The cerebellar vermis, long associated with axial motor control, has been implicated in a surprising range of neuropsychiatric disorders and cognitive and affective functions. Remarkably little is known, however, about the specific cell types and neural circuits responsible for these diverse functions. Here, using single-cell gene expression profiling and anatomical circuit analyses of vermis output neurons in the mouse fastigial (medial cerebellar) nucleus, we identify five major classes of glutamatergic projection neurons distinguished by gene expression, morphology, distribution, and input-output connectivity. Each fastigial cell type is connected with a specific set of Purkinje cells and inferior olive neurons and in turn innervates a distinct collection of downstream targets. Transsynaptic tracing indicates extensive disynaptic links with cognitive, affective, and motor forebrain circuits. These results indicate that diverse cerebellar vermis functions could be mediated by modular synaptic connections of distinct fastigial cell types with posturomotor, oromotor, positional-autonomic, orienting, and vigilance circuits.

Although the overall topography of the cerebellar corticonuclear projection formed by Purkinje cell (PC) axons has been described, only a few studies have dealt with the organization of this projection at the level of individual PC axons. Thus, we reconstructed 65 single PC axons that were labeled with biotinylated dextran amine in the rat. We then analyzed the relationship between the projections of these PCs and the compartmentalization of the cerebellar cortex and nuclei based on the topography of olivocerebellar projection and aldolase C expression in PCs. After giving rise to short local recurrent collaterals near the soma, a PC axon formed a terminal arbor in a specific small area in the cerebellar nuclei (CN). The terminal arbors of vermal PCs were spread more widely than those of hemispheric PCs and sometimes extended to extracerebellar targets. PCs located in any of the aldolase C-positive (Groups I and II) and -negative (Groups III and IV) stripes consistently projected to the caudoventral and rostrodorsal parts of the CN, respectively, precisely in accordance with the compartmentalization of the cortex and nuclei. Mediolateral segregation and rostrocaudal convergence were seen between projections of separate PCs in a single aldolase C compartment. The results revealed a tight link between the projection patterns of individual PC axons, the topography of the olivocerebellar pathway, and the aldolase C expression pattern. Their overall correspondence seems to reflect a basic aspect of cerebellar organization, although some area-dependent variation in the relationship of these three entities was also present.

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Aldolase C (Aldoc, also known as “zebrin II”), a brain type isozyme of a glycolysis enzyme, is expressed heterogeneously in subpopulations of cerebellar Purkinje cells (PCs) that are arranged longitudinally in a complex striped pattern in the cerebellar cortex, a pattern which is closely related to the topography of input and output axonal projections. Here, we generated knock-in Aldoc-Venus mice in which Aldoc expression is visualized by expression of a fluorescent protein, Venus. Since there was no obvious phenotypes in general brain morphology and in the striped pattern of the cerebellum in mutants, we made detailed observation of Aldoc expression pattern in the nervous system by using Venus expression in Aldoc-Venus heterozygotes. High levels of Venus expression were observed in cerebellar PCs, cartwheel cells in the dorsal cochlear nucleus, sensory epithelium of the inner ear and in all major types of retinal cells, while moderate levels of Venus expression were observed in astrocytes and satellite cells in the dorsal root ganglion. The striped arrangement of PCs that express Venus to different degrees was carefully traced with serial section alignment analysis and mapped on the unfolded scheme of the entire cerebellar cortex to re-identify all individual Aldoc stripes. A longitudinally striped boundary of Aldoc expression was first identified in the mouse flocculus, and was correlated with the climbing fiber projection pattern and expression of another compartmental marker molecule, heat shock protein 25 (HSP25). As in the rat, the cerebellar nuclei were divided into the rostrodorsal negative and the caudoventral positive portions by distinct projections of Aldoc-positive and negative PC axons in the mouse. Identification of the cerebellar Aldoc stripes in this study, as indicated in sample coronal and horizontal sections as well as in sample surface photos of whole-mount preparations, can be referred to in future experiments.