Cerebellar choline acetyltransferase positive mossy fibres and their granule and unipolar brush cell targets: A model for central cholinergic nicotinic neurotransmission

A subset of cerebellar messy fibres is rich in choline acetyltransferase, the rate-limiting enzyme for the synthesis of acetylcholine. These choline acetyltransferase-positive messy fibres are concentrated in the vestibule cerebellum and originate predominantly from the medial vestibular nucleus. The granular layer of the vestibulocerebellum is also enriched in unipolar brush cells, an unusual type of small neuron that form giant synapses with messy fibres. In this immunocytochemical light and electron microscopic study, we explored whether choline acetyltransferase-positive messy fibres innervate unipolar brush cells of the rat cerebellum. We utilized monoclonal antibodies to rat choline acetyltransferase of proven specificity, and immunoperoxidase procedures with 3,3'-diaminobenzidine tetrahydrochloride as the chromogen. A high density of choline acetyltransferase-positive fibres occurred in the nodulus and ventral uvula, where they showed an uneven, zonal distribution. Immunostained messy fibre rosettes contained high densities of round synaptic vesicles and mitochondria. They formed asymmetric synaptic junctions with dendritic profiles of both granule cells and unipolar brush cells. The synaptic contacts between choline acetyltransferase-immunoreactive messy fibres and unipolar brush cells were very extensive, and did not differ from synapses of choline acetyltransferase-negative messy fibres with unipolar brush cells. Analysis of a total area of 1.25 mm(2) of the nodulus from three rats revealed that 14.2% of choline acetyltransferase-immunoreactive messy fibre rosettes formed synapses with unipolar brush cells profiles. Choline acetyltransferase-positive rosettes accounted for 21.7% of the rosettes forming synapses with unipolar brush cells. Thus, the present data demonstrate that unipolar brush cells are innervated by a heterogeneous population of messy fibres, and that some unipolar brush cells receive cholinergic synaptic input from the medial vestibular nucleus. The ultrastructure of these synapses is compatible with the possibility that choline acetyltransferase-positive messy fibres co-release acetylcholine and glutamate. As the granular layer of the vestibulocerebellum contains nicotinic binding sites, the choline acetyltransferase-positive messy fibres may be a model for studying nicotinic neurotransmission in the CNS.