The localization of 5′-nucleotidase, an adenosine-producing ectoenzyme, was studied by a cytochemical method in the superior cervical ganglion of the adult cat and guinea pig. The following subcellular sites of enzymatic activity were detected: (1) the surface of Schwann and satellite cells including the extracellular space between these cells and neuronal profiles; (2) the plasmalemma and pinocytotic vesicles of capillary endothelial cells; and (3) the synaptic clefts between cholinergic preganglionic axon terminals and sympathetic neurons. The simultaneous presence of 5′-nucleotidase at both glial elements and synapses within the adult peripheral nervous system (PNS) constitutes a novel distribution pattern for this enzyme which does not apply to the mature central nervous system (CNS), but which is rather typical for the developing CNS. These distributions of 5′-nucleotidase activity may reflect specific cellular requirements for nucleosides involved in parenchymal metabolism, in vascular transport processes and, possibly, in synaptic plasticity.