The valvula cerebelli of the spiny eel,Macrognathus aculeatus,receives primary lateral-line afferents from the rostrum of the upper jaw

Summary In the spiny eel, Macrognathus aculeatus, anterodorsal and (to a lesser degree) anteroventral lateralline nerves project massively to the granular layer of the valvula cerebelli, throughout its rostrocaudal extent. The posterior lateral-line nerve terminates in the corpus cerebelli. Thus, valvula and corpus cerebelli are supplied with mechanosensory input of different peripheral origins. An analysis of the taxonomic distribution of experimentally determined primary lateral-line input to the three parts of the teleostean cerebellum reveals that the eminentia granularis always receives such input, and that the corpus cerebelli is the recipient of primary lateral-line input in many teleosts. The valvula, however, receives primary lateral-line afferents in only two examined species. In M. aculeatus, the massive lateral-line input to the valvula probably originates in mechanoreceptors located in the elongated rostrum of the upper jaw, a characteristic feature of mastacembeloid fishes. This projection to the valvula may therefore represent a unique specialization that arose with the evolution of the peculiar rostrum.     

Comparative aspects of cerebellar cytoarchitecture in the European eel life cycle

The major anatomical divisions of the cerebellum of the European eel, i.e., corpus cerebelli, lobus vestibulolateralis, and valvula, were studied morphologically and morphometrically. There were differences in cerebellar cytoarchitecture and gross morphology in two stages of the eel life cycle, the trophic stage (yellow eel), and the reproductive stage (silver eel), which are characterized by different degrees of swimming activity. The principal differences between silver and yellow eels in the cytoarchitecture of the corpus cerebelli and the lobus vestibulolateralis were in distribution of Purkinje or Purkinje-like cells in the molecular layer, which is wider in silver eels, in part because of a decreased thickness of the granular cell layer. In the silver eel, the scattering of Purkinje cells was more evident in the lobus vestibulolateralis where the molecular layer is also thicker. The data indicate the transition from the yellow eel to the silver eel is characterized by a migration of granule cells from the ganglionic cell layer to the internal granular layer and by a further development of molecular layer components, e.g., parallel fibers, Purkinje-cell dendrites, etc. In contrast, the thickness of the granular layer and of the Purkinje cell layer, limited to the lower part of the valvula, decreased. There is also a slight increase of cerebellar volume in the silver eel. The volume of the lobus vestibulolateralis was constant. Hypertrophy of the valvula and eminentiae granulares is observed and is due to the migration of cells from the granular layer of the corpus cerebelli whose volume slightly decreases. Perhaps the lobus vestibulolateralis also contributes to the increased volume of eminentiae granulares. Our findings suggest that the cerebellum continues to develop during the passage from the trophic to the reproductive stage of the eel. The appearance of new afferents from the lateral line which becomes more visible in the silver eel probably completes cerebellar ontogeny.     

Early development of the cerebellum in teleost fishes: a study based on gene expression patterns and histology in the medaka embryo

The cerebellar structures of teleosts are markedly different from those of other vertebrates. The cerebellum continues rostrally into the midbrain ventricle, forming the valvula cerebelli, only in ray-finned fishes among vertebrates. To analyze the ontogenetic processes that underlie this morphological difference, we examined the early development of the cerebellar regions, including the isthmus (mid/hindbrain boundary, MHB), of the medaka (Oryzias latipes), by histology and in-situ hybridization using two gene (wnt1 and fgf8) probes. Isthmic wnt1 was expressed stably in the caudalmost mesencephalic region in the neural tube at all developmental stages examined, defining molecularly the caudal limit of the mesencephalon. The wnt1-positive mesencephalic cells became located rostrally to the isthmic constriction at Iwamatsu's stages 25-26. Isthmic fgf8 expression changed dynamically and became restricted to the rostralmost metencephalic region at stage 24. The rostralmost part (prospective valvula cerebelli) of the fgf8-positive rostral metencephalon protruded rostrally into the midbrain ventricle, bypassing the isthmic constriction, at stages 25-26. Thus, the isthmic constriction shifted caudally with respect to the molecularly defined MHB at stages 25-26. Paired cerebellar primordia were formed from the alar plates of the fgf8-positive rostral metencephalon and the fgf8-negative caudal metencephalon in the medaka neural tube. Our results show that cerebellar development differs between teleosts and murines: both the rostral and caudal metencephalic alar plates develop into the cerebellum in medaka, whereas in the murines only the caudal metencephalic alar plate develops into the cerebellum, and the rostral plate is reduced to a thin membrane.     

Adult neurogenesis in the brain of the Mozambique tilapia, Oreochromis mossambicus

Although the generation of new neurons in the adult nervous system ('adult neurogenesis') has been studied intensively in recent years, little is known about this phenomenon in non-mammalian vertebrates. Here, we examined the generation, migration, and differentiation of new neurons and glial cells in the Mozambique tilapia (Oreochromis mossambicus), a representative of one of the largest vertebrate taxonomic orders, the perciform fish. The vast majority of new cells in the brain are born in specific proliferation zones of the olfactory bulb; the dorsal and ventral telencephalon; the periventricular nucleus of the posterior tuberculum, optic tectum, and nucleus recessi lateralis of the diencephalon; and the valvula cerebelli, corpus cerebelli, and lobus caudalis of the cerebellum. As shown in the olfactory bulb and the lateral part of the valvula cerebelli, some of the young cells migrate from their site of origin to specific target areas. Labeling of mitotic cells with the thymidine analog 5-bromo-2'-deoxyuridine, combined with immunostaining against the neuron-specific marker protein Hu or against the astroglial marker glial fibrillary acidic protein demonstrated differentiation of the adult-born cells into both neurons and glia. Taken together, the present investigation supports the hypothesis that adult neurogenesis is an evolutionarily conserved vertebrate trait.     

Zonal distribution of Purkinje cells in the zebrafish cerebellum: analysis by means of a specific monoclonal antibody

We have isolated a monoclonal antibody that recognizes a 42-kDa protein from adult zebrafish brain. The antibody stains the typical drop-shaped perikaryon of Purkinje cells and their dendrites. The cerebellum of teleosts has complex features. It is composed of three parts; the valvula cerebelli (Va), the corpus cerebelli (CCe), and the crista cerebellaris (CC). In higher vertebrates, the molecular layer is always found as the most outer layer of the cerebellum, but in teleosts, some of the granular cells are located on the surface of the Va. In higher vertebrates, the boundary between the granular and molecular layers always contains Purkinje cells, but this does not occur in teleosts. The Purkinje cells are found only in a part of the boundary in Va. We have found that the layer containing Purkinje cells forms a continuous zone in the cerebellum in the zebrafish. The complex structure of the cerebellum is more easily understood with the aid of the concept of a "Purkinje zone". The Purkinje zone starts at the caudal end of Val (lateral division of Va), turns at the edge of Va toward Vam (medial division of Va), connects to CCe, and ends at the bottom of CCe. The dendrites are found only on one side of the zone. The dendrites of the Purkinje cells in Vam are planar and are packed regularly, similar to those of higher vertebrates. However, the dendrites in Val and the posterior part of CCe are not planar and are irregularly packed.     

Postnatal differentiation of the Golgi apparatus and the dendrites of Purkinje cells of the rat cerebellum

Summary The morphology of postnatal differentiation of the Golgi apparatus, the nucleus, the perikaryon, and the dendrites was studied in Purkinje cells of the rat cerebellum for 30 days after birth using histochemical, histological, and electron microscopic methods.The Golgi apparatus during differentiation undergoes morphological and positional changes. From the 1st to 7th postnatal day, the Golgi apparatus is found in a supranuclear position, and is connected with the axes of differentiating primary dendrites by beam-like processes. From days 8 to 11 this connection disappears, and most of the Golgi apparatus assumes a lateronuclear and infranuclear position. After the 11th or 12th day, the Golgi apparatus is found in perinuclear and peripheral cytoplasmic positions. The formation of granular endoplasmic reticulum occurs in the vicinity of the perinuclear Golgi apparatus. The differentiation of cell and nuclear forms requires approximately 20 days. The morphological changes of differentiation are discussed in relation to the participation of the Golgi apparatus in the differentiation of dendrites and in the formation of the granular endoplasmic reticulum.     

The zebrafish cerebellar rhombic lip is spatially patterned in producing granule cell populations of different functional compartments

The upper rhombic lip, a prominent germinal zone of the cerebellum, was recently demonstrated to generate different neuronal cell types over time from spatial subdomains. We have characterized the differentiation of the upper rhombic lip derived granule cell population in stable GFP-transgenic zebrafish in the context of zebrafish cerebellar morphogenesis. Time-lapse analysis followed by individual granule cell tracing demonstrates that the zebrafish upper rhombic lip is spatially patterned along its mediolateral axis producing different granule cell populations simultaneously. Time-lapse recordings of parallel fiber projections and retrograde labeling reveal that spatial patterning within the rhombic lip corresponds to granule cells of two different functional compartments of the mature cerebellum: the eminentia granularis and the corpus cerebelli. These cerebellar compartments in teleosts correspond to the mammalian vestibulocerebellar and non-vestibulocerebellar system serving balance and locomotion control, respectively. Given the high conservation of cerebellar development in vertebrates, spatial partitioning of the mammalian granule cell population and their corresponding earlier-produced deep nuclei by patterning within the rhombic lip may also delineate distinct functional compartments of the cerebellum. Thus, our findings offer an explanation for how specific functional cerebellar circuitries are laid down by spatio-temporal patterning of cerebellar germinal zones during early brain development.     

MALNUTRITION AND BRAIN DEVELOPMENT: CEREBELLAR WEIGHT, DNA, RNA, PROTEIN AND HISTOLOGICAL CORRELATIONS

Abstract— Rat pups were reared in litters of 20 and litters of 6 to study effects of malnutrition on cerebellar development. Cell production and cell content were determined by assaying for DNA, as a measure of cell number, and RNA and protein, as indicators of cell constituents. By comparing DNA contents at 3, 4, 8, 11, 14, 17, 21, and 28 days after birth, we concluded that (a) there is little nutritional reserve at birth since significant differences appear by day 4, (b) most relative differences between groups appeared by day 8, with absolute differences increasing to day 21, and (c) there is partial recovery of cell number and cell constituents in the malnourished rats between 21 and 28 days.
Areal measurements of histological preparations showed that malnutrition resulted in less total area in cerebellar midsagittal sections at days 8, 11. and 14. In malnourished animals, the germinal matrix area of the cerebellum, the external granular layer, was smaller on the 8th postnatal day, the same on the eleventh day, and larger on the fourteenth day when compared with that of well fed animals. At all three ages alterations could be discerned in the distribution of cells between the mitotic external mantle and nonmitotic internal matrix portions of the external granular layer.
Further studies involving exchanging animals between large and small litters at various ages indicated that the time around days 4 to 8 is most sensitive to malnutrition. The results suggest a process in which malnutrition exerts its maximum effect by a slowing of cell production in the external granular layer in the initial exponential growth phase. It is likely that an adaptation occurs immediately in the external granular layer which subsequently permits a partial recovery of cerebellar growth between days 21 and 28.     

Neurotrophins and their Trk‐receptors in the cerebellum of zebrafish

Neurotrophins (NTs) and their specific Trk‐receptors are key molecules involved in the regulation of survival, proliferation, and differentiation of central nervous system during development and adulthood in vertebrates. In the present survey, we studied the expression and localization of neurotrophins and their Trk‐receptors in the cerebellum of teleost fish Danio rerio (zebrafish). Teleostean cerebellum is composed of a valvula, body and vestibulolateral lobe. Valvula and body show the same three‐layer structure as cerebellar cortex in mammals. The expression of NTs and Trk‐receptors in the whole brain of zebrafish has been studied by Western blotting analysis. By immunohistochemistry, the localization of NTs has been observed mainly in Purkinje cells; TrkA and TrkB‐receptors in cells and fibers of granular and molecular layers. TrkC was faintly detected. The occurrence of NTs and Trk‐receptors suggests that they could have a synergistic action in the cerebellum of zebrafish. J. Morphol. 277:725–736, 2016. © 2016 Wiley Periodicals, Inc.     

Comparative aspects of cerebellar organization. From mormyrids to mammals.

Recent progress in the comparative analysis of the vertebrate cerebellar organization shows that the cerebella of different tetrapods have a basically similar intrinsic organization, whereas the cerebellum of fishes displays a number of fundamental differences in this respect. Clear examples of teleostean cerebellar specializations are present in the gigantocerebellum of mormyrids, including a valvula cerebelli, the absence of a parasagittal zonal organization, the presence of eurydendroid projection neurons instead of deep cerebellar nuclei, a precerebellar nucleus lateralis valvulae, olivocerebellar fibers that do not climb into the molecular layer, uni- and bilateral locations of granule cells, parallel fibers without a T-shaped bifurcation and with a coextensive distribution in the transverse plane, and different Purkinje cell arrangements including a dendritic palisade pattern. A theoretical exploration of the possible significance of these configurations suggests that they all might be involved in a single main cerebellar function, i.e. coincidence detection of parallel fiber activity by Purkinje cells.     

Development of longitudinal patterns in the cerebellum of the chicken (Gallus domesticus): a cytoarchitectural study on the genesis of cerebellar modules.

The cytoarchitecture of the cerebellum has been studied in chicken embryos from day 3-20 using serial sections stained with cresylviolet, haematoxylin-eosin and toluidine blue. Three periods have been distinguished in cerebellar development on a basis of cytoarchitectonic characteristics. Of these periods the middle one, which lasts from the 8th to the 15th day, is marked by two subsequent transient longitudinal cytoarchitectonic patterns in the cortical anlage. The first pattern, which exists between days 8 and 11, consists of 4 longitudinal Purkinje cell clusters (of the first order) at either side of the midline. The second pattern, which is most distinct and complete during embryonic days 12-14, is caused by specific localizations of otherwise few, early inwardly migrating granule cells from the external cerebellar matrix (so-called granule raphes), which pass through the layer of Purkinje cell clusters of the first order and thus subdivide these latter into smaller entities: Purkinje cell clusters of the second order. The number of these latter (6 or 7 and 11 or 12 in the anterior and posterior lobes, respectively) correspond to the number of parasagittal modules, which can be discerned on a basis of the organization of fiber connections of the adult cerebellar cortex. Thanks to this similarity various hypotheses can be formulated concerning the significance of the transient cytoarchitectonic patterns in the primitive cortex for the genesis of the modular organization of the cerebellum.     

Microbial carbohydrate specific antibodies distinguish between different stages of differentiating mouse cerebellum

High titered anticarbohydrate antibodies were used to identify cell surface carbohydrates during different stages in histogenesis of mouse cerebellum in a micro tissueculture system which mimics selected features of in vivo cerebellum development. Blockage of fiber formation within the first few days in vitro and inhibition of cell migrations by carbohydrate-specific antibodies served as an assay system for possible contributions of surface carbohydrates to the behavior of developing cerebellar cells. Microbial strains were selected on the basis of carbohydrate structures of their cell wall antigens, and anticarbohydrate antibodies were raised against treated whole bacteria and yeast in rabbits. We found that antibodies to mannan were active at all stages of development tested (embryonic day 13, E13; the day of birth, P0; and postnatal day 7, P7). Antibodies to sialic acids prepared against strains B and C of Neisseria meningitidis distinguish different subterminal structures: anti-B reacted with E 13 and P0 cerebellar cells, and anti-C mostly with cells older than P7. Antifetuin antibody recognized E 13 and P0 but not P7 cell populations. Pneumococcus C strain R36A-specific antibodies were effective only after coating cells to C type carbohydrate before application of the antibody. The results demonstrate that antimicrobial carbohydrate antibodies cross-react with mammalian cell surface carbohydrate structures and therefore can be used as a powerful tool in tissue culture to analyze those structures which might control cell behaviors pertinent to cerebellar development.     

Chromatin changes in frog neurons after eighth nerve transection

Fourteen days after unilateral eighth nerve transection in the frog, Purkinje neurons of the lobus vestibulolateralis and corpus of the cerebellum and medium-sized neurons of the vestibular nuclear complex showed changes in metabolic activity. In the ipsilateral parts, and to a lesser extent in the contralateral parts, of operated frogs, the Feulgen-DNA values were higher and the nuclear areas larger, associated with decondensation of chromatin. The cytoplasmic basophilia was also less. These changes could be due to anabolic responses of the neuronal populations during regeneration. The anabolic reaction of the corpus cerebelli and contralateral vestibular nuclear complex is only partially non-specific and ascribable to the surgical trauma (comparison between sham-operated and unoperated frogs). The results indicate clear patterns of connection between the ipsilateral and contralateral parts and between the cerebellar and vestibular nuclear complex neurons.     

Adaptive processing in electrosensory systems: Links to cerebellar plasticity and learning

The first central stage of electrosensory processing in fish takes place in structures with local circuitry that resembles the cerebellum. Cerebellum-like structures and the cerebellum itself share common patterns of gene expression and may also share developmental and evolutionary origins. Given these similarities it is natural to ask whether insights gleaned from the study of cerebellum-like structures might be useful for understanding aspects of cerebellar function and vice versa. Work from electrosensory systems has shown that cerebellum-like circuitry acts to generate learned predictions about the sensory consequences of the animals’ own behavior through a process of associative plasticity at parallel fiber synapses. Subtraction of these predictions from the actual sensory input serves to highlight unexpected and hence behaviorally relevant features. Learning and prediction are also central to many current ideas regarding the function of the cerebellum itself. The present review draws comparisons between cerebellum-like structures and the cerebellum focusing on the properties and sites of synaptic plasticity in these structures and on connections between plasticity and learning. Examples are drawn mainly from the electrosensory lobe (ELL) of mormyrid fish and from extensive work characterizing the role of the cerebellum in Pavlovian eyelid conditioning and vestibulo-ocular reflex (VOR) modification. Parallels with other cerebellum-like structures, including the gymnotid ELL, the elasmobranch dorsal octavolateral nucleus (DON), and the mammalian dorsal cochlear nucleus (DCN) are also discussed.     

泥鳅脑的形态构造观察

观察了泥鳅（Misgurnus Anguillicaudatus）脑的显微结构。结果表明,泥鳅的脑组织基本结构与多数鱼类相一致,其脑轮廓狭长,包括端脑、间脑、中脑、小脑及延脑五部分,视叶隆起,小脑瓣突入中脑室内,延脑有核团的分化。同时与其生存环境和捕食习性相适应,泥鳅脑具有一些原始的特征,包括：嗅叶及嗅束分化较明显,大脑呈长椭圆状,仍保留鲤科（Cyprinidae）鱼类脑的原始特征;中脑视叶壁偏薄,与视觉不甚发达相关;小脑不发达,与其喜静的生活习性相适应;延脑前部稍稍隆起,面叶发达与其须感知食物的习性相符。     

Effects of methylazoxymethanol given at different stages of postnatal life on development of the rat brain. Comparison with those of thyroid deficiency

Newborn rats were treated at different stages of their development with low doses of methylazoxymethanol acetate. The postnatal increase of the DNA content of the cerebrum did not differ from that of controls. In the cerebellum, the DNA content was transitorily reduced, but later, the external granular layer became thicker and DNA deposition increased in comparison with controls; finally, the cerebellar DNA returned to a normal value. Morphological abnormalities of the cerebellum, abnormal orientation of migrating cells, scattering of Purkinje cell bodies within the internal granule cells and specially striking abnormalities of the morphology and orientation of Purkinje cell dendrites were noted in rats treated with MAM from birth to day 3. The effects on the Purkinje cell morphogenesis persisted but were much less marked when MAM was given from 4 to 7 or from 8 to 11 days. Neonatal thyroid deficiency, as MAM-treatment between days 0 and 3, leads to an abnormal position of Purkinje cell bodies within the cerebellar cortex; it also leads to morphological abnormalities of their dendritic arborization which closely resemble those observed after MAM-treatment during the second postnatal week. It also alters the cell formation in the cerebellum. Thyroid deficiency probably exerts its effect on cell formation earlier than previous biochemical studies have shown. On another hand, the morphological abnormalities of Purkinje cell arborizations in the thyroid-deficient animals may be partly due to the perturbations of cell formation which persist later in the cerebellum.     

A 3H-thymidine autoradiographic study of the development of the cerebellum of the White Leghorn (Gallus domesticus): "evidence for longitudinal neuroblast generation patterns"

Dynamic aspects of the development of the cerebellum of the white Leghorn (Gallus domesticus) were examined using 3H-thymidine autoradiography. A single dose of 25 microCi 3H-thymidine was given to embryos varying in age from 0-18 days. The embryos were all studied before hatching, either after short or after long survival times. It appeared that the superficial part of the outer mantle layer (OML-s, or called external granular layer, egl) is histologically present at day 6, but starts its activity as a secondary matrix at day 7, probably due to activation by a secondary influx of cells from the ventricular neuro-epithelium. It was also demonstrated that Purkinje cells and cells of the central cerebellar nuclei are produced by the ventricular neuro-epithelium mainly at days 3, 4 and 5. No birth of these cells was observed beyond day 5. In addition, a relationship was found between the birth patterns of Purkinje cells and cells of the central cerebellar nuclei and the longitudinal cytoarchitectonic patterns as observed in normal Nissl stained material. It was concluded that these birth patterns are an early contribution to the establishment of the longitudinal organization of the cerebellum.     

Transgenic mice expressing F3/contactin from the TAG-1 promoter exhibit developmentally regulated changes in the differentiation of cerebellar neurons

F3/contactin (CNTN1) and TAG-1 (CNTN2) are closely related axonal glycoproteins that are differentially regulated during development. In the cerebellar cortex TAG-1 is expressed first as granule cell progenitors differentiate in the premigratory zone of the external germinal layer. However, as these cells begin radial migration, TAG-1 is replaced by F3/contactin. To address the significance of this differential regulation, we have generated transgenic mice in which F3/contactin expression is driven by TAG-1 gene regulatory sequences, which results in premature expression of F3/contactin in granule cells. These animals (TAG/F3 mice) display a developmentally regulated cerebellar phenotype in which the size of the cerebellum is markedly reduced during the first two postnatal weeks but subsequently recovers. This is due in part to a reduction in the number of granule cells, most evident in the external germinal layer at postnatal day 3 and in the inner granular layer between postnatal days 8 and 11. The reduction in granule cell number is accompanied by a decrease in precursor granule cell proliferation at postnatal day 3, followed by an increase in the number of cycling cells at postnatal day 8. In the same developmental window the size of the molecular layer is markedly reduced and Purkinje cell dendrites fail to elaborate normally. These data are consistent with a model in which deployment of F3/contactin on granule cells affects proliferation and differentiation of these neurons as well as the differentiation of their synaptic partners, the Purkinje cells. Together, these findings indicate that precise spatio-temporal regulation of TAG-1 and F3/contactin expression is critical for normal cerebellar morphogenesis.     

Developmental Expression and Functional Characterization of the 4C5 Antigen in the Postnatal Cerebellar Cortex

Abstract: The monoclonal antibody 4C5 recognizes a neuron-specific surface antigen (4C5 antigen) in the CNS and PNS of the rat. In the present study we investigated the expression of 4C5 antigen in the developing cerebellum of the rat and the functional role of this molecule during cerebellar morphogenesis. Immunoblotting and immunohistochemistry in sections of cerebellar cortex revealed an age-dependent decrease in the expression of the 4C5 antigen. In cerebellar primary cell cultures, 4C5 immunoreactivity was detected both on granule and on Purkinje neurons. Granule cell migration was inhibited in cerebellar explants derived from 8-day-old rats and cultured for 2 days in the presence of antibodies against the 4C5 antigen. Electron microscope immunocytochemistry revealed that in 8-day-old rat cerebellum, 4C5 immunoreactivity was localized on the cell bodies of granule neurons in the external and internal granular layers and on parallel fibers in the developing molecular layer as well as at contact sites between these cellular elements. It was not detected on Bergmann glia. These results suggest strongly that the 4C5 antigen is involved in granule cell migration during cerebellar development, possibly via neuron-neuron interactions.