Glutamyl aminopeptidase (EAP, EC 3.4.11.7) selectively hydrolyzes N-terminal glutamyl and aspartyl residues from oligopeptides and is present in the brain. (Asp1)Angiotensin II (Ang II) is a substrate for EAP, and increasing evidence suggests that des(Asp1)angiotensin II (Ang III) is an active angiotensin peptide in the brain. To determine whether a relationship exists between EAP and Ang II/III in rat brain, we compared their immunocytochemical distributions. EAP-like immunoreactivity was localized primarily to the adventitial surface of cerebral microvessels throughout the forebrain. Endothelial cells, neurons and glial cells were not labeled. The immunocytochemical staining of microvessel adventitium with EAP antiserum was suggestive of labeling of perivascular pericytes since intravenous horseradish peroxidase resulted in a similar adventitial pattern of staining, in addition to pericyte cell bodies. EAP immunoreactivity was highest within circumventricular organs, areas known to contain high levels of Ang II receptors. Positively stained EAP microvessels were also concentrated in areas containing Ang II/III immunoreactive neurons or nerve terminals, including the hypothalamic paraventricular nucleus and the median eminence. The immunocytochemical localization of EAP suggests that it may be involved in a wide variety of functions within the brain, including: (i) metabolism of circulating peptides in brain areas devoid of a blood-brain barrier, (ii) metabolism of circulating peptides as a component of the blood-brain barrier, (iii) metabolism of intravascularly synthesized peptides, (iv) metabolism of hypothalamic peptides released into the portal circulation, (v) metabolism/conversion of neuronally released Ang II to Ang III in the interstitial space, and (vi) metabolism of neuronally released neuropeptides with vasoactive properties.(ABSTRACT TRUNCATED AT 250 WORDS)