78; Exp. Brain Res. 34:189-195, '79; Neuroscience 6:377-397, '81). Fibers with NT-LI were distributed massively to the globus pallidus and ventral midbrain regions, but not to the entopeduncular nucleus. In the ventral midbrain regions, many fine varicose fibers with NT-LI were distributed to the pars compacta and pars lateralis of the substantia nigra, ventral tegmental area, and retrorubral area. In the pars reticulata of the substantia nigra, however, fibers with NT-LI were rather sparse. Examination of consecutive sections immunostained for NT, enkephalin (Enk), GABA, and substance P (SP) revealed that 50% of neurons with NT-LI in the caudate nucleus and nucleus accumbens exhibited Enk-LI, 15% showed GABA-LI, and 5% manifested both Enk-LI and GABA-LI; no NT-positive neurons in the striatum and nucleus accumbens showed SP-LI. No morphological differences were found between NT-positive neurons with Enk-LI and/or GABA-LI and those without Enk-LI and GABA-LI. Most neurons with NT-LI in the striatum and nucleus accumbens were retrogradely labeled with True Blue injected into the globus pallidus, pars compacta and pars lateralis of the substantia nigra, and ventral tegmental area. After hemitransection severing neuronal connections between the ventral midbrain regions and the forebrain structures, fibers with NT-LI and those with Enk-LI in the ventral midbrain regions were markedly reduced in number.(ABSTRACT TRUNCATED AT 400 WORDS)