Histochemical techniques have been used to examine the development of the enzyme acetylcholinesterase (AChE) in normal and transplanted rat superior colliculus (SC). At birth (P0), relatively little AChE activity was found in SC in situ; however, there was a gradual increase in the intensity of AChE staining in the SC over the first 4 postnatal weeks. In the superficial layers, an increase in AChE activity was first seen in rostromedial SC at P6 and was found throughout the upper tectal layers by P10. An increase in AChE activity in the intermediate layers was apparent by P12 and the adult pattern, characterized by periodic bands of AChE staining, was established by P22. In tectal grafts, the development of AChE activity followed a time course similar to that found in normal SC. Mature tectal grafts contained moderate AChE activity with AChE‐positive cells scattered throughout the neuropil. There were however, localized, often spherically shaped areas which displayed relatively intense AChE activity. These AChE‐dense areas had a characteristic appearance in adjacent sections stained for Nissl or neurofibrils. Significantly, host retinal input, where present, was always restricted to the AChE‐dense regions and it seems certain that these areas are homologous to the superficial layers of normal SC. AChE‐rich regions were also present, however, in grafts which received no retinal input and in general the pattern of AChE activity in tectal grafts was strikingly similar, irrespective of their location or connections with the host brain. It would appear, therefore, that much of the AChE activity in tectal transplants, and presumably in SC in situ, is intrinsic to that region and not derived from or dependent upon extrinsic innervation.