Although the whole cerebellar cortex receives cholinergic afferents, the source of origin of this projection has been clarified only for some corticocerebellar regions. Experiments were performed in kittens to investigate whether the two major cholinergic groups of the brainstem, the pedunculopontine (PPT) and laterodorsal tegmental nuclei (LDT), contribute to the cholinergic innervation of the cerebellar cortex, in particular the vermal cortex. Tegmento-cerebellar projecting neurons were identified by injecting the retrograde tracer rhodamine-labeled latex microspheres in the lobules V to VII of the cerebellar vermis. Subsequently, some of these tegmento-cerebellar neurons were demonstrated to be cholinergic by using the immunohistochemical technique for choline acetyltransferase (ChAT). Only a small portion of the ChAT-positive tegmental neurons projected to the cerebellar vermis. However, among the whole population of the retrogradely labeled tegmental neurons about one third were cholinergic. These cholinergic tegmento-cerebellar neurons were located in the PPT, LDT, and also within the locus coeruleus (LC) complex, where noradrenergic neurons predominate. Since the LC complex sends noradrenergic afferents to the cerebellar cortex, it appears that the dorsal pontine area contributes to the tegmento-cerebellar projections not only with noradrenergic but also with cholinergic afferents. The physiological significance of this cholinergic projection to the cerebellar cortex has been discussed.