Characterization and autoradiographic distribution of hemicholinium-3 high-affinity choline uptake sites in mammalian brain

3H]hemicholinium-3 (HC-3) binding characteristics have been investigated using membrane binding assays and in vitro receptor autoradiography. In rat brain membrane preparations, 3H]HC-3 binds with high affinity to an apparent single class of sites. 3H]HC-3 binding is Na+-dependent. The ligand selectivity pattern strongly suggests that 3H]HC-3 selectivity labels the high affinity choline uptake (HACU) in brain membranes (HC-3 greater than choline greater than carbamylcholine greater than acetylcholine). This hypothesis is also supported by quantitative autoradiographic data which demonstrate that the discrete distribution of 3H]HC-3 binding sites correlates very well with the known distribution of other cholinergic markers such as choline acetyltransferase (ChAT), acetylcholinesterase (AChE), HACU, and 3H]AH-5183 (blocker of the vesicular transport of acetylcholine). For example, high densities of labelling are observed for these different markers in the interpeduncular nucleus, anteroventral nucleus of the thalamus, striatum, basolateral nucleus of the amygdala, and an exquisite laminar distribution in the hippocampus. Similar autoradiographic distributions of 3H]HC-3 binding sites are observed in other mammalian species such as guinea pig and monkey. Finally, 7-day unilateral kainic acid lesions of the nucleus basalis magnocellularis (nbm) decrease cortical 3H]HC-3 binding and ChAT activity, although not to a similar extent. In summary, these results demonstrate that 3H]HC-3 is a selective ligand of the HACU in mammalian brain. Thus, it is now possible to characterize precisely various structural components of the cholinergic synapses using markers such as 3H]HC-3, ChAT, HACU, 3H]AH-5183, and selective muscarinic and nicotinic receptor radioligands.