Conserved and divergent development of brainstem vestibular and auditory nuclei.

Vestibular function was established early in vertebrates and has remained, for the most part, unchanged. In contrast, each group of tetrapods underwent independent evolutionary processes to solve the problem of hearing on land, resulting in a remarkable mixture of conserved, divergent and convergent features that define extant auditory systems. The vestibuloacoustic nuclei of the hindbrain develop from a highly conserved ground plan and provide an ideal framework on which to address the participation of developmental processes to the evolution of neuronal circuits. We employed an electroporation strategy to unravel the contribution of two dorsoventral and four axial lineages to the development of the chick hindbrain vestibular and auditory nuclei. We compare the chick developmental map with recently established genetic fate-maps of the developing mouse hindbrain. Overall, we find considerable conservation of developmental origin for the vestibular nuclei. In contrast, a comparative analysis of the developmental origin of hindbrain auditory structures echoes the complex evolutionary history of the auditory system. In particular, we find that the developmental origin of the chick auditory interaural time difference circuit supports its emergence from an ancient vestibular network, unrelated to the analogous mammalian counterpart.

Chondroitin sulfates in the developing rat hindbrain confine commissural projections of vestibular nuclear neurons.

BACKGROUND: Establishing correct neuronal circuitry is crucial to proper function of the vertebrate nervous system. The abundance of chondroitin sulfate (CS) proteoglycans in embryonic neural environments suggests that matrix proteoglycans regulate axonal projections when fiber tracts have not yet formed. Among the early-born neurons, the vestibular nucleus (VN) neurons initiate commissural projections soon after generation at E12.5 and reach the contralateral target by E15.5 in the rat hindbrain. We therefore exploited 24-hour cultures (1 day in vitro (DIV)) of the rat embryos and chondroitinase ABC treatment of the hindbrain matrix to reveal the role of CS moieties in axonal initiation and projection in the early hindbrain. RESULTS: DiI tracing from the VN at E12.5(+1 DIV) showed contralaterally projecting fibers assuming fascicles that hardly reached the midline in the controls. In the enzyme-treated embryos, the majority of fibers were unfasciculated as they crossed the midline at 90°. At E13.5(+1 DIV), the commissural projections formed fascicles and crossed the midline in the controls. Enzyme treatment apparently did not affect the pioneer axons that had advanced as thick fascicles normal to the midline and beyond, towards the contralateral VN. Later projections, however, traversed the enzyme-treated matrix as unfasciculated fibers, deviated from the normal course crossing the midline at various angles and extending beyond the contralateral VN. This suggests that CSs also limit the course of the later projections, which otherwise would be attracted to alternative targets. CONCLUSIONS: CS moieties in the early hindbrain therefore control the course and fasciculation of axonal projections and the timing of axonal arrival at the target.

Electrophysiological properties of morphologically-identified medial vestibular nucleus neurons projecting to the abducens nucleus in the chick embryo.

Neurons in the medial vestibular nucleus (MVN) show a wide range of axonal projection pathways, intrinsic firing properties, and responses to head movements. To determine whether MVN neurons participating in the vestibulocular reflexes (VOR) have distinctive electrophysiological properties related to their output pathways, a new preparation was devised using transverse brain slices containing the chicken MVN and abducens nucleus. Biocytin Alexa Fluor was injected extracellularly into the abducens nucleus so that MVN neurons whose axons projected to the ipsilateral (MVN/ABi) and contralateral (MVN/ABc) abducens nuclei were labeled selectively. Whole-cell, patch-clamp recordings were performed to study the active and passive membrane properties, sodium conductances, and spontaneous synaptic events in morphologically-identified MVN/AB neurons and compare them to MVN neurons whose axons could not be traced (MVN/n). Located primarily in the rostral half of the ventrolateral part of the MVN, MVN/AB neurons mainly have stellate cell bodies with diameters of 20-25 µm. Compared to MVN/n neurons, MVN/ABi and MVN/ABc neurons had lower input resistances. Compared to all other MVN neuron groups studied, MVN/ABc neurons showed unique firing properties, including type A-like waveform, silence at resting membrane potential, and failure to fire repetitively on depolarization. It is interesting that the frequency of spontaneous excitatory and inhibitory synaptic events was similar for all the MVN neurons studied. However, the ratio for miniature to spontaneous inhibitory events was significantly lower for MVN/ABi neurons compared to MVN/n neurons, suggesting that MVN/ABi neurons retained a larger number and/or more active inhibitory presynaptic neurons within the brain slices. Also, MVN/ABi neurons had miniature excitatory postsynaptic currents (mEPSCs) with slower decay time and half width compared to MVN/n neurons. Altogether, these findings underscore the diversity of electrophysiological properties of MVN neuron classes distinguished by axonal projection pathways. This represents the first study of MVN/AB neurons in brain slice preparations and supports the concept that the in vitro brain slice preparation provides an advantageous model to investigate the cellular and molecular events in vestibular signal processing.

Dye coupling in developing vestibular nuclei.

The chick tangential nucleus is a major vestibular nucleus whose principal cells participate in the vestibular reflexes. During development, most mature vestibular nucleus neurons must acquire repetitive firing of action potentials on depolarization and spontaneous spike activity to process signals effectively. In the chicken, these properties emerge gradually in the embryo, starting the week before (E13, E16) and continuing through the first week after hatching (H7). Since gap junction-mediated cell coupling may influence the emergence of neuronal excitability, we investigated whether neuron-neuron and neuron-glia coupling are present in this morphologically distinctive vestibular nucleus during the period for establishing signal processing. In brain slices, principal cells were injected with biocytin in the whole-cell configuration and visualized via confocal imaging at E13, E16, and H7. The incidence of dye coupling between the injected principal cell and neurons was 42% at E13, 75% at E16, and 7% at H7, whereas the incidence of dye coupling with glia was 100% at both embryonic ages but decreased to 27% by H7. For each injected principal cell at E13, one coupled neuron and 35 coupled glia were detected, whereas three coupled neurons and 12 coupled glia were observed at E16, and few if any coupled neurons and glia were detected at H7. These results suggest that neuron-neuron and neuron-glia coupling are developmentally regulated and present before, but not after, the onset of mature signal processing by these neurons. Thus, transient neuron-neuron and neuron-glia coupling may both play roles in establishing excitability in vestibular nucleus neurons during development.

Fate-mapping the mammalian hindbrain: segmental origins of vestibular projection neurons assessed using rhombomere-specific Hoxa2 enhancer elements in the mouse embryo.

As a step toward generating a fate map of identified neuron populations in the mammalian hindbrain, we assessed the contributions of individual rhombomeres to the vestibular nuclear complex, a major sensorimotor area that spans the entire rhombencephalon. Transgenic mice harboring either the lacZ or the enhanced green fluorescent protein reporter genes under the transcriptional control of rhombomere-specific Hoxa2 enhancer elements were used to visualize rhombomere-derived domains. We labeled functionally identifiable vestibular projection neuron groups retrogradely with conjugated dextran-amines at successive embryonic stages and obtained developmental fate maps through direct comparison with the rhombomere-derived domains in the same embryos. The fate maps show that each vestibular neuron group derives from a unique rostrocaudal domain that is relatively stable developmentally, suggesting that anteroposterior migration is not a major contributor to the rostrocaudal patterning of the vestibular system. Most of the groups are multisegmental in origin, and each rhombomere is fated to give rise to two or more vestibular projection neuron types, in a complex pattern that is not segmentally iterated. Comparison with studies in the chicken embryo shows that the rostrocaudal patterning of identified vestibular projection neuron groups is generally well conserved between avians and mammalians but that significant species-specific differences exist in the rostrocaudal limits of particular groups. This mammalian hindbrain fate map can be used as the basis for targeting genetic manipulation to specific subpopulations of vestibular projection neurons.

The development of descending projections from the brainstem to the spinal cord in the fetal sheep.

BACKGROUND: Although the fetal sheep is a favoured model for studying the ontogeny of physiological control systems, there are no descriptions of the timing of arrival of the projections of supraspinal origin that regulate somatic and visceral function. In the early development of birds and mammals, spontaneous motor activity is generated within spinal circuits, but as development proceeds, a distinct change occurs in spontaneous motor patterns that is dependent on the presence of intact, descending inputs to the spinal cord. In the fetal sheep, this change occurs at approximately 65 days gestation (G65), so we therefore hypothesised that spinally-projecting axons from the neurons responsible for transforming fetal behaviour must arrive at the spinal cord level shortly before G65. Accordingly we aimed to identify the brainstem neurons that send projections to the spinal cord in the mature sheep fetus at G140 (term = G147) with retrograde tracing, and thus to establish whether any projections from the brainstem were absent from the spinal cord at G55, an age prior to the marked change in fetal motor activity has occurred. RESULTS: At G140, CTB labelled cells were found within and around nuclei in the reticular formation of the medulla and pons, within the vestibular nucleus, raphe complex, red nucleus, and the nucleus of the solitary tract. This pattern of labelling is similar to that previously reported in other species. The distribution of CTB labelled neurons in the G55 fetus was similar to that of the G140 fetus. CONCLUSION: The brainstem nuclei that contain neurons which project axons to the spinal cord in the fetal sheep are the same as in other mammalian species. All projections present in the mature fetus at G140 have already arrived at the spinal cord by approximately one third of the way through gestation. The demonstration that the neurons responsible for transforming fetal behaviour in early ontogeny have already reached the spinal cord by G55, an age well before the change in motor behaviour occurs, suggests that the projections do not become fully functional until well after their arrival at the spinal cord.

Emergence of action potential generation and synaptic transmission in vestibular nucleus neurons.

Principal cells of the chick tangential nucleus are vestibular nucleus neurons in the hindbrain. Although detailed information is available on the morphogenesis of principal cells and synaptogenesis of primary vestibular fibers, this is the first study of their early functional development, when vestibular terminals emerge at embryonic days 10 and 13 (E10 and E13). At E10, 60% of principal cells generated spikes on depolarization, whereas 50% exhibited excitatory postsynaptic currents (EPSCs) on vestibular-nerve stimulation. The frequency was 0.2 Hz for glutamatergic spontaneous EPSCs (sEPSCs) at -60 mV, and 0.6 Hz for spontaneous inhibitory postsynaptic current (sIPSC) at +10 mV and completely GABAergic. All of these synaptic events were TTX-insensitive, miniature events. At E13, 50% of principal cells generated spikes on depolarization and 82% exhibited EPSCs on vestibular-nerve stimulation. The frequency was 0.7 Hz for sEPSCs at -60 mV, and 0.8 Hz for sIPSCs at +10 mV. Most principal cells had sIPSCs composed of both GABAergic (75%) and glycinergic (25%) events, but a few cells had only GABAergic sIPSCs. TTX decreased the frequency of EPSCs by 12%, and the IPSCs by 17%. In summary, at E10, some principal cells generated immature spikes on depolarization and EPSCs on vestibular-nerve stimulation. At E10, GABAergic events predominated, AMPA events had low frequencies, and glycinergic activity was absent. By E13, glycinergic events first appeared. This data were compared systematically to that obtained from the late-term embryo and hatchling to reveal the long-term sequence of changes in synaptic events and excitability and offer a broader understanding of how the vestibular system is assembled during development.

Maturation of firing pattern in chick vestibular nucleus neurons.

The principal cells of the chick tangential nucleus are vestibular nucleus neurons participating in the vestibuloocular and vestibulocollic reflexes. In birds and mammals, spontaneous and stimulus-evoked firing of action potentials is essential for vestibular nucleus neurons to generate mature vestibular reflex activity. The emergence of spike-firing pattern and the underlying ion channels were studied in morphologically-identified principal cells using whole-cell patch-clamp recordings from brain slices of late-term embryos (embryonic day 16) and hatchling chickens (hatching day 1 and hatching day 5). Spontaneous spike activity emerged around the perinatal period, since at embryonic day 16 none of the principal cells generated spontaneous action potentials. However, at hatching day 1, 50% of the cells fired spontaneously (range, 3 to 32 spikes/s), which depended on synaptic transmission in most cells. By hatching day 5, 80% of the principal cells could fire action potentials spontaneously (range, 5 to 80 spikes/s), and this activity was independent of synaptic transmission and showed faster kinetics than at hatching day 1. Repetitive firing in response to depolarizing pulses appeared in the principal cells starting around embryonic day 16, when <20% of the neurons fired repetitively. However, almost 90% of the principal cells exhibited repetitive firing on depolarization at hatching day 1, and 100% by hatching day 5. From embryonic day 16 to hatching day 5, the gain for evoked spike firing increased almost 10-fold. At hatching day 5, a persistent sodium channel was essential for the generation of spontaneous spike activity, while a small conductance, calcium-dependent potassium current modulated both the spontaneous and evoked spike firing activity. Altogether, these in vitro studies showed that during the perinatal period, the principal cells switched from displaying no spontaneous spike activity at resting membrane potential and generating one spike on depolarization to the tonic firing of spontaneous and evoked action potentials.

Intrinsic membrane properties of vertebrate vestibular neurons: function, development and plasticity.

Central vestibular neurons play an important role in the processing of body motion-related multisensory signals and their transformation into motor commands for gaze and posture control. Over recent years, medial vestibular nucleus (MVN) neurons and to a lesser extent other vestibular neurons have been extensively studied in vivo and in vitro, in a range of species. These studies have begun to reveal how their intrinsic electrophysiological properties may relate to their response patterns, discharge dynamics and computational capabilities. In vitro studies indicate that MVN neurons are of two major subtypes (A and B), which differ in their spike shape and after-hyperpolarizations. This reflects differences in particular K(+) conductances present in the two subtypes, which also affect their response dynamics with type A cells having relatively low-frequency dynamics (resembling "tonic" MVN cells in vivo) and type B cells having relatively high-frequency dynamics (resembling "kinetic" cells in vivo). The presence of more than one functional subtype of vestibular neuron seems to be a ubiquitous feature since vestibular neurons in the chick and frog also subdivide into populations with different, analogous electrophysiological properties. The ratio of type A to type B neurons appears to be plastic, and may be determined by the signal processing requirements of the vestibular system, which are species-variant. The membrane properties and discharge pattern of type A and type B MVN neurons develop largely post-natally, through the expression of the underlying ion channel conductances. The membrane properties of MVN neurons show rapid and long-lasting plastic changes after deafferentation (unilateral labyrinthectomy), which may serve to maintain their level of activity and excitability after the loss of afferent inputs.

Role of Phox2b and Mash1 in the generation of the vestibular efferent nucleus.

The inner ear (vestibular and cochlear) efferent neurons are a group of atypical motor-like hindbrain neurons which innervate inner ear hair cells and their sensory afferents. They are born in the fourth rhombomere, in close association with facial branchial motor neurons, from which they subsequently part through a specific migration route. Here, we demonstrate that the inner ear efferents depend on Phox2b for their differentiation, behaving in that respect like hindbrain visceral and branchial motor neurons. We also show that the vestibular efferent nucleus is no longer present at its usual site in mice inactivated for the bHLH transcription factor Mash 1. The concomitant appearance of an ectopic branchial-like nucleus at the location where both inner ear efferents and facial branchial motor neurons are born suggests that Mash1 is required for the migration of a subpopulation of rhombomere 4-derived efferents.

Differential expression of connexin 43 in the chick tangential vestibular nucleus.

The chick tangential nucleus is a major vestibular nucleus whose principal cells receive convergent inputs from primary vestibular and nonvestibular fibers and participate in the vestibular reflexes. During development, the principal cells gradually acquire the mature firing pattern in part by losing a specific potassium current around hatching (H). Here we focus on characterizing the expression of connexin 43 (Cx43), a gap junction protein found mainly between astrocytes in the mature brain. The astrocytic syncytium plays an important role in maintaining extracellular potassium ion balance in the brain. Accordingly, it is important to characterize the potential of this syncytium to communicate during the critical developmental age of hatching. Using fluorescence immunocytochemistry, we investigated whether Cx43 staining was concentrated in specific cellular compartments at H1 by applying well-known markers for astrocytes (glial fibrillary acidic protein; GFAP), oligodendrocytes (antimyelin), neurons (microtubule-associated protein 2), and synaptic terminals (synaptotagmin). GFAP-positive astrocytes and GFAP-negative nonneuronal cells around the principal cell bodies were labeled with Cx43, suggesting that Cx43 was expressed exclusively by nonneuronal cells near the neuronal elements. Next, the developmental pattern of expression of Cx43 was studied at embryonic day 16 (E16), H1, and H9. At E16, Cx43 was present weakly as random small clusters in the tangential nucleus, whereas, at H1, overall staining became localized, with increases in size, brightness, and number of immunostained clusters. Finally, at H9, Cx43 staining decreased, but cluster size and location remained unchanged. These results suggest that Cx43 is developmentally regulated with a peak at birth and is associated primarily with astrocytes and nonneuronal cells near the principal cell bodies.

[Segmentary organisation of the efferents of the vestibular complex in chicken embryos: is this an example of the general case?]. / Organización segmentaria de las eferencias del complejo vestibular en el embrión de pollo: ejemplo del caso general?

INTRODUCTION: In this paper we present, from the perspective of the embryological segmentary layout of the hindbrain, the topographic layout of the vestibular projection neurons that sustain the vestibulospinal, vestibulo ocular and vestibulocerebellous efferents, in correlation with the classic vestibular nuclei. AIMS: Four vestibular nuclei are usually described superior, lateral, medial and inferior. These originate in at least nine successive rhombomeric segments or pseudosegments, which suggests the possibility of a more precise analysis of their neuronal populations and of their respective connections and functions. It has recently been observed that the vestibular projection neurons identified for a particular target tend to appear aggregated in discrete accumulations, which have been proved to correlate either with rhombomeric units, where they apparently develop, or with internal subdivisions within them. Each projection has its own particular organisation. Comparing them with the resulting connective mosaic in different species shows that various aspects of this organisation are conserved throughout evolution in vertebrates. It is argued that certain genes that control the development of the rhombomeric units in the brain stem may determine, among other aspects, the specific properties of the different neuronal subpopulations related with their axonal navigation and synaptogenesis. CONCLUSIONS: This type of analysis furthers our understanding of how the functional circuitry of a complex system, such as the vestibular system, is generated and is a line of reasoning that in principle can be applied to the whole neural tube.

Development of sensory systems in zebrafish (Danio rerio).

Zebrafish possess all of the classic sensory modalities: taste, tactile, smell, balance, vision, and hearing. For each sensory system, this article provides a brief overview of the system in the adult zebrafish followed by a more detailed overview of the development of the system. By far the majority of studies performed in each of the sensory systems of the zebrafish have involved some aspect of molecular biology or genetics. Although molecular biology and genetics are not major foci of the paper, brief discussions of some of the mutant strains of zebrafish that have developmental defects in each specific sensory system are included. The development of the sensory systems is only a small sampling of the work being done using zebrafish and provides a mere glimpse of the potential of this model for the study of vertebrate development, physiology, and human disease.

Development of the gravity sensing system.

The utricle and saccule contain hair cells, which are the peripheral sensors of change in gravity that transmit signals regarding these changes to the neural components of the vestibular system. Although the fundamental neural pathways, especially the vestibular reflex pathways, have been investigated extensively, the principals underlying the functional development of this system are under study at present. The objective of this review is to identify the gravity-sensing components of the vestibular system and to present an overview of the research performed on their development. The second part of this review is focused on one important aspect of development, the emergence of electrical excitability using the chick tangential vestibular nucleus as a model. The importance of this research to understanding vestibular compensation and vestibular disturbance during spaceflight is considered. Because there is a conservation of the fundamental pathways and function in vertebrate phylogeny from birds through mammals, findings from studies on avians should contribute significantly to understanding the mechanisms operating in mammals. Also, we expect that as the events and basic mechanisms underlying normal vestibular development are revealed, these will provide practical tools to investigate the pattern of recovery from dysfunction of the vestibular system. This is related to the evidence suggesting that recovery of function in different systems and cell lines, including neurons, involves repeating certain patterns established during development.

Rostrocaudal nuclear relationships in the avian medulla oblongata: a fate map with quail chick chimeras.

We present a correlative fate map of the nonsegmented caudal hindbrain down to the medullospinal boundary (medulla oblongata), as a companion to a previous fate mapping study of the hindbrain rhombomeres r2-r6 in quail chick chimeras at stages HH10/11 [Marín and Puelles (1995) Eur J Neurosci 7:1714-1738]. For reproducibility and equivalent precision of analysis, successive portions of the medulla-called pseudorhombomeres "r7" to "r11"-were delimited by transverse planes through the center of adjacent somites at stages HH10/11. These units were each grafted homotopically and isochronically from quail donors into chick hosts. The chimeric specimens were fixed at stages HH35/36 and alternate Nissl-stained sagittal sections were compared to adjacent sections in which quail cells were detected immunocytochemically. This analysis in general showed that there is little intermixing between adjacent pseudorhombomeric domains, although some neuronal populations in the vestibular and trigeminal columns, as well as in the reticular formation and pontine nuclei, do migrate selectively into the host hindbrain. Contralateral migration was scarce up to the stages examined. Several motor nuclei, i.e., the vagal motor complex, or sensory nuclei, i.e., the medial vestibular nucleus, show cytoarchitectonic limits that coincide with pseudorhombomeric ones; however, most conventional grisea were found to originate across several pseudorhombomeres. The inferior olivary complex originated between "r8" and "r11" (between the centers of somites 1 and 5). The medullospinal boundary coincided precisely with the center of the fifth somite, slightly caudal to the obex and the end of the choroidal roof, and correlated with the end of many medullary cytoarchitectonic units. In contrast, the dorsal column nuclei and the caudal subnucleus of the descending trigeminal column fell within the spinal cord. On the whole, the patterns observed were very similar to those found before within the overtly segmented part of the hindbrain, suggesting that some underlying common mechanism may account for the transverse cytoarchitectonic boundaries.

A 5' segment of the mouse Zic1 gene contains a region specific enhancer for dorsal hindbrain and spinal cord.

Zic1 encodes a zinc finger protein, which is required for the development of the dorsal neural tissue. The gene is a mammalian homologue of the Drosophila odd-paired. We examined the regulatory elements in the 5' flanking region of the Zic1 gene as an initial step to understanding how the Zic1 expression is restricted to the dorsal neural tissue. When a 2.9-kb fragment of the 5' flanking segment of the mouse Zic1 gene was linked to the E. coli beta-galactosidase gene, the enzyme was consistently expressed in the dorsal half of the embryonic spinal cord and in the vestibulocochlear nucleus in all four transgenic mouse lines. The transgene expression mimics the Zic1 expression with respect to the region where it occurs. But this is not so for the neuronal cell types. This suggests that the segment contains a region-specific enhancer. In vivo and in vitro deletion analyses indicated that there are essential regions between -2.0 and -0.9 kb and within the proximal 0.9 kb. The distal element is necessary for the transgene expression in the embryonic dorsal spinal cord whereas the adult vestibulocochlear nucleus expression is regulated by both elements. In these regions, there are sequences similar to the binding sequences for potential regulatory proteins.

Developing vestibular ganglion neurons switch trophic sensitivity from BDNF to GDNF after target innervation.

Recent evidence showing a distinctive cell loss in vestibular and cochlear ganglia of brain-derived neurotrophic factor (BDNF) versus neurotrophin-3 (NT-3) null mutant mice demonstrates that these neurotrophins play a critical role in inner ear development. In this study, biological functions of BDNF and NT-3 in the chick vestibular and cochlear ganglion development was assessed in vitro and compared to those of other neurotrophic factors. The embryonic day (E)8-12 vestibular ganglion neurons showed an extensive outgrowth in response to BDNF with less outgrowth to NT-3. In contrast, NT-3 had stronger neurotrophic effects on the E12 cochlear ganglion neurons compared to BDNF. These results support previous evidence that neurotrophins play important roles in the vestibular and cochlear ganglion neuron development. However, the responsiveness to the neurotrophins declined and became undetectable by E16. Unexpectedly, glial cell line-derived neurotrophic factor (GDNF) promoted neurite outgrowth from vestibular ganglia at E12-16, later than the stages at which BDNF had neurotrophic effects. The time of switching sensitivity of the vestibular ganglion neurons from BDNF to GDNF correlated with the time of completion of synaptogenesis on their peripheral and central targets. Furthermore, a factor released from E12 inner ears exerted neurotrophic effects on late-stage vestibular ganglion neurons that were not responsive to the E4 otocyst-derived factor. These results raise the possibility that the vestibular ganglion neurons become responsive to GDNF upon target innervation and that the changes in sensitivity are regulated by changes in available factors released from their peripheral targets, the inner ear epithelia.

Lack of biocytin transfer at gap junctions in the chicken vestibular nuclei.

In vivo experiments were designed to test for functional gap junctions at 'mixed' synapses that were morphologically characterized between the large-diameter, primary vestibular fibers and second-order vestibular neurons in the chicken, Gallus gallus. In previous intracellular recordings and dye injections into these neurons from brain slice preparations of chick embryos (E15/16) and also newborn hatchlings (HI-2), no evidence was obtained for functional gap junctions. Therefore, biocytin, a low molecular weight tracer that permeates gap junction channels, was extracellularly applied to either the ampullary nerves or to the vestibular ganglion of 3-6 day old hatchlings and adult chickens (9 months). This procedure resulted in the uptake of the dye and heavy staining of both the thick and thin fibers composing the vestibular nerve and in loading of vestibular efferent neurons. However, no dye transfer was observed between the large-diameter, primary vestibular fibers and second-order vestibular neurons. This observation, which was performed using a relatively non-invasive approach on intact animals, suggests that the gap junctions at these mixed synapses are probably not functional under the conditions of these experiments.

Development of the human lateral vestibular nucleus: a morphometric evaluation.

The development of the human lateral vestibular nucleus was studied on serial sections of the brain of 8 fetuses and neonates at 12-40 weeks of gestation, an infant at 2 months of age and an adult of 63 years using a microscope with a drawing tube and an image-analysing computer system. A morphometric analysis revealed that the lateral vestibular nucleus, whose neurons were distinguished from glia after 16 weeks of gestation, divided cytoarchitectonically into the medial and the lateral subnuclei at 21 weeks of gestation onwards, and showed the moderate development in terms of the columnar length and volume, neuronal size and neuropil.

[Development and cell differentiation of the vestibulocochlear nuclei in cattle]. / Entwicklung und Zelldifferenzierung der Nuclei vestibulocochleares beim Rind.

Based upon light- and electron microscopic examinations (50 embryos ranging from 1 to 53 cm crown-rump-length, CRL) the origin of the nuclei of the cranial nerve VIII is described with special regard to neurogenesis. The ventricular matrix lateral to the sulcus limitans represents the alar plate with its sensory areas. Up to 2.7 cm CRL migrating neurons from the vestibular nuclei can be detected, the bigger neuronal elements of which are the early formed lateral vestibular nucleus. From 6.7 cm CRL onward all nuclear groups of the vestibular nerve can be identified. At 1 cm CRL the recess plate represents the primordium of the cochlear nuclear complex. Identification of the definitive nuclei is possible at 3.8 cm CRL. Subsequently from 7.6 cm CRL onward the process of lamination can be observed in the dorsal cochlear nucleus. Due to the proceeding maturation the nuclei of the cranial nerve VIII correspond at 53 cm CRL topographically and cytologically to the characteristics of adult animals. Electron microscopic examinations are documenting characteristic features of cytogenesis of sensory neurons (vestibular nucleus) during early embryonic stages (2.5 cm and 3.6 cm CRL). At 2.5 cm CRL the elements exhibiting features of migrating neurons are predominating, whereas at 3.6 cm CRL an increased differentiation is typical for neurons localized in their ultimate position. At this stage synapses can be identified for the first time.