Neuropeptide Y (NPY) was tested for an ability to directly influence the release of insulin using an in vitro isolated rat pancreatic islet system. NPY, at doses ranging from 100 pg/ml to 1 microgram/ml, had no significant effect on the basal release (5.5 mM glucose) of insulin. However, NPY treatment resulted in a significant, dose-dependent (1 ng/ml to 1 microgram/ml) inhibition of glucose-stimulated (11 mM) insulin release. When tested in a perfused rat pancreas preparation in situ, NPY administration led to a marked inhibition of both basal and stimulated insulin release followed by a postinhibitory rebound which exceeded the control insulin levels by 3-fold. In contrast, the intracerebroventricular (ICV) microinjection of NPY (5 micrograms) produced a significant but delayed (30 min) elevation of circulating insulin. It is therefore suggested that the direct action of NPY on insulin release is inhibitory while the central action of NPY indirectly results in an increase in plasma insulin. Thus, NPY may be added to the growing list of peptidergic agents which may affect the endocrine pancreas by acting as neurotransmitters and/or neuromodulators.