The content of acetylcholine (ACh) in nerve terminals or the dorsal hippocampus was examined after intraventricular, intraseptal, or intrahippocampal administration of a variety of endorphin/corticotropin neuropeptides. beta-Lipotropin, alpha-endorphin, gamma-endorphin, alpha-melanotropin, beta-melanotropin, adrenocorticotropin-1-39 (ACTH1-39), and ACTH4-10 (1, 3, 10, or 30 micrograms each) did not affect levels of ACh in the hippocampus 30 min after injection into the lateral ventricle. beta-Endorphin, administered intraventricularly (1, 3, 10, or 30 micrograms) or intraseptally (1 microgram), increased levels of ACh, while ACTH1-24, injected similarly, decreased levels of the neurotransmitter. ACh concentrations remained unchanged after direct application of beta-endorphin or ACTH1-24 (1, 3, 10, of 30 micrograms each) into Ammon's horn. Acute unilateral transection of the fimbria/superior fornix resulted in a time-related decrease in hippocampal ACh concentrations. Levels of ACh did not change 1 hr after transection; however, concentrations of hippocampal ACh decreased significantly 1 d or 1 week after deafferentation. ACh levels in the contralateral hippocampus remained unaffected at all times tested. Fimbrial transection blocked fully both endorphin- and corticotropin-induced changes in hippocampal ACh after the neuropeptides were injected into the lateral ventricle or the septal region. Naloxone, which, after subcutaneous (1 mg/kg) or intraventricular (100 micrograms) injection alone, failed to change levels of hippocampal ACh, antagonized the effects of intraventricular or intraseptal beta-endorphin or ACTH1-24 or hippocampal ACh levels. The results suggest a site of endorphin/corticotropin receptor interaction at the level of cholinergic cell bodies in the septal region for regulating the activity of septohippocampal cholinergic neurons.