Cerebellum Lecture: the Cerebellar Nuclei-Core of the Cerebellum

Abstract

The cerebellum is a key player in many brain functions and a major topic of neuroscience research. However, the cerebellar nuclei (CN), the main output structures of the cerebellum, are often overlooked. This neglect is because research on the cerebellum typically focuses on the cortex and tends to treat the CN as relatively simple output nuclei conveying an inverted signal from the cerebellar cortex to the rest of the brain. In this review, by adopting a nucleocentric perspective we aim to rectify this impression. First, we describe CN anatomy and modularity and comprehensively integrate CN architecture with its highly organized but complex afferent and efferent connectivity. This is followed by a novel classification of the specific neuronal classes the CN comprise and speculate on the implications of CN structure and physiology for our understanding of adult cerebellar function. Based on this thorough review of the adult literature we provide a comprehensive overview of CN embryonic development and, by comparing cerebellar structures in various chordate clades, propose an interpretation of CN evolution. Despite their critical importance in cerebellar function, from a clinical perspective intriguingly few, if any, neurological disorders appear to primarily affect the CN. To highlight this curious anomaly, and encourage future nucleocentric interpretations, we build on our review to provide a brief overview of the various syndromes in which the CN are currently implicated. Finally, we summarize the specific perspectives that a nucleocentric view of the cerebellum brings, move major outstanding issues in CN biology to the limelight, and provide a roadmap to the key questions that need to be answered in order to create a comprehensive integrated model of CN structure, function, development, and evolution.

Keywords: Cerebellar ataxias; Cerebellar modules; Cerebellar nuclear afferents; Cerebellar nuclear anatomy; Cerebellar nuclear cell types; Cerebellar nuclear efferents; Cerebellum; Chick; Connectivity; Development; Evolution; Migration; Molecular specification; Mouse; Neurogenesis; Pathology; Rhombic lip; Ventricular zone.

Fig. 1

Panel a shows the conventional, cortico-centric, model by which the cerebellar circuitry is described. Afferent inputs, conveyed by CF (climbing fibers) and MF (mossy fibers) terminate predominantly in the cerebellar cortex, with collateral copies to the CN, which are thought to be of lesser importance. The cerebellar cortex processes the signal, which then passes from the Purkinje cells (PC) to the cerebellar nuclei (CN) and out of the cerebellum. Panel b illustrates the nucleo-centric perspective. The primary pathway is for cerebellar afferents to synapse in the CN, where the cerebellar efferents originate. In parallel, afferent copies are sent to the cerebellar cortex, where a complementary inhibitory signal is generated that enters the CN via the corticonuclear pathway and modulates the cerebellar efferent output. Please note that the thickness of the arrows reflects the relative importance of the information flow, not anatomical size or signal strength. Furthermore, blue arrows denote information passing through the CN circuits, without reference to specific cell types

Fig. 2

3D-reconstructions based on serial sections of the CN of the mouse, rat, cat, macaque, bonobo and human. Each horizontal panel depicts a rostral (anterior) view, a dorsal view, and a dorsal view with separated individual nuclei. Note that the relative size and shape of the various nuclei can vary considerably. The dentated appearance of the Lat can only be appreciated in the bonobo and human. In apes and humans, the gyration of the Lat can be divided into a caudoventral macrogyric (red) and rostrodorsal microgyric (purple) part. These dentated sheets of cells fold over the hilus that, in a rostral and medioventral direction, gives access to the scp. Scale bars indicate 1 mm (mouse, rat, cat and macaque), 2 mm (bonobo) and 10 mm (human). Reconstructions were made with Neurolucida (MBF Bioscience)

Fig. 3

Series of equidistant (80 µm) photomicrographs of transverse, thionine-stained Sects. (40 µm) of the mouse CN from its caudal-most level (panel 1) to its rostral-most level (panel 18). Midline is at the left-hand margin of each panel. The four main nuclei are indicated by thin lines. Dashed lines indicate equivocal nuclear borders. Arrows in panel 18 denote medial (M), dorsal (D), lateral (L), and ventral (V) directions. Scale bar in panel 1 equals 500 µm. Abbreviations: CoN, cochlear nuclei; dLat, dorsal part of the Lat; icp, inferior cerebellar peduncle; IntA, anterior cerebellar nucleus; IntDL, dorsolateral hump; IntDM, dorsomedial crest; IntIC, interstitial cell groups; IntP, posterior interposed nucleus; IV, inferior vestibular nucleus; Lat, lateral cerebellar nucleus; LC, locus coeruleus; LV, lateral vestibular nucleus; Med, medial cerebellar nucleus; MedDL, dorsolateral hump; MV, medial vestibular nucleus; MVm, magnocellular part of MV; MVp, parvocellular part of MV; scp, superior cerebellar peduncle; SV, superior vestibular nucleus; un, uncinate fascicle; vLat, ventral part of the Lat; Y, group Y

Fig. 4

Overview of the CN of the mouse and the fiber bundles connecting them to the rest of the brain. a The location of CN and fiber bundles in a sagittal schematic of the mouse brain. b A depiction of the CN within the cerebellum. Arrows indicate the primary directions of axonal projections within the bundles. Dark green and light blue connections via the superior cerebellar peduncle indicate ascending and descending connections. The two arrows feeding into the icp and mcp indicate a combination of inputs arriving from ascending pathways (e.g., from the spinal cord or inferior olive) and descending ones (e.g. via the basal pontine nuclei). Note that, for clarity, the brainstem is not shown. Abbreviations: CB, cerebellum; Med, medial nucleus; Int, interposed nucleus; Lat, lateral nucleus; scp, superior cerebellar peduncle; mcp, middle cerebellar peduncle; icp, inferior cerebellar peduncle; unc, uncinate fibers; IO, inferior olive

Fig. 5

Schematic depiction of the 5 classes of CN afferent inputs, indicated by numerals: 1, GABAergic axons of the PCs converging on CN neurons; 2, glutamatergic axons of the IO neurons; 3, glutamatergic non-IO-originating axons that also branch as mossy fibers in the cerebellar cortex; 4, glutamatergic non-IO-originating axons that do not contribute to the cerebellar cortical mossy fibers; and 5, modulatory afferents. Abbreviations: MF, mossy fibers; PC, Purkinje cells; GC, granule cell; PF, parallel fiber; CF, climbing fiber; mcp, middle cerebellar peduncle; icp, inferior cerebellar peduncle. The arrows indicate approximate image directions: D, dorsal; V, ventral; L, lateral; M, medial. The Med, Int, and Lat are colored as in Fig. 2. For details regarding the distribution of the afferent axons among the nuclei, refer to Table 2

Fig. 6

Illustration of modular connections in the rat cerebellum. a Iontophoretically applied injection of cholera toxin centered on the IntA, without involvement of surrounding nuclei. b1 A stripe-like band of retrogradely labeled PC’s in lobules IV and V of the anterior lobe. b2 Detail of cortical labeling showing retrogradely labeled PC somata aligned with CF terminals running like railroad tracks perpendicular to the surface in the molecular layer. c Retrogradely labeled olivary cells are only observed in the ventral fold of the dorsal accessory olive. Inset shows detail with labeled olivary neurons (asterisks) and dense labeling of fine terminal arborizations of nucleo-olivary afferents in the neuropil (between arrowheads). d 3D reconstruction (Neurolucida™) showing the white matter (blue) of the anterior part of the cerebellum (seen from the anterior) with the location of labeled PCs (yellow) and labeled CF (red). Note the near-perfect correspondence of both types of labeling indicating the modularity of the olivo-cortico-nuclear connections. Scale bar equals 250 μm in a, b1, c (10 μm in inset), 25 μm in b2. Abbreviations: III, IV, V, cerebellar lobules III, IV, V; C1, C3, PC stripes projecting to IntA; IntA, anterior interposed nucleus; IntDL, dorsolateral hump; IODvf, ventral fold of dorsal accessory olive; IOPdm, dorsomedial group of the principal olive; IOP, principal olive; Lat, lateral cerebellar nucleus; Med, medial cerebellar nucleus; rIOM, medial accessory olive, rostral part; scp, superior cerebellar peduncle; v4; fourth ventricle [Modified from 30]

Fig. 7

Schematic representation of the olivo-cortico-nuclear interconnectivity. Note that each module, apart from their interconnected parts, also connects to the rest of the brain by their output (double arrowheads). Also note that the cortical part of each module is formed by one or several stripes of either zebrin II + or zebrin II- Purkinje cells. The mediolateral order of the represented modules, indicated from left to right in the diagram, is based on the mediolateral position of the CN and is not related to the mediolateral position of the cortical components. For example, the cortical A2 modules are located lateral to the cortical B-module [see 138]. IntDL receives input from zebrin II-negative PCs of cortical module D0, which is interspersed between zebrin II ⁺ stripes D1 and D2 and receives its olivary input from a part of the principal olive (IOPdm). Different shades of yellow, blue, and purple refer to Med, Int, and Lat modules, respectively. The green module is related to the lateral vestibular nucleus (LV). Note that modules of the vestibulocerebellum, i.e., with cortical input form nodulus and flocculus, are not indicated in this scheme. Abbreviations. Inferior olive (IO) per module from left to right: cIOMa, group a of caudal medial accessory olive (cIOM); cIOMb, group b of cIOM; cIOMbe, group beta of cIOM; cIOMd, group d of cIOM; cIOMc, group c of cIOM; IODdf, dorsal fold of dorsal inferior olive (IOD); IODvf, ventral fold of IOD; iIOM, intermediate part of medial accessory olive (IOM); rIOM, rostral part of IOM; IODvf, ventral fold of IOD; IOPvf, ventral fold of principal olive (IOP); IOPdm, dorsomedial group of IOP; IOPdf, dorsal fold of IOP. CX-zone: sagittally oriented zones of Purkinje cells in the cerebellar cortex indicated by capital letters A to D followed by either a number or a lowercase ‘x’ and related to zebrin-positive or zebrin-negative stripes. Cerebellar nuclei (CN) from left to right: vMed, ventral part of medial cerebellar nucleus (Med); rMed, rostral part of Med; cMed, caudal part of Med; MedcDL, caudal part of dorsolateral protuberance of Med; MedrDL, rostral part of dorsolateral protuberance of Med; IntIC, interstitial cell groups; LV, lateral vestibular nucleus; IntA, anterior interposed nucleus; IntIC, interstitial cell groups; IntP, posterior interposed nucleus; IntA, anterior interposed nucleus; vLat, ventral part of lateral cerebellar nucleus (lat); IntDL, dorsolateral hump; dLat, dorsal part of the Lat. Based on [20, 72, 139]

Fig. 8

3D representations of CN projections to the brainstem and thalamus in the mouse visualized using recombinant anterogradely transported adeno-associated virus (rAAV) as a tracer injected in different parts of the CN. Panels from top to bottom depict a lateral, dorsal, and caudal transparent 3D view with a final section showing the injection site. a Injection centered on the Med, depicting prominent bilateral terminal labeling in the vestibular nuclei and medial reticular formation. Note the conspicuous intracerebellar course of the uncinate fascicle. b Injection centered on the IntA. Apart from the course of labeled fibers to the midbrain and thalamus by way of the scp, note the prominent course of ipsilaterally descending fibers, which seems to be a special feature of rodent connectivity. c Injection centered on the Lat. Here, aspects of an ipsilateral descending tract together with a contralateral descending tract terminating in the medullary reticular formation can be appreciated. See text for further explanation. Yellow, cyan, and magenta circles indicate the approximate sites of the Med, IntA and Lat injections respectively, in the 3D representations. Double arrowheads in the third row point to the nucleocortical projections seen in all cases. Abbreviations: cdt, contralateral descending tract; dfbt, direct fastigiobulbar tract; IC, inferior colliculus; idt, ipsilateral descending tract; IntA, anterior interposed nucleus; Lat, lateral cerebellar nucleus; Med, medial cerebellar nucleus; SC, superior colliculus; scp, superior cerebellar peduncle; Th, thalamus; un, uncinate fascicle; based on material from [145], experiment numbers 268389532, 120, 493, 315, 127, 650, 431)

Fig. 9

Schematic representation of the CN cell types and their connectivity. Numbers refer to descriptions in Table 3. Abbreviations: ML, molecular layer; GCL, granular layer; GoC, Golgi cell; PC, Purkinje cell; GC, granule cell; Med, medial CN (hosting the “exceptional” glycinergic projection neuron type labeled 6); VN, vestibular nucleus; ipsilat, ipsilateral; IO, inferior olive; contralat, contralateral; scp, superior cerebellar peduncle. The question mark indicates unknown targets of local interneuron axons

Fig. 10

In situ hybridization demonstrating expression patterns of astroglial markers, Aqp4 and Gat-3. Sagittal sections through the lateral vermis. CN are either negatively outlined by staining for Aqp4, or positively by staining for Gat-3. In contrast, astroglial cells in the white matter and the granule cell layer, and also Bergman glial cells, strongly express the mRNA for Aqp4, but not that for Gat-3. Arrows give orientation (r, rostral, c, caudal, d, dorsal, v, ventral. Scale bar = 1 mm. [145]

Fig. 11

Schematic depiction of the conceptual differences between an approach that considers the cerebellar cortex and the nuclei as independent functional units (a) and a view of cerebellar computation where information processing is not segmented into “cortical” and “nuclear” parts (b). The latter scheme is a natural extension of the nucleocentric view promoted in this review. From the viewpoint of the whole brain, the cerebellar system appears as a unified and modular computational system. CX, cerebellar cortex; CN, cerebellar nuclei; NC, nucleocortical pathway; MF, mossy fibers; CF, climbing fibers; IO, inferior olive; NO, nucleo-olivary fibers (cf. Figure 1)

Fig. 12

Schematic drawing showing migratory steps of GABAergic and glutamatergic neurons composing the cerebellar nuclei. All sagittal sections are oriented with rostral to the right. a Between E10.5 and E11.5 glutamatergic projection neurons (light blue cells) originate from the ATOH1 ⁺ and PAX6 ⁺ progenitors (light blue spheroids) localized in the Rhombic Lip (Rl, Triangular shape). They migrate apposed to the pia mater, forming the subpial stream (SPS, curved line) away from the Rl (gray arrow) expressing the genes indicated in Box 1. Few neurons reach the nuclear transitory zone (Ntz). GABAergic neurons differentiate from ASCL1 ⁺ and PTF1A ⁺ common progenitors (green spheroids) in the ventricular zone (Vz, rectangular shape) from which NEUROG2 ⁺ or PAX2 ⁺ transit amplifier populations originate (green spheroids). Postmitotic NEUROG2 ⁺ or PAX2 ⁺ GABAergic (green cells) neurons leave the Vz expressing IRX3 or Dmbx1. b Between E12.5 and E13.5 glutamatergic projection neurons continue their migration expressing several markers (Box 2) and start to reach the Ntz, where they express other markers (Box 3). GABAergic neurons move towards the Ntz and express SOX14 and DMBX1 while IRX3 is downregulated. Neurogenesis of NEUROG2 ⁺ and PAX2 ⁺ neurons appears to continue until E12.25-E12.5. c By E14.5 all the prospective glutamatergic CN neurons are located in the Ntz surrounded by a GABAergic population, where they further mature and prepare to descend into their final position in the central mass (grey arrows). d By E18.5 all the neurons are in the central mass, occupying different territories: glutamatergic neurons localize dorsally while NEUROG2 ⁺ GABAergic neurons are in ventral and lateral positions. A few PAX2 ⁺ GABAergic neurons are intermingled in the neuronal mass, while the prospective PAX2 ⁺ GABAergic interneurons surround the cerebellar nuclei mass. e By P4, the GABAergic and glutamatergic neurons continue small movements to reach their terminal destination. Color fonts of expressed molecules during development indicate the known destination of the cells. The numbers of cells illustrated do not reflect the actual cell numbers, which are mostly unknown

Fig. 13

In situ hybridization of molecular markers that allow the identification of excitatory (Slc17a6, also known as Vglut2) or various inhibitory [Dmbx1, Sox14, Slc6a5 (also known as GlyT2), Pax2] CN neurons at E13.5 and E15.5. For comparison, PCs that express Calbindin 1 (also known as Calbindin D28k) are also shown in the last panel. The top and middle rows show sagittal sections taken from midway between the midline and the lateral border of the cerebellar anlage. The bottom row shows images taken more laterally. At E15.5, the areas occupied by Sox14⁺ /Dmbx1⁺ cells appear to overlap with those occupied by Pax2⁺ cells, but not with territories in which excitatory (Slc17a6⁺) or Calbindin1⁺ Purkinje cells are found. Arrows give orientation (r, rostral, c, caudal, d, dorsal, v, ventral. Scale bars = 0.5 mm (top row for E13.5; bottom row for E15.5) [145]

Fig. 14

In situ hybridization showing expression of genes that allow the identification of subsets of cells at E13.5 that assemble into the CN. Tbr2, Pax5, and Lmo3 mark distinct but apparently partly overlapping sets of cells in the classical NTZ, considered precursors of glutamatergic CN neurons. Dmbx1 and Sox14 are markers of inhibitory neurons projecting to the inferior olive. Pax2 ⁺ precursors contribute inhibitory interneurons to the CN. Ret and Kit are representative of genes expressed in the CN from at least E13.5 onward and into adulthood. Numbers in individual panels refer to the scheme illustrated in the lower right corner. Arrows give orientation (r, rostral, c,caudal, d, dorsal, v, ventral. Scale bar = 0.5 mm [145]

Fig. 15

Proposed diversification of the CN in vertebrates, visualizing the appearance of new CN subdivisions during evolution alongside increasing behavioral complexity. Note that in several shark species the CN is clearly divided into two parts, but their functional independence in terms of downstream connectivity has not been examined. SPS, substitutions per site in the dendrogram, reflecting relative amount of genetic changes since the previous branch. Dendrogram based on [130] and [422]