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The regional distribution of mRNA coding for the m5 muscarinic acetylcholine receptor subtype was investigated in tissue sections of rat brain by in situ hybridization histochemistry. The highest hybridization signal was observed in the hippocampus, but restricted to the ventral subiculum, pyramidal cells of the CA1 and, with lower intensity, of the CA2 subfields. Significant levels of hybridization were also seen in the substantia nigra pars compacta, ventral tegmental area, lateral habenula, ventromedial hypothalamic nucleus and mammillary bodies. An involvement of the m5 muscarinic receptors in the regulation of the dopaminergic nigrostriatal pathway is suggested.