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The morphology of the trigeminal ganglion in human fetus was investigated by means of the tract-tracing method using the lipophilic dye DiI-C18-(3) (1,1'-double octadecane 3,3,3'3'-tetramethyl indole carbonyl cyanine-perchlorate), hematoxylin-eosin (HE) stain, and three-dimensional computer reconstruction models. The trigeminal ganglion was flat in the dorsoventral direction, and DiI staining revealed that the trigeminal ganglion cells were somatotopically distributed in the ganglion in a way that reflected the mediolateral order of the three branches. Ganglion cells of the ophthalmic nerve were distributed in the anteromedial part of the trigeminal ganglion, those of the mandibular nerve were in the posterolateral part, and those of the maxillary nerve were localized in the intermediate part. DiI labeled both ganglion cells and nerve fibers in the trigeminal ganglion; the ganglion cells varied in size and appeared as round- or oval-shaped, the neurites connected the cell soma, and some bipolar neurons were also observed. The number of embryonic trigeminal ganglion cells did not significantly change with gestational age, but the cell diameter, area, and perimeter significantly increased. The motor root leaves the pons, runs along the sensory root, passes the ventral surface of the ganglion, and finally runs together with the mandibular nerve. The findings reported here elucidate the morphology, development, and somatotopic organization of the trigeminal ganglion and reveal the trigeminal nerve motor root pathway along the trigeminal ganglion and mandibular nerve in the human fetus.

The development of facial nuclei in animal models of disease is poorly understood, but autism is sometimes associated with facial palsy. In the present study, to investigate migration of facial neurons and initial facial nucleus formation in an animal model of autism, rat embryos were treated with valproic acid (VPA) in utero at embryonic day (E) 9.5 and their facial nuclei were analyzed by in situ hybridization at E13.5, E14.5 and E15.5. Signals for Tbx20, which is expressed in early motor neurons, appeared near the floor plate at the level of the vestibular ganglion and extended caudolaterally, where they became ovoid in shape. This pattern of development was similar between control and VPA-exposed embryos. However, measurements of the migratory pathway and the size of the facial nuclei revealed that exposure to VPA hindered the caudal migration of neurons to the facial nuclei. Signals for cadherin 8, which is expressed in mature facial nuclei, revealed that exposure to VPA caused a significant reduction in the size of the facial nuclei. Our findings provide the first quantitative description of tangential migration and nucleus formation in the developing hindbrain in a rat model of autism.

Background and Objective The optical stimulation of neurons from pulsed infrared lasers has appeared over the last years as an alternative to classical electric stimulations based on conventional electrodes. Laser stimulation could provide a better spatial selectivity allowing single-cell stimulation without prerequisite contact. In this work we present relevant physical characteristics of a non-lethal stimulation of cultured mouse vestibular and retinal ganglion neurons by single infrared laser pulses. Study Design/Materials and Methods Vestibular and retinal ganglion neurons were stimulated by a 100–400 mW pulsed laser diode beam (wavelengths at 1,470, 1,535, 1,875 nm) launched into a multimode optical fiber positioned at a few hundred micrometers away from the neurons. Ionic exchange measurements at the neuron membrane were achieved by whole-cell patch-clamp recordings. Stimulation and damage thresholds, duration and repetition rate of stimulation and temperature were investigated. Results All three lasers induced safe and reproducible action potentials (APs) on both types of neurons. The radiant exposure thresholds required to elicit APs range from 15 ± 5 to 100 ± 5 J cm−2 depending on the laser power and on the pulse duration. The damage thresholds, observed by a vital dye, were significantly greater than the stimulation thresholds. In the pulse duration range of our study (2–30 milliseconds), similar effects were observed for the three lasers. Measurements of the local temperature of the neuron area show that radiant exposures required for reliable stimulations at various pulse durations or laser powers correspond to a temperature increase from 22°C (room temperature) to 55–60°C. Stimulations by laser pulses at repetition rate of 1, 2, and 10 Hz during 10 minutes confirmed that the neurons were not damaged and were able to survive such temperatures. Conclusion These results show that infrared laser radiations provide a possible way to safely stimulate retinal and vestibular ganglion neurons. A similar temperature threshold is required to trigger neurons independently of variable energy thresholds, suggesting that an absolute temperature is required. Lasers Surg. Med. 44: 736–745, 2012. © 2012 Wiley Periodicals, Inc.

Objectives/Hypothesis: Vestibular neuritis is a common cause of both acute and chronic vestibular dysfunction. Multiple pathologies have been hypothesized to be the causative agent of vestibular neuritis; however, whether herpes simplex type I (HSV1) reactivation occurs within the vestibular ganglion has not been demonstrated previously by experimental evidence. We developed an in vitro system to study HSV1 infection of vestibular ganglion neurons (VGNs) using a cell culture model system. Study Design: basic science study. Results: Lytic infection of cultured rat VGNs was observed following low viral multiplicity of infection (MOI). Inclusion of acyclovir suppressed lytic replication and allowed latency to be established. Upon removal of acyclovir, latent infection was confirmed with reverse-transcription polymerase chain reaction and by RNA fluorescent in situ hybridization for the latencyassociated transcript (LAT). A total of 29% cells in latently infected cultures were LAT positive. The lytic ICP27 transcript was not detected by reverse-transcription polymerase chain reaction (RT-PCR). Reactivation of HSV1 occurred at a high frequency in latently infected cultures following treatment with trichostatin A (TSA), a histone deactylase inhibitor. Conclusions: VGNs can be both lytically and latently infected with HSV1. Furthermore, latently infected VGNs can be induced to reactivate using TSA. This demonstrates that reactivation of latent HSV1 infection in the vestibular ganglion can occur in a cell culture model, and suggests that reactivation of HSV1 infection a plausible etiologic mechanism of vestibular neuritis.

The cochleo-vestibular ganglion contains neural crest-derived glial cells and sensory neurons that are derived from the neurogenic otic placode. Little is known about the molecular mechanisms that regulate the tightly orchestrated development of this structure. Here, we report that Sox10, a high-mobility group DNA-binding domain transcription factor that is required for the proper development of neural crest cell derivatives, is specifically expressed in post-migratory neural crest cells in the cochleo-vestibular ganglion. Using Sox10-deficient mice, we demonstrate that this transcription factor is essential for the survival, but not the generation, of the post-migratory neural crest cells within the inner ear. In the absence of these neural crest-derived cells, we have investigated the survival of the otocyst-derived auditory neurons. Surprisingly, auditory neuron differentiation, sensory target innervation and survival are conserved despite the absence of glial cells. Moreover, brain-derived neurotrophic factor expression is increased in the hair cells of Sox10-deficient mice, a compensatory mechanism that may prevent spiral ganglion neuronal cell death. Taken together, these data suggest that in the absence of neural crest-derived glial cells, an increase trophic support from hair cells promotes the survival of spiral ganglion neurons in Sox10 mutant mice.

The espins are Ca(2+)-resistant actin-bundling proteins that are enriched in hair cell stereocilia and sensory cell microvilli. Here, we report a novel localization of espins to a large proportion of rat type I spiral ganglion neurons (SGNs) and their projections to the cochlear nucleus (CN). Moreover, we show that a fraction of these espins is in the nucleus of SGNs owing to the presence of splice-isoforms that contain a functional nuclear localization signal (NLS). Espin antibody labeled approximately 83% of type I SGNs, and the labeling intensity increased dramatically during early postnatal development. Type II SGNs and vestibular ganglion neurons were unlabeled. In the CN, espin-positive auditory nerve fibers showed a projection pattern typical of type I SGNs, with intense labeling in the nerve root region and posteroventral CN (PVCN). The anteroventral CN (AVCN) showed moderate labeling, whereas the dorsal CN showed weak labeling that was restricted to the deep layer. Espin-positive synaptic terminals were enriched around nerve root neurons and octopus cells in the PVCN and were also found on globular bushy cells and multipolar neurons in the PVCN and AVCN. SGNs expressed multiple espin transcripts and proteins, including splice-isoforms that contain a nonapeptide, which is rich in positively charged amino acids and creates a bipartite NLS. The nonapeptide was necessary to target espin isoforms to the nucleus and was sufficient to target an unrelated protein to the nucleus when joined with the upstream di-arginine-containing octapeptide. The presence of cytoplasmic and nuclear espins in SGNs suggests additional roles for espins in auditory neuroscience.

Bipolar neurons (BNs) in the vestibular ganglion (VG) connect vestibular hair cells with the central nervous system (CNS). Disturbed function and cell loss in central vestibular target areas or in the vestibular periphery involve BNs either retro- or anterogradely. However, the impact of central vestibular disturbances or hair cell defects on the maintenance of BNs is poorly understood. In the present study the volume of the VG, the size and total number of BNs, and the number of BNs expressing the calcium-binding protein calretinin (Calr) were quantified stereologically in the cerebellar mutants purkinje cell degeneration (pcd/pcd), weaver (wv/wv), and Lurcher (Lc/+), and in the vestibular mutants jerker (je/je), shaker-1 (sh/sh), and Varitint-waddler (Va/+). In all the different mutant mice investigated the total number of BNs did not differ from that of wildtypes. In contrast, the number of Calr-positive BNs was significantly reduced in je/je (23%) and sh/sh (33%) mutants. Reduced cell size was apparent in sh/sh mutants and the volume of the VG significantly decreased in je/je mice. Calr was virtually absent from calyx endings in the vestibular periphery of je/je, sh/sh, and Va/+ mutants, whereas in wildtypes and cerebellar mutants many calyces displayed intense Calr labeling. These results imply that the survival of BNs is apparently unaffected by the peripheral and central target defects found in the mutants investigated. Whether the decrease in Calr expression may reflect biochemical adaptations in response to input disturbances or a specific loss of large BNs is discussed.

Deterioration of balance with advancing age is a well-known fact of life. Some investigators have reported a 50% prevalence of dizziness in the elderly. Clinically, progressive dysequilibrium of aging presents as gradually worsening balance due to age-related decline in function of the peripheral vestibular system, central nervous system, vision, and musculoskeletal system. Vestibular function testing has shown clear evidence of age-related changes in peripheral and central sites. Histopathologic changes in the vestibular sensory organs include progressive hair cell degeneration, otoconial degeneration in the otolith organs, and decreasing number of Scarpa's ganglion neurons. Recently, a new quantitative method of assessing vestibular otopathology has been described, utilizing Nomarski differential interference contrast microscopy. This technique has been applied to 67 human temporal bones of individuals from birth to age 100 to create a normative database of total, type I, and type II hair cell counts as a function of age. Results show a highly significant continuous decrease in all counts from birth to age 100, best fit by a linear regression model. Type I hair cell counts in all three semicircular canal cristae decrease at a similar rate, significantly faster than the degeneration observed in type I hair cells of the maculae. Type II hair cell counts decline at the same rate for all 5 sensory epithelia. These normative data provide the basis for comparisons to hair cell counts made in temporal bones from subjects with known vestibular disorders. They also provide a basis for drawing correlations between vestibular function testing and vestibular otopathology.

The acquisition of ion channels is critical to the formation of neuronal pathways in the peripheral and central nervous systems. This study describes the different types of inward currents (I i ) recorded from the soma of isolated cochleovestibular ganglion (CVG) cells of the embryonic chicken, Gallus gallus. Cells were isolated for whole-cell tight-seal recording from embryonic day (ED) 3, an age when the CVG is a cell cluster, to ED 9, an age when the cochlear and vestibular ganglia (CG, VG) are distinct structures. Results show Na + and Ca + currents (I Na and I Ca ) are acquired by ED 3, although / Na dominates with greater density levels that peak by ED 6-7 in VG neurons. In the CG, / Na acquisition is slower, reaching peak values by ED 8-9. Isolation of I Ca , using Ba 2+ as the charge carrier, showed both transient (I BaT )- and sustained (I BaL )-type currents on ED 3. Unlike I Na , I Ba density varied with age and ganglion. Total I Ba increased steadily, showing a decline only in CG cells on ED 8-9 as a result of a decrease in I BaT . I BaL density increased over time, reaching a maximum on ED 6-7 in VG cells, followed by a decline on ED 8-9. In comparison, I BaL in CG neurons, did not increase significantly beyond mean values measured on ED 5. The early onset of these currents and the variations in Ca 2+ channel expression between the ganglia suggests that intracellular signals relevant to phenotypic differentiation begin within these early time frames.

Vestibular tissues (cristae ampullares, macular otolithic organs, and Scarpa's ganglia) in chinchilla, rat, and guinea pig were examined for immunoreactivity to the alpha9 nicotinic acetylcholine receptor (nAChR) subunit. The alpha9 antibody was generated against a conserved peptide present in the intracellular loop of the predicted protein sequence of the guinea pig alpha9 nAChR subunit. In the vestibular periphery, staining was observed in calyces around type I hair cells, at the synaptic pole of type II hair cells, and in varying levels in Scarpa's ganglion cells. Ganglion cells were also triply labeled to detect alpha9, calretinin, and peripherin. Calretinin labels calyx-only afferents. Peripherin labels bouton-only afferents. Dimorphic afferents, which have both calyx and bouton endings, are not labeled by calretinin or peripherin. In these experiments, alpha9 was expressed in both calyx and dimorphic afferents. A subpopulation of small ganglion cells did not contain the alpha9 nAChR but did stain for peripherin. We surmise that these are bouton-only afferents. Bouton (regularly discharging) afferents also show efferent responses, although they are qualitatively different from those in irregularly discharging (calyx and dimorphic) afferents, much slower and longer lasting. Thus, regular afferents are probably more affected via a muscarinic cholinergic or a peptidergic mechanism, with a much smaller superimposed fast nicotinic-type response. This latter response could be due to one of the other nicotinic receptors that have been described in studies from other laboratories.

The metabolism of glutamate, the most likely neurotransmitter of vestibular ganglion cells, includes synthesis from glutamine by the enzyme glutaminase. We used microdissection combined with a fluorometric assay to measure glutaminase activity in the vestibular nerve root and nuclei of rats with unilateral vestibular ganglionectomy. Glutaminase activity in the lesioned-side vestibular nerve root decreased by 62% at 4 days after ganglionectomy and remained at similar values through 30 days. No change occurred in the contralateral vestibular nerve root. Glutaminase activity changes in the vestibular nuclei were lesser in magnitude and more complex, including contralateral increases as well as ipsilateral decreases. At 4 days after ganglionectomy, glutaminase activity was 10-20% lower in individual lesioned-side nuclei compared with their contralateral counterparts. By 14 and 30 days after ganglionectomy, there were no statistically significant differences between the nuclei on the two sides. This transient asymmetry of glutaminase activities in the vestibular nuclei contrasts with the sustained asymmetry in the vestibular nerve root and suggests that intrinsic, commissural, or descending pathways are involved in the recovery of chemical symmetry. This recovery resembles our previous finding for glutamate concentrations in the vestibular nuclei and may partially underlie central vestibular compensation after peripheral lesions.

Objective/Hypothesis This study was designed to investigate the hypothesis that Meniere's disease is associated with herpes simplex virus (HSV) reactivation in the vestibular ganglion. Study Design Case control study. Methods Vestibular ganglia were obtained from archival surgical pathology specimens from patients undergoing vestibular neurectomy for vertigo caused by Meniere's disease. All patients met criteria for classification as definite Meniere's disease according to American Academy of Otolaryngology/Head and Neck Surgery (AAO-HNS) criteria. Control specimens were obtained from willed body donors. Sections from each ganglion were studied for prevalence of viral DNA using a nested polymerase chain reaction designed to amplify the HSV DNA polymerase gene. Quantitative analysis determined the number of viral copies per standard unit of ganglionic DNA. Results HSV DNA was more prevalent in paraffin embedded ganglia from patients with Meniere's disease (100%) than in fresh-frozen control ganglia (81%) (P = .02). Fixation and paraffin embedding substantially reduced recovery of HSV virus in selected control specimens. Quantitative analysis found no correlation between viral copy number in control ganglia processed frozen versus formalin fixed and paraffin embedded. Conclusions HSV is more commonly isolated from vestibular ganglia of patients with Meniere's disease than the general population. The routine histologic preparation of formalin fixation and paraffin embedding significantly altered the quantity of virus detected though not in a predictable manner. The study provides supportive evidence for a viral etiology in Meniere's disease.

In order to investigate the mechanisms responsible for adaptation to altered gravity, we assessed the changes in mRNA expression of glutamate receptors in vestibular ganglion cells, medial vestibular nucleus, spinal vestibular nucleus/lateral vestibular nucleus, cerebellar flocculus, and uvula/nodulus from rats exposed to hypergravity for 2 h to 1 week using real-time quantitative RT-PCR methods. The mRNA expression of GluR2 and NR1 receptors in the uvula/nodulus and NR1 receptors in the medial vestibular nucleus increased in animals exposed to 2 h of hypergravity, and it decreased gradually to the control level. The mRNA expression of GluR2 receptors in vestibular ganglion cells decreased in animals exposed to 1 week of hypergravity. Neither the metabotropic glutamate receptor 1 nor delta2 glutamate receptor in flocculus and uvula/nodulus was affected by a hypergravity load for 2 h to 1 week. It is suggested that the animals adapted to the hypergravity by enhancing the cerebellar inhibition of the vestibular nucleus neurons through activation of the NR1 and GluR2 receptors on the Purkinje cells in uvula/nodulus especially at the early phase following hypergravity. In the later phase following hypergravity, the animals adapted to the hypergravity by reducing the neurotransmission between the vestibular hair cells and the primary vestibular neurons via down-regulation of the postsynaptic GluR2 receptors in the vestibular periphery.

Dysequilibrium of aging in humans has been speculated to arise from progressive deterioration within anatomical components of the vestibular system. An integral part of this system is vestibular ganglions, which are bipolar neurons that relay peripheral vestibular information to the central nervous system. To assess the effect of aging on the number of human vestibular ganglion neurons, assumption-free stereology in the form of the optical fractionator was used on 20 serially sectioned archival human temporal bone specimens. Donors had no history of vestibular pathology and ranged in age from 2 to 88 years. An average of 25,812 (coefficient of variation = 0.13) vestibular ganglion neurons was found throughout this age range, a significant departure from the results of past studies. Logistics-based regression analysis pointed to a nonlinear pattern of decline in the neuronal population: the number of cells remained roughly constant at about 28,952 cells in youth and then declined gradually between 30 and 60 years of age before leveling off at approximately 23,349 cells in older individuals. This study confirmed the existence of an age-related decline in the primary neurons of the human vestibular system, thus providing one anatomical basis for the increased incidence of imbalance seen with age. J. Comp. Neurol. 431:437–443, 2001. © 2001 Wiley-Liss, Inc.

Eph receptors and their ligands, termed ephrins, have been implicated in axon guidance, neuron-target interactions, regional compartmentalization, and synaptic functions in nervous systems. These activities of the Eph family molecules prompted us to investigate whether these molecules play roles in the maintenance, regeneration and plasticity in the mature peripheral vestibular system. Using reverse transcription-polymerase chain reaction (RT-PCR), Western blot and immunocytochemical analyses, we identified distinct and reciprocal expression patterns of full-length isoforms of EphA5, EphA6, EphA7, EphB1, ephrin-A2 and ephrin-B1 that correlated with structural features of the peripheral vestibular system in adult rats. All of the Eph receptors and ephrins examined were localized in the cell bodies of vestibular ganglion neurons in vivo and in vitro, and were readily detected in their outgrowing neurites in vitro. In the utricle, these molecules were localized in distinct cellular and subcellular compartments corresponding to discrete features of utricular afferent innervation, e.g. defasciculation, branching and synapse formation. Taken together, these results identify the Eph receptors and ephrins as candidate molecular substrates for defining some aspects of the structural organization of the adult peripheral vestibular system.

Vestibular neuritis is a common cause of partial unilateral vestibular paralysis, which usually spares posterior semicircular canal function. The cause is assumed to be a viral reactivation of latent herpes simplex virus type 1 (HSV-1) in human vestibular ganglia. The existence of an anastomosis between the intermediate nerve and the superior vestibular nerve suggests the question of whether selective affliction of the superior vestibular nerve is the result of migration of HSV-1 from the geniculate ganglion along this faciovestibular anastomosis. We determined the distribution of HSV-1 among geniculate ganglia, vestibular ganglia, and within Scarpa's ganglion by examining 35 human temporal bones by polymerase chain reaction. HSV-1 was found in 66% of geniculate ganglia and 60% of vestibular ganglia; all examined parts of vestibular ganglia were almost equally HSV-1 infected. Our data provided no support for viral migration along this anastomosis or for a preferential latency of HSV-1 in the superior vestibular nerve. We suggest that the common double innervation of the posterior ampulla by two nerves running in two separate bony canals could offer an alternative explanation for the regular sparing of posterior canal function in vestibular neuritis.

The effects of intra-labyrinthine application of kainic acid on afferent vestibular nerve terminals and vestibular ganglion cells were investigated in this study. Results showed 10 nM kainic acid can induce scattered swelling of the calyx nerve endings surrounding type I vestibular hair cells. The swelling became much more severe when kainic acid concentration increased to 50 nM. The calyx swelling is also the earliest pathological change in the vestibular epithelium after a short period of anoxia. Degeneration of vestibular ganglion cells was found 1 week after kainic acid treatment. These results suggested, like in the cochlea, excitotoxicity can be mediated by overactivation of KA receptors. This is also an indirect evidence support L-glutamate as the neurotransmitter used in the peripheral afferent vestibular system.

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. © 1999 John Wiley & Sons, Inc. J Neurobiol 38: 414–427, 1999

The subpopulation of primary vestibular afferents (PVA) displaying immunoreactivity for the calcium binding protein Calbindin D-28k (Calb) is constituted of particularly large bipolar neurons in the vestibular ganglion (VG) that innervate the central regions of the vestibular end organs exclusively via calyx endings on type I vestibular hair cells. These large-diameter PVA are characterized by irregular spontaneous discharge patterns and predominantly phasic firing properties with respect to natural vestibular stimulation. The present study describes the complete course and terminations of Calb+ PVA in the cerebellar cortex, the cerebellar (CN) and vestibular nuclei (VN) of the mouse. To eliminate the two sources of Calb+ fibers in the cerebellum, i.e., the Calb+ primary vestibular input and the axons of cerebellar Purkinje cells (PC), in their totality, a unilateral eighth nerve transection was performed in the PC-deficient mutant mice, Purkinje cell degeneration (pcd/pcd) and Lurcher (Lc/+). Neurectomy in these mutants results in a complete ipsilateral loss of Calb+ fibers in the cerebellar cortex, the CN and VN. The Calb+ primary vestibular input on the contralateral side terminates solely in the rostral half of the ventral uvula and in the nodulus of the cerebellar cortex. Calb+ fibers traverse all three subdivisions of the CN, but terminations were found only in the lateral and medial cerebellar nuclei. In the VN, Calb+ PVA terminations were restricted to the superior, the ventral part of the lateral, the lateral portion of the medial, and the inferior vestibular nuclei. Calb + terminals were also present in the small cell group Y and Cajal's interstitial nucleus of the vestibular nerve as well as in defined areas of the reticular formation. All Calb + PVA are strictly unilateral. The results show that the Calb+ subpopulation of VG neurons is the sole source of Calb+ fibers and terminals in the PC-deficient cerebellum and the VN. The central input of this distinct subgroup of PVA is distributed in narrow posterior vermal areas and parts of the CN and VN. The cerebellar mutants, Purkinje cell degeneration and Lurcher, provide excellent tools to selectively investigate the subgroup of Calb+ PVA in the mouse in its entirety.

On the basis that neurotrophins (NTs) affect neuronal synaptic plasticity, are expressed in various cell types of the vestibular system, and exert a trophic influence on statoacoustic neurons, the authors hypothesized a role for NTs in vestibular compensation. To test this hypothesis, they performed unilateral surgical labyrinthectomy in 11 heterozygous (+/-) neurotrophin 3 (NT3) and brain-derived neurotrophic factor (BDNF) knockout mice and in two neurotrophin 4 (NT4) homozygous (-/-) knockout mice, each with a control (+/+) sibling, for a total of 26 mice. Four BDNF(+/-) and four NT3(+/-) mice with their (+/+) controls each were allowed to recover in a normal lighted room for 3, 7, 14, and 30 days following labyrinthectomy. Two BDNF(+/-) and two NT4(-/-) mice with controls were kept in total darkness for 1- and 16-day survival periods. One NT3(+/-) mouse without a control (which died in surgery) was sacrificed after 16 days in darkness. The behavior of all mice was videorecorded to monitor their recovery. Compared with normal (+/+) littermate controls, NT3(+/-) mice demonstrated a delay in compensation (8 to 10 days) in light surround, whereas NT4(-/-) mice showed only a minor delay in dark surround. Despite a 40% lower vestibular ganglion cell population in BDNF(+/-) mice compared with (+/+) controls, BDNF(+/-) mice did not reveal a detectable delay in recovery following labyrinthectomy. These findings suggest that a 50% loss of NT3 protein significantly affects vestibular recovery in adult mice. Perhaps variations in achieving vestibular compensation in humans may be partly secondary to genetically different NT3 levels in vestibular pathways.

NaG, a member of subfamily 2, was originally characterized as a “glial” sodium channel, and is expressed at high levels in Schwann cells in vivo. However, NaG has also been shown by in situ hybridization to be highly expressed in rat dorsal root ganglion (DRG) neurons, which, like Schwann cells, are neural crest derivatives. In the present study, we used non-isotopic in situ hybridization with a riboprobe for NaG, in conjunction with RT-PCR, to determine whether NaG is expressed in tissues related to the DRG either by neural crest origin or sensory function. We found that the expression of significant levels of NaG mRNA was restricted to derivatives of the neural crest (neurons of DRG, trigeminal ganglion and mesencephalic nucleus of the trigeminal nerve, and Schwann and satellite cells); the absence of NaG from superior cervical ganglion neurons and adrenal medulla chromaffin cells indicates that not all neural crest derived neural elements express NaG. NaG was not observed in sensory neurons (retina, vestibular ganglion, spiral ganglion, olfactory epithelium) that are not of neural crest origin. Our results indicate that NaG mRNA is expressed not only in Schwann cells, but also in a spectrum of neuronal cell types with afferent function of neural crest origin. NaG thus appears to represent a transcript encoding a sodium channel or sodium channel-like protein that is uniquely expressed by both Schwann cells and afferent neurons of neural crest origin. GLIA 21:269–276, 1997. © 1997 Wiley-Liss, Inc.

The effects of the application of brain-derived neurotrophic factor (BDNF) and neurotrophin-3 (NT-3) neurotrophins on the intracellular calcium level ([Ca2+]i) were studied in vestibular ganglion neurons (VGNs) from postnatal day 3 (P3) rats cultured for 50 hr. We first assessed the expression of trkB and trkC mRNA receptors in cultured VGNs. Immunobloting and immunocytochemistry confirmed the presence of the neurotrophin receptors on neurons. Both neurotrophins induced transient [Ca2+]i elevations in VGNs: BDNF-treated neurons responded in 65% and NT-3-treated neurons in 56%. The responses could be inhibited by anti-BDNF or anti-NT-3 antibodies. The [Ca2+]i elevation was dependent on extracellular calcium since it was abolished in calcium-free medium but also implicates the release of calcium from intracellular stores as tested by prior depletion with thapsigargin. Our results suggest the implication of a short-term calcium regulation in VGNs, which could reflect specific fast effects of neurotrophins in the early postnatal rat vestibular system.

The protein kinase C family of enzymes is composed of at least ten different isoforms that display a variety of distinct biochemical specificities. Many of these isoforms are highly expressed in brain, and some show regional specificity in their distribution, suggesting that they may serve specific functions. By using immunocytochemistry to localize the betaI, betaII, gamma, or delta isoforms of protein kinase C in the central vestibular system of the adult rat, we found the vestibular ganglion and its peripheral and central processes of the eighth nerve to be heavily labeled with protein kinase C betaI immunoreactivity. Labeled axons and terminals were also found in all four vestibular nuclei. Some neurons of the vestibular ganglion were weakly stained with the antibody to protein kinase C betaII, as were scattered axons in the eighth nerve, and scattered axons and terminals were found in all four vestibular nuclei among weakly labeled neurons. A few axons in the vestibular portion of the eighth nerve were labeled with protein kinase C gamma immunoreactivity, and neurons of the spinal, lateral, and superior vestibular nuclei were heavily decorated with synapses, presumably derived from Purkinje neurons, which were also strongly immunoreactive. Neurons of the medial vestibular nucleus were not as heavily innervated. With the antibody to protein kinase C delta, we found scattered, weakly immunoreactive neurons in the vestibular portion of the eighth nerve. Myelinated fiber bundles of the spinal vestibular nucleus contained moderate numbers of labeled axons, and the other vestibular nuclei were well innervated by protein kinase C delta axons and terminals. Most of these probably derive from Purkinje cells, which were labeled in longitudinal bands interspersed with bands of labeled basket cells. These data suggest that particular protein kinase C isoforms play specific roles in vestibular and cerebellar function.

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 (H1-2), 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.

The presence and the activity of alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA) glutamate receptors were investigated in mouse cultured vestibular ganglion neurons using immunocytochemistry and measurement of intracellular calcium concentration ([Ca2+]i) by spectrofluorimetry. Cultures of dissociated vestibular ganglia from 18 gestation day mouse embryos were grown in vitro for 3-4 days. Immunocytochemical labelling of AMPA receptor subunits GluR2/R3 and GluR4 was detected in neuron cell bodies and proximal neurites and more lightly in glial cells. There was no clear selective subcellular localization of the different subunits. For the GluR1 subunit a signal was observed only in some neurons and neurites and was weak. Vestibular ganglion neurons responded to fast application of 1 mM glutamate and 10 mM aspartate through unknown receptors by a transient increase in [Ca2+]i. The mean amplitude of this rapid increase was about nine times the resting level and recovery was complete within 30-45 s after the application. If separated by an interval of at least 10 min, consecutive applications produced similar calcium responses. AMPA (1 mM) application induced the same type of responses. Five minutes prior to the AMPA exposure, the application of a specific AMPA antagonist, 6,7-dinitroquinoxaline-2,3-dione (DNQX, 1.5 mM), in the external medium inhibited the response to AMPA. Chelation of external calcium by EGTA (1.5 mM) abolished the responses to drug applications, indicating that an influx of external calcium is involved in the [Ca2+]i increase. These observations suggest that heteromeric AMPA receptors are expressed in vestibular ganglion neurons in culture and play a functional role in their glutamate-induced depolarization. Experiments are in progress using specific AMPA and NMDA antagonists to characterize the participation of the two types of ionotropic glutamate receptors in the glutamate/aspartate-induced intracellular calcium response.

Postmortem examination of the brain and temporal bones of a patient with well-documented vestibular neuritis showed selective neuronal loss in Scarpa's ganglia on the side with absent caloric response. There was loss of hair cells and an “epithelialization” of the utricular macule and semicircular canal cristae on the deafferented side, and synaptic density in the vestibular nuclei on the deafferented side was decreased compared with that on the normal side. All findings were consistent with an isolated viral infection of Scarpa's ganglia. This is the first description of the effects of chronic deafferentation on the vestibular sensory epithelia and the vestibular nuclei in a human being. (Otolaryngol Head Neck Surg 1996;114:586-92.)

Presence and distribution of glutamate, glycine, GABA and beta-alanine in VIIIth nerves of frogs and rats were investigated with postembedding immunocytochemical methods on serial semithin sections. In Scarpa's ganglion of the frog, all cell bodies were glutamate immunoreactive. About 17% of the cells per section were also glycine immunoreactive, but none were GABA or beta-alanine immunoreactive. The mean diameter of glycine-positive cell bodies (26.7 +/- 6.9 microns; N = 130) was significantly (P0.0001) larger than that of glycine-negative cell bodies (15.7 +/- 5.4 microns; N = 272). The intensity of glutamate immunostaining decreased with cell diameter, whereas the intensity of glycine immunostaining increased with cell diameter. As a result, the staining intensities for glutamate and glycine in a given cell were negatively correlated. Glycine immunoreactivity was also present in a size-related manner in distal and proximal afferent fibers. The majority of thin fibers (4 microns) was glycine negative, whereas most of the thick fibers (10 microns) were glycine positive. Glycine-positive fibers were observed in the sensory epithelial of all end organs in the inner ear. The saccular macula and its nerve, however, contained only few glycine immunoreactive structures. In Scarpa's ganglion of the rat, all cells were immunoreactive for glutamate, about 12% for colocalized glycine, and none for GABA or beta-alanine. Glycine-positive cell bodies were significantly (P0.0001) larger (32.2 +/- 5.2 microns; N = 82) than glycine-negative cell bodies (25.1 +/- 5.3 microns; N = 274). Cell bodies in the spiral ganglion were only glutamate immunoreactive, whereas staining for glutamate, glycine, and GABA was dense in the ventral cochlear nucleus. These results demonstrate that thicker vestibular afferent fibers represent a particular subpopulation that differs from the majority of thinner afferents due to their glycine immunoreactivity.

Nitric oxide is known to function as a neurotransmitter in the central nervous system. It is also known to be involved in the central nervous system excitatory amino acid neurotransmission cascade. Activation of excitatory amino acid receptors causes an influx of calcium, which activates nitric oxide synthase. The resulting increase in intracellular nitric oxide activates soluble guanylate cyclase, leading to a rise in cyclic guanosine monophosphate. The excitatory amino acids glutamate and aspartate are found in the vestibular system and have been postulated to function as vestibular system neurotransmitters. Although nitric oxide has been investigated as a neurotransmitter in other tissues, no published studies have examined the role of nitric oxide in the vestibular system. Neuronal NADPH-diaphorase has been characterized as a nitric oxide synthase. This enzyme catalyzes the conversion of L-arginine to L-citrulline, producing nitric oxide during the reaction. We used a histochemical stain characterized by Hope et al. (Proc Natl Acad Sci 1991;88:2811) as specific for neuronal nitric oxide synthase to localize the enzyme in the rat vestibular system. An immunocytochemical stain was used to examine rat inner ear tissue for the presence of the enzyme's end product, L-citrulline, thereby demonstrating nitric oxide synthase activity. Staining of vestibular ganglion sections showed nitric oxide synthase presence and activity in ganglion cells and nerve fibers. These results indicate the presence of active nitric oxide synthase in these tissues and suggest modulation of vestibular neurotransmission by nitric oxide.

We examined the ultrastructural distribution of calcitonin gene-related peptide immunoreactivity in the peripheral vestibular system of the chinchilla to study the innervation patterns of this efferent neuropeptide. Immunoelectron microscopic localization of calcitonin gene-related peptide immunoreactive terminals in the maculae and cristae revealed an extensive innervation pattern on the afferent vestibular pathway. Calcitonin gene-related peptide immuno-reactive terminals made synaptic contacts with the unmyelinated portions of the primary afferent vestibular dendrites innervating both type I and type II hair cells. Abundant synaptic contact between calcitonin gene-related peptide immunoreactive terminals and the chalices surrounding type I hair cells was observed. Direct contact between calcitonin gene-related peptide immunoreactive terminals and type II hair cells was observed. In addition, vesiculated efferent terminals without calcitonin gene-related peptide immunoreactivity were seen synapsing on the chalices of type II hair cells and on the surrounding type I hair cells. The primary afferent somata in the vestibular ganglion of Scarpa did not contain calcitonin gene-related peptide immunoreactivity. Unmyelinated calcitonin gene-related peptide immunoreactive axons passed among the primary afferent fibers in Scarpa's ganglion, and these fibers continued through the subepithelial regions of the vestibular end-organs. The calcitonin gene-related peptide immunoreactive axons ramified to produce numerous calcitonin gene-related peptide immunoreactive terminals throughout the neurosensory epithelium of the maculae and cristae. These data suggest that calcitonin gene-related peptide-mediated modulation of the afferent vestibular system is functionally important.

The vestibular primary afferent projection to the cerebellum of the rabbit was studied with retrograde and orthograde tracers. We injected individual lobules of the cerebellum with horseradish peroxidase (HRP) or wheat germ agglutinin-HRP (WGA-HRP). Following these injections the numbers of labeled and unlabeled cells in Scarpa's ganglion were counted. Approximately 64–89% of the cells in Scarpa's ganglion were labeled retrogradely following uvula-nodular injections. About 2% of the cells in the ipsilateral Scarpa's ganglion were labeled after injections of the flocculus. Virtually no cells were labeled following injections of the ventral paraflocculus. The vestibular primary afferent projection to the uvula-nodulus is so extensive that it must be part of a collateral system that also innervates the vestibular nuclei. This collateral projection pattern was confirmed by using fluorescent tracers injected into the uvula-nodulus and vestibular complex. Fluorogold was injected into the uvula-nodulus and peroxidaserhodamine isothiocyanate was injected into the vestibular complex. More than 50% of the neurons in Scarpa's ganglion were double labeled by these subtotal injections. The dense vestibular primary afferent projection to the uvula-nodulus was confirmed by using the C fragment of tetanus toxin (TTC) injected into the labyrinth as an orthograde tracer. With the TTC technique, the vestibular primary afferent projection to the uvula-nodulus terminated exclusively in the ipsilateral granule cell layer of lobules 9d and 10. Much sparser vestibular primary afferent projections were found in the banks of major cerebellar sulci. A barely detectable projection was found to the flocculus and ventral paraflocculus. © 1993 Wiley-Liss, Inc.

Sensory nerve fibers of the cochleo-vestibular ganglion (CVG) innervate the otic epithelium in the early chick embryo by directed growth. To see if the target tissue could exert a tropic influence, we co-cultured CVGs from chick embryos (Hamburger-Hamilton stages 16-30) in a 3D collagen matrix with their normal target epithelium or with other epithelial tissues taken from the same or different stages of development. The pattern of neurite outgrowth and the viability of the CVG after five days in vitro were assessed histologically with a silver method. On the basis of the patterns of neurite outgrowth directed toward the epithelium, the cultures were classified as having slightly, mostly, exclusively, or no directed outgrowth. Of 49 cultures containing otic epithelium, 33 had mostly or exclusively directed growth patterns. This effect did not depend on any particular stage difference between co-cultures or on their viability in vitro. Cultures of non-sensory otic epithelium (endolymphatic duct) also presented directed growth patterns. Co-cultures with ectoderm from forelimb or visceral arch had little, if any, directed growth. The directed growth could not be explained simply as a result of guidance by non-neuronal cells or of the viability of the explants. The results are consistent with the hypothesis that the otic epithelium provides a tropic factor that attracts growing CVG fibers.

Cobaltic-lysine complex was used to label the afferent and efferent components of the glossopharyngeal nerve in the ganglion and brainstem of the Mexican salamander, axolotl (Ambystoma mexicanum). The distribution of afferent cell bodies in the combined glossopharyngeal-vagus ganglion (the IX-X ganglion) was reconstructed from serial sections, and the sizes of the cell bodies were measured. The central projection of afferents and the location of efferent cell bodies were determined by the tracer. The afferent cell bodies in the ganglion were medium-sized (ca. 25 μm). Cell bodies with a single process were seen. The ganglion was not clearly divided into superior and inferior ganglia, as is observed in mammals and frogs, but comprised a single ganglion. Labelled cells were diffusely distributed in the rostral part of the IX-X ganglion. A few labelled cells also were seen in the caudal part, where the vagus nerve fibers and cell bodies were mainly distributed. Double labellings of the glossopharyngeal and vagus nerves with HRP and cobaltic-lysine demonstrated that the ganglion cells of each nerve are not clearly separated in the IX-X ganglion. In the brainstem, the majority of afferent fibers formed thick ascending and descending limbs in the solitary fasciculus. The remaining afferent fibers formed a thin bundle in the spinal tract of the trigeminal nerve, which had a short ascending limb and a long descending limb. These two bundles had terminal areas in the ipsilateral brainstem: in the dorsal gray matter for the solitary fasciculus and in the lateral funiculus for the spinal tract of the trigeminal nerve, respectively. The cell bodies of the efferent neurons possessed developed dendritic arborizations in the ventrolateral white matter, and formed a longitudinal cell column in the ventrolateral margin of the gray matter. Thus, the glossopharyngeal nerve system in the axolotl assumes a primordial form in its ganglions, but its topographical organization in the brainstem is basically similar to that in anurans.

The purpose of the present study was to place horseradish peroxidase on the distal processes of the three ampullary nerves in 6–8-week-old chickens so that we could identify the ganglion cells associated with each nerve and trace the specific central pathways taken by each nerve in the brainstem. We are especially interested in the pathways of the colossal vestibular fibers, which may play a role in a fast reflex pathway as suggested by their large caliber and electrotonic mode of transmission in the tangential vestibular nucleus. The cells of origin of each ampullary nerve occupy discrete portions of the vestibular ganglion. Those vestibular ganglion cells giving rise to the posterior ampullary nerve (PAN) occupy the posterior portion of Scarpa's ganglion; the ganglion cells producing the anterior (AAN) and lateral (LAN) ampullary nerves occupy the anterior ganglion, within the dorsal and ventral portions, respectively. Centrally the vestibular fibers occupy discrete portions of the tangential vestibular nucleus before bifurcating into ascending and descending tracts. The tangential nucleus receives afferents from the colossal fibers, which form spoon endings, and also from the fine vestibular fibers, which form small terminals. The ascending fibers of the posterior ampullary nerve are associated with the nucleus piriformis; the ascending fibers of the anterior and lateral ampullary nerves occupy discrete cell groups of the vestibulo-cerebellar nucleus. All three ampullary nerves have descending branches that course through the retrotangential nucleus into the descending vestibular nucleus (DVN). Within the descending vestibular nucleus, the descending fibers of the posterior ampullary nerve run dorsally and centrally, whereas fibers of the anterior ampullary nerve course ventromedially, and the lateral ampullary nerve fibers take a ventrolateral course until all three fiber bundles converge in the posterior tip of the descending vestibular nucleus. The ascending and descending fibers of each ampullary nerve form collaterals that pass to the ventrolateral and dorsomedial parts of the medial vestibular nucleus. These collaterals are derived exclusively from the fine and medium diameter vestibular fibers. Some of these ascending fibers form a distinctive tract that courses posteriorly within medial regions of the dorsomedial part of the medial vestibular nucleus. The colossal vestibular fibers, which are found within all three ampullary nerves, conform to the ampullary pathways as described, excluding the innervation of the medial vestibular nucleus.

The reported examinations were carried out on turtles, Japanese quails and chinchillas of either sex. The trigeminal nerve was studied with the modified thiocholine method and routine histological techniques. The trigeminal ganglion of the turtle is a single, compact structure connected with the trigeminal nerve. Analysis of consecutive sections showed a presence of ganglionic neurocytes only at the medial side of the trigeminal nerve. A small number of nerve cells and large number of fibres is characteristic for the investigated structure. The trigeminal ganglion of Japanese quail is a structure connected with two branches (ophthalmic and mandibular) of the trigeminal nerve. Both parts of the ganglion distinctly differ in their morphology. The part connected with the ophthalmic branch – V1 forms a thickening of the nerve, which becomes narrow approaching the orbital cavity. The second part connected with the mandibular branch – V2 is of regular shape. This part of the ganglion is located in the vicinity of a delicate maxillary branch. Histological investigations allow us to observe a characteristic arrangement of neurocytes with only one layer of nerve cells on the surface of the ganglion. The trigeminal ganglion in chinchilla is a single, irregular cluster of neurocytes closely connected with trigeminal nerve where this nerve divides into ophthalmic and maxillary nerves. Agglomerations of neurocytes allow us to distinguish two parts of trigeminal ganglion. From the medial aspect, the nasal part of ganglion is connected with this part of the trigeminal nerve, which is not distinctly divided into ophthalmic and maxillary branches. The caudal part of the trigeminal ganglion is connected with the mandibular branch of trigeminal nerve. These two parts are separated from each other by the motor radix of the trigeminal nerve. From the lateral side all parts of the ganglion form one structure and it is difficult to distinguish precisely different cell aggregations corresponding to the particular branches of trigeminal nerve.

Using a computer-aided three-dimensional reconstruction and measurement method, the authors studied the shape and dimensions of Scarpa's ganglion and interrelations of the ganglion to nearby important anatomical structures in 10 normal human temporal bones obtained from 10 individuals of different ages (12 gestational weeks to 72 years). In the postnatal cases, Scarpa's ganglion was shaped like a distorted hourglass. Although the shape of the ganglion varied somewhat among specimens, a superior division, inferior division, and isthmus ganglionaris were distinguishable in each case. Volume of the ganglion was relatively consistent after birth, so that width and height of the ganglion were inversely correlated with each other (linear regression analysis, P less than .05). There seemed to be, however, a possible postnatal increase in the distances from the ganglion to neighboring structures and in the length of the internal auditory canal; these increases were probably the result of postnatal growth of the temporal bone. Moreover, the width of the ganglion in any single specimen was found to correlate significantly positively with the distances from the ganglion to the vestibular end-organs and the length of the internal auditory canal in that specimen (linear regression analysis, P less than .01). This finding suggests that Scarpa's ganglion may be elongated secondary to the elongation of the vestibular nerve due to the growth of the temporal bone. In fetuses, each portion of the ganglion was less clearly identifiable than that portion in postnatal cases. The dimensions of Scarpa's ganglion were found to have reached maturity around the time of birth.

The central projections of the external gustatory system in bullhead catfish were examined using orthograde degeneration and retrograde transport of horseradish peroxidase (HRP) techniques. Both large and small cells were observed in the anterior ganglion which contains a mixture of elements from the trigeminal, facial and anterior lateral line nerves. Some of the large cells on the lateral margin of the ganglion were found to belong to the lateral line system. No separation of trigeminal and facial nerve connections could be made. Using HRP, the relation between the barbels and specific ganglion regions was determined. The dorsalmost portion of the ganglion received recurrens nerve inputs (from taste buds on the trunk); the rostromedial portion of the ganglion, from the nasal barbel nerve; and the ventral portion of the ganglion, from the maxillary and mandibular barbel nerves. The facial lobe (similar to part of the n. solitarius) was found to be divided into lobules by fascicles of nerve fibers. The lateral lobule received input only from the dorsal-most part of the ganglion (recurrens nerve: trunk receptors); the intermediate lobule from the rostro-lateral part of the ganglion (nasal barbel); and the medial lobule from the ventral areas of the ganglion (maxillary and mandibular barbels). Thus a topographical relationship exists between the different taste receptor groups and their locus of representation in the facial lobe. The trunk receptors connect to the lateral lobule; the nasal barbel receptors to the intermediate lobule, and the maxillo-mandibular receptors to the medial lobule.

Superior vestibular neurons were penetrated with horseradish peroxidase (HRP)-loaded glass microelectrodes in anesthetized cats and identified electrophysiologically following electrical stimulation of the vestibular nerves and oculomotor complex. Neurons that were not antidromically activated from the oculomotor complex were stained by intracellular injection of horseradish peroxidase. Three types of neurons are identified according to their initial axonal trajectories into the cerebellum, the dorsal pontine reticular formation, or the brachium conjunctivum. Ipsilateral vestibular nerve input to all neurons is primarily monosynaptic and excitatory, whereas the contralateral is inhibitory. The neurons are located in the periphery of the superior vestibular nucleus. Soma diameters range from 20.5 micrometers to 44 micrometers. Most neurons exhibit globular and ovoid cell bodies. The dendritic arbors are intermediate between iso- and allodendritic branching patterns. The few spines and dendritic appendages present are distributed mainly distally on the dendrites. Soma size does not correlate with axon diameter, number of dendrites, or dendritic territories.

The sections in this article are: 1 Anatomy of Primary Auditory Neurons 1.1 General Features of Primary Neurons 1.2 Nerve Endings in Cochlea 1.3 Nerve Fibers in Cochlea 1.4 Neuronal Cell Bodies in Spiral Ganglion 1.5 Auditory Nerve Fibers in Internal Auditory Meatus 1.6 Primary Innervation of Cochlear Nucleus 2 Physiological Data on Auditory Nerve 2.1 Conceptual Stages of Peripheral Auditory System 2.2 Methodology in Stimulus-Response Relationships of Auditory Nerve Units 2.3 Stimulus-Response Relationships For Auditory Nerve Units 2.4 Relationship Between Gross Neural Potentials and Unit Activity 2.5 Effects of Stimulating Efferent Fibers to Cochlea 2.6 Responses in Abnormal Ears 2.7 Electric Stimulation of Auditory Nerve Fibers 3 Implications of Data on Auditory Nerve 3.1 Peripheral Mechanisms Generating Auditory Nerve Activity 3.2 Central Processing of Auditory Information 3.3 Addendum

This report documents and quantitates the naturally occurring death of neurons in the cochlear and vestibular ganglia during the normal development of the chick embryo. The data are compared with the amount of cell loss in the cochlear and vestibular nuclei to evaluate the possibility that the death of the neurons in the same sensory pathway may be related to the formation of their connections. Neurons of the cochlear and vestibular ganglia of the chick were counted and their cell bodies measured with light microscopic methods at the beginning of their period of synapse formation, embryonic day 8 (E8), at the end of the period of synapse formation (E14), and again at a more mature stage, post-hatching day 14 (P14). The number of neurons declined between E8 and E14 in the cochlear ganglion by 25% (from 11170 to 8353) and in the vestibular ganglion by 24% (from 12687 to 9613). The neuronal population remained relatively stable between E14 and P14. Cell counts at each age were accompanied by morphological descriptions of the neurons and of their synaptic targets in the sensory epithelia of the inner ear. During the period of cell loss the mean diameters of the cell bodies in the cochlear ganglion increased slightly, from 9.5 to 111 μm and reached 17.6 μm at P14, while those in the vestibular ganglion increased from 10.4 to 13.3 μm and reached 20.9 μm at P14. The period of cell loss in the cochlear and vestibular ganglia coincides with the stages in their development when the sensory neurons have already contacted their target cells in the primary sensory nuclei and in the receptor epithelium. It is during this period that the transformation from the immature axonal endings to the definitive types of synapses begins. Thus the present data are consistent with the view that the formation of central as well as peripheral synapses is involved in determining the extent of neuronal death during the development of sensory neurons.

Bipolar vestibular ganglion cells give rise to the colossal vestibular fibers in the chicken. These fibers form the largest calycine endings in the cristae ampullares and also the spoon endings in the tangential vestibular nucleus of the medulla oblongata. Because these synaptic endings are two of the largest and most distinctive in the vertebrate nervous system, they are especially suitable for comparisons of the development of synapses and synaptic endings of a specific cell type. An ultrastructural study of the spoon endings and quantitative data on their synapses were available from material of 15-day-old chick embryos, hatchlings, and 3-yr-old chickens. Here we provide similar data on the large calyces. Briefly, large calyces exhibited no ultrastructural changes corresponding to the changes in the spoon endings apparent when they retract from their target cell surfaces around hatching time. However, the concentration of the ribbon synapses at the large calyces decreased around hatching, when the concentration of the chemical synapses at the spoon endings declined. Moreover, the concentration of the ribbon synapses at the large calyces corresponded closely to the concentration of the chemical synapses at the spoor endings at the same age. Thus at the developmental ages studied, there were similar concentrations in the peripheral and central synapses formed at two different synaptic endings, both derived from one cell type and participating in the same neural pathway. These findings raise the issue of how synapses are regulated locally, but also suggest the possibility for central-peripheral interactions to produce correlative changes in parallel.

Stimulation of preganglionic axons arising from different levels of the thoracic spinal cord causes different effects on end-organs supplied by the superior cervical ganglion (Langley, 1892; Nja & Purves, 1977a; Lichtman, Purves & Yip, 1979). For example, stimulation of the first thoracic ventral root (T1) causes pupillary dilatation and widening of the palpebral fissure; stimulation of T4, on the other hand, has little effect on the eye, even though axons arising from this level innervate about as many superior cervical ganglion cells as those from T1. Thus ganglion cell innervation is selective. (1) Three months after crushing the major post-ganglionic branches of the superior cervical ganglion this differential effectiveness is lost: T1 and T4 stimulation have approximately equal effects on the end-organs of the eye. (2) In normal animals, the cellular counterpart of selective end-organ effects is the innervation of each ganglion cell by a contiguous subset of the spinal segments that innervate the ganglion as a whole. One of these segments is usually dominant, the strength of innervation from adjacent segments falling off as a function of distance from the dominant one (Nja & Purves, 1977a). Intracellular recordings from ganglion cells 3 months after post-ganglionic axotomy showed that this selective pattern is re-established. (3) Since the innervation of ganglion cells appears normal, the abnormal end-organ responses after post-ganglionic axotomy suggest that ganglion cell axons are not limited to their original targets during peripheral re-innervation. This suggestion is supported by the finding that ganglion cells sending axons to different peripheral destinations via the second and third cervical spinal nerves were no longer distinguishable on the basis of their segmented inputs 3 months after post-ganglionic axotomy. (4) Similar results were obtained when the preganglionic cervical trunk was cut at the same time as the post-ganglionic axons were crushed; the pattern of end-organ responses was abnormal, whereas individual ganglion cells were re-innervated according to the rules of contiguity and segmental dominance. (5) These results indicate that ganglion cells do not undergo a compensatory change in the segmental innervation they receive when their axons regenerate to targets different from, or in addition to those they originally innervated, even when an entirely new set of ganglionic connexions is formed. This suggests that ganglion cells, or some aspect of their immediate environment, possess a permanent label that determines the segmental innervation they receive.

The central distribution of afferents that innervate the macula of the saccule and the crista of the posterior canal was assessed in the gerbil following the direct injection of horseradish peroxidase (HRP) separately into the sensory neuroepithelia of each peripheral receptor organ. Ganglion cells innervating the posterior canal were located in the caudal part of the inferior ganglion, while those cells innervating the saccule were located in the rostral part of the inferior ganglion, scattered in the superior ganglion, and concentrated at the junction (isthmus) between the two. The paths of the central axons of these two groups of ganglion cells through the vestibular root and their division into ascending or descending pathways were similar. However, the distributions of their terminals were different. The posterior canal projected to medial parts of the vestibular nuclear complex. Terminals were found in the medial and superior vestibular nuclei. The posterior canal also projected to the uvula of the cerebellum. The saccule projected to more lateral-lying brainstem areas. Terminal fields were located in the lateral and descending vestibular nuclei and cell group y. Saccule projections outside the vestibular complex were observed to the lateral cuneate nucleus, the N. gigantocellularis, and the cerebellar cortex. Of the eight areas receiving primary afferent projections from these two organs, only within the medial and descending vestibular nuclei and the cerebellar cortex were overlapping projections observed.