Choline acetyltransferase immunoreactivity is localized to four types of synapses in the rat interpeduncular nucleus

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Abstract
The cholinergic synapses of the rat interpeduncular nucleus (IPN) were demonstrated by immunostaining that utilized a monoclonal antibody directed against choline acetyltransferase. The rostral, central, intermediate and lateral subnuclei of the IPN each contained a single type of immunoreactive terminal. Immunoreactivity was localized to synaptic vesicle membranes, inner and outer mitochondrial membranes, the cytoplasmic surface of plasma membranes (especially at the contact zones), and longitudinal microtubules in preterminal portions of axons. Terminals were identified by comparison to previous studies of the synaptic organization of the IPN. In the rostral subnucleus, the immunoreactive terminals were characterized by their content of spherical vesicles, 45 nm in diameter, intermixed with moderate numbers of dense-cored vesicles, 75–100 nm in diameter. These terminals formed asymmetrical contacts. They correspond to the more numerous of the two types of axodendritic terminals described in this subnucleus, i.e. those which degenerate after lesions of the habenula. The moderate number of immunoreactive terminals in the lateral subnucleus contained pleomorphic vesicles, 30–45 nm in diameter. Up to three of these formed symmetrical contacts with individual dendrites, which ranged in diameter from 0.35 to 0.55μm. The other types of axodendritic terminal in this subnucleus, which often contacted the same dendrites, were unstained. These latter terminals have been interpreted as being those which contain substance P. The immunoreactive terminals in the central subnucleus consisted of moderate numbers of S terminals. These contained spherical vesicles, 40–60 nm in diameter, and formed markedly asymmetricalen passant contacts with small dendritic processes or spines. The immunoreactive terminals in the intermediate subnucleus had the same presynaptic and contact morphology. Many were clearly crest synapses. The remainder appeared to be such, but seen only partially within the plane of section. In the intermediate subnucleus there were up to several hundred immunostained terminals per grid square in some sections. These findings are consistent with the existence of a dense cholinergic projection to the IPN. The crest and S synapses, both of which degenerate after lesions of the habenular region, are shown to be cholinergic as previously suggested. Additionally, demonstration of the cholinergic innervation of other subnuclei of the IPN increases understanding of the relation of cholinergic to other transmitters localized to various portions of this nucleus.