Little is known about the role that neuropeptides such as substance P play in cell-to-cell interactions in the striatum. The effect of locally perfused substance P on extracellular acetylcholine (ACh) in the dorsal striatum of awake, freely moving rats was examined using microdialysis. Neostigmine (1 microM) was included in the perfusate to improve recovery of ACh. Basal extracellular ACh was sensitive to Na(+)-channel blockade with tetrodotoxin (0.3 microM) and Ca(2+)-channel blockade with MgCl2 (10 mM) and therefore largely neuronal in origin. Local perfusion with 10 and 25 microM substance P for 20 min elevated extracellular ACh by 30% and 51%, respectively. The NK1 receptor antagonist, CP 96,345 (10 microM), which by itself had no effect on extracellular ACh, prevented the substance P-induced increase in extracellular ACh. These results suggest that stimulation of NK1 receptors by substance P enhances ACh release in the dorsal striatum and is consistent with anatomical evidence of a substance P-cholinergic circuit in this region.