The topographical distribution of choline acetyltransferase, muscarinic receptor binding and high affinity choline uptake was studied in 21 separate areas of the rat striatum. The areas of the nucleus chosen represented dorsolateral, dorsomedial, ventrolateral and ventromedial regions along the rostrocaudal aspect of the striatum, such that a three-dimensional distribution of the cholinergic parameters could be obtained. Within any given rostrocaudal section, no significant dorsoventral differences were noted for any of the cholinergic parameters. On the other hand, marked differences were found in a comparison of the medial and lateral striatum. Choline acetyltransferase, muscarinic receptor binding and high affinity choline uptake were more concentrated in the lateral striatum than the medial striatum, and the magnitude of this medio-lateral disparity increased from rostral to caudal regions of the nucleus. The lateral striatum exhibited no significant rostrocaudal variations in the cholinergic parameters; however, the medial portion of the striatum did exhibit differences along its rostrocaudal extent, with the rostral-most sections being enriched relative to the more caudal sections. These results suggest that the cholinergic system in the striatum is heterogeneously distributed within this nucleus, with the lateral portion possessing a greater cholinergic innervation than the medial portion. They further suggest that future neurochemical studies of cholinergic alterations in the striatum should include a consideration of the possibility of regional effects within the nucleus rather than treating the striatum as a homogeneous tissue.