We compared the development of enkephalin (Enk) expression in normal rats and rats in which target contact was transiently disrupted with 6- hydroxydopamine (6-OHDA). During the first 3 postnatal weeks, there was a striking increase in Enk immunoreactivity (-IR) in superior cervical ganglia (SCG) assayed by radioimmunoassay (RIA). This increase was correlated with the appearance of Enk-IR in postganglionic neurons. In the caudal region of the SCG, the proportion of Enk-IR neurons and their immunoreactivity increased until one-third of the neurons possessed Enk-IR between postnatal days (P) 14 and 21. After P21, the number of Enk-IR neurons and their immunofluorescence decreased. By 6 weeks, only occasional neurons possessed moderate Enk-IR. The increases in Enk-IR were correlated with increased ganglionic proenkephalin A mRNA detected by in situ hybridization. The decrease in IR after P21 was not, however, paralleled by a comparable decrease in proenkephalin A mRNA. To determine whether interactions between SCG neurons and their target tissues influence Enk expression, we disrupted them by treating neonatal rats with a single dose of 6-OHDA at P0. This treatment transiently reduced sympathetic fiber density in the submandibular gland, one target of Enk-IR neurons, over 90%. Two weeks later, the fiber density in glands of treated animals was not different from control. Following 6-OHDA, the concentration of Enk-IR in SCG extracts and the number of Enk-IR neurons and their immunofluorescence intensity failed to increase. SCG from treated rats also contained fewer neurons with proenkephalin A mRNA. In contrast, the content of neuropeptide Y (NPY) and the proportion of NPY-IR neurons were not decreased by 6-OHDA treatment. Our results indicate that the developmental history of Enk expression differs from that of other neuropeptides in rat sympathetic ganglia, suggesting that distinct mechanisms regulate the expression of individual neuropeptides. Further, they provide evidence that target contact during a critical period is important for the induction of Enk.