Cholinergic neurons expressing substance P receptor (SPR, NK(1)) were examined in the rat brain using double immunofluorescence. The distribution of SPR-like immunoreactive (SPR-LI) neurons completely overlapped with that of choline acetyltransferase (ChAT)-LI neurons in the medial septal nucleus, the nucleus of diagonal band of Broca, the magnocellular preoptic nucleus, the substantia innominata of basal forebrain, the caudate-putamen, and the ventral pallidum of the basal ganglia. In the mesopontine tegmentum and the cranial motor nuclei of the brainstem, the distribution of SPR-LI and ChAT-LI neurons was partially overlapping. Neurons showing both SPR-like and ChAT-like immunoreactivities, however, were predominantly found above basal forebrain regions and 82-90% of these ChAT-LI neurons displayed SPR-like immunoreactivity, in addition to the confirmatory observation that 100% of the ChAT-LI neurons exhibit SPR-like immunoreactivity in the basal ganglia. In contrast, neurons double-labeled for SPR-like and ChAT-like immunoreactivities were hardly detected in aforementioned regions of the brainstem. The present study has provided morphological evidence for direct physiological modulation of cholinergic neurons by tachykinins through substance P receptor in the basal forebrain of the rat.