Characterization of opioid receptor types modulating acetylcholine releasein septal regions of the rat brain

Presynaptic opioid receptors of the delta- and mu-types have been shown to inhibit the release of acetylcholine (ACh) in the rat striatum and hippocam pus, respectively, but it is unknown whether opioid receptors modulate the release of ACh also in the region of origin of the hippocampal cholinergic innervation, the septum. To answer this question, slices (350 mu m) of the medial septal area and of the diagonal band of Broca, as well as (for compa rison) of the hippocampus, were prepared from adult male Wistar rats. The s lices were incubated with [H-3]choline, superfused in the presence of hemic holinium-3 (10 mu M) and stimulated twice (S-1, S-2) by electrical fields ( 360 pulses, 3 Hz, 2 ms, 60 mA); opioid receptor agonists were present durin g S-2. The preferential mu-agonist [D-Ala(2),N-Me-Phe(4),Gly-ol(5)]enkephal in (DAMGO) inhibited the evoked ACh release by maximally about 40% in hippo campal slices and acted even more strongly in the medial septal area, or th e diagonal band of Broca (about 60% or 75% maximal inhibition, respectively ). These effects: were reduced or abolished by the preferential mu-antagoni st naloxone, which showed no effects when given alone. Using naloxone in th e presence of a cocktail of peptidase inhibitors, no evidence for an endoge nous tone of opioid peptides was found in the medial septal area, diagonal band of Broca or the hippocampus. Using the preferential delta-agonist [D-P en(2), D-Pen(5)]enkephalin (DPDPE) and the delta-antagonist naltrindole, a delta-opioid receptor inhibiting evoked ACh release was clearly detectable both in the medial septal area and the diagonal band of Broca, but not in t he hippocampus, whereas the preferential kappa-agonist trans-3.4-dichloro-N -methyl-N-[2-(1-pyrrolidinyl)cyclo-hexyl] benzeneacetamide (U50,488H) had o nly weal; or no effects, In addition to the functional experiments, double in-situ hybridization studies were performed, in which cells containing mRN A for choline acetyltransferase (ChAT) were labeled by an antibody linked e nzymatic staining procedure, whereas mRNAs for mu- or delta-opioid receptor s were detected with radioactive probes. These experiments revealed that in the septal region mainly mu-opioid receptors were expressed by neurons pos itive for ChAT mRNA, whereas in the rat striatum the expression of delta-op ioid receptors prevailed in those neurons. We conclude that in the septal a rea of the rat brain, in contrast to the rat striatum and hippocampus, both presynaptic mu- and delta-opioid receptors,rs modulate the evoked release of ACh, Whether presynaptic mu- and delta-opiold receptors occur on the sam e or on different septal cells or axon terminals remains to be clarified.