The organization of three of the efferent projections of substantia nigra has been investigated in rat by means of combined injections of two fluorescent retrograde tracers: red fluorescent Evans Blue and blue fluorescent DAPI-primuline mixture. First the distributions of the retrogradely labeled neurons in substantia nigra after injections of each of the two tracers in the striatum, thalamus and superior colliculus were compared with the distributions of the retrogradely labeled nigral neurons after HRP injections in these same structures. The findings in these sets of experiments were the same. This indicates that the two fluorescent retrograde tracers are as effective as HRP in tracing the efferent fiber connections of the substantia nigra. Subsequently the retrograde labeling of the substantia nigra neurons was studied after combined injections of the two fluorescent retrograde tracers in two of the above structures, in different combinations. In these experiments both single labeled neurons fluorescing either red or blue, as well as double labeled neurons fluorescing both red and blue were found. Double labeled neurons were present only in pars reticulata and occurred only after injections of the two tracers in thalamus and superior colliculus respectively. From this it has been concluded that in rat the various efferent projections from the substantia nigra are mainly derived from different neurons, but that at least part of the neurons which distribute fibers to superior colliculus also distributes collaterals to the thalamus.