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Vertebrates possess paired cranial sensory ganglia derived from two embryonic cell populations, neural crest and placodes. Cranial sensory ganglia arose prior to the divergence of jawed and jawless vertebrates, but the developmental mechanisms that facilitated their evolution are unknown. Using gene expression and cell lineage tracing experiments in embryos of the sea lamprey, Petromyzon marinus, we find that in the cranial ganglia we targeted, development consists of placode-derived neuron clusters in the core of ganglia, with neural crest cells mostly surrounding these neuronal clusters. To dissect functional roles of neural crest and placode cell associations in these developing cranial ganglia, we used CRISPR/Cas9 gene editing experiments to target genes critical for the development of each population. Genetic ablation of SoxE2 and FoxD-A in neural crest cells resulted in differentiated cranial sensory neurons with abnormal morphologies, whereas deletion of DlxB in cranial placodes resulted in near-total loss of cranial sensory neurons. Taken together, our cell-lineage, gene expression, and gene editing results suggest that cranial neural crest cells may not be required for cranial ganglia specification but are essential for shaping the morphology of these sensory structures. We propose that the association of neural crest and placodes in the head of early vertebrates was a key step in the organization of neurons and glia into paired sensory ganglia., (© 2020 Wiley Periodicals, Inc.)

After exiting the hindbrain, branchial motor axons reach their targets in association with sensory ganglia. The trigeminal ganglion has been shown to promote motor axon growth from rhombomeres 2/3 and 4/5, but it is unknown whether this effect is ganglion specific and through which signals it is mediated. Here, we addressed these questions by co-cultures of ventral rhombomere 8 explants with cranial and spinal sensory ganglia in a collagen gel matrix. Our results show that all cranial sensory ganglia and even a trunk dorsal root ganglion can promote motor axon growth and that ganglia isolated from older embryos had a stronger effect on the axonal growth than younger ones. We found that brain-derived neurotrophic factor (BDNF) and nerve growth factor (NGF) are necessary and sufficient for this effect. Altogether, our results demonstrate that the promoting effect of sensory ganglia on cranial motor axon growth is stage dependent, but not ganglion specific and is mediated by BDNF and NGF signals.

The primary vestibular neurons convey afferent information from hair cells in the inner ear to the vestibular nuclei and the cerebellum. The intrinsic firing properties of vestibular ganglion cells (VGCs) are heterogeneous to sustained membrane depolarization, and undergo marked developmental changes from phasic to tonic types during the early postnatal period. Previous studies have shown that low-voltage-activated potassium channels, Kv1 and Kv7, play a critical role in determining the firing pattern of VGCs. In the present study, we explored the developmental changes in the properties of hyperpolarization-activated current (Ih) in rat VGCs and the role played by Ih in determining the firing properties of VGCs. Tonic firing VGCs showed a larger current density of Ih as compared to phasic firing VGCs, and tonic firing VGCs became phasic firing in the presence of ZD7288, an Ih channel blocker, indicating that Ih contributes to control the firing pattern of VGCs. The amplitude of Ih increased and the activation kinetics of Ih became faster during the developmental period. Analysis of developmental changes in the expression of hyperpolarization-activated cyclic nucleotide-gated cation (HCN) channels revealed that expression of HCN1 protein and its mRNA increased during the developmental period, whereas expression of HCN2-4 protein and its mRNA did not change. Our results suggest that HCN1 channels as well as Kv1 channels are critical in determining the firing pattern of rat VGCs and that developmental up-regulation of HCN1 transforms VGCs from phasic to tonic firing phenotypes., (Copyright © 2014 IBRO. Published by Elsevier Ltd. All rights reserved.)

The dysfunction of the ubiquitin proteasome system has been related to a broad array of neurodegenerative disorders in which the accumulation of misfolded protein aggregates causes proteotoxicity. The ability of proteasome inhibitors to induce cell cycle arrest and apoptosis has emerged as a powerful strategy for cancer therapy. Bortezomib is a proteasome inhibitor used as an antineoplastic drug, although its neurotoxicity frequently causes a severe sensory peripheral neuropathy. In this study we used a rat model of bortezomib treatment to study the nucleolar and Cajal body responses to the proteasome inhibition in sensory ganglion neurons that are major targets of bortezomib-induced neurotoxicity. Treatment with bortezomib induced dose-dependent dissociation of protein synthesis machinery (chromatolysis) and nuclear retention of poly(A) RNA granules resulting in neuronal dysfunction. However, as a compensatory response to the proteotoxic stress, both nucleoli and Cajal bodies exhibited reactive changes. These include an increase in the number and size of nucleoli, strong nucleolar incorporation of the RNA precursor 5'-fluorouridine, and increased expression of both 45S rRNA and genes encoding nucleolar proteins UBF, fibrillarin and B23. Taken together, these findings appear to reflect the activation of the nucleolar transcription in response to proteotoxic stress Furthermore, the number of Cajal bodies, a parameter related to transcriptional activity, increases upon proteasome inhibition. We propose that nucleoli and Cajal bodies are important targets in the signaling pathways that are activated by the proteotoxic stress response to proteasome inhibition. The coordinating activity of these two organelles in the production of snRNA, snoRNA and rRNA may contribute to neuronal survival after proteasome inhibition. This article is part of a Special Issue entitled: Role of the Nucleolus in Human Disease., (Copyright © 2013 Elsevier B.V. All rights reserved.)

The neural system appears before the vascular system in the phylogenetic tree. During evolution, vascular system generation takes advantage of the pre-existing vascular endothelial growth factor (VEGF) in order to form its networks. Nevertheless, the role of VEGF in neuronal and glial cells is not yet completely understood. In order to support the hypothesis of a neural role for VEGF, we searched for VEGF- and VEGF receptor (VEGFR)-like immunoreactivities (immunohisto/cytochemistry and Western blotting) in the eyestalk of the invertebrate Ucides cordatus (Crustacea, Brachyura, Ucididae). Our results showed that both neurons and glial cells expressed VEGF-immunoreactivity, and that VEGFR was evidenced in neural cells. This is the first report about the VEGF/VEGFR-like immunoreactivities in the nervous tissue of a crustacean, and enables U. cordatus to be included in the repertoire of animal models used for ascertaining the role of VEGF in the nervous system., (Copyright © 2014 Elsevier Inc. All rights reserved.)

Neurochemical features in sympathetic and afferent neurons are subject to change during development. Nitric oxide (NO) plays a developmental role in the nervous system. To better understand the neuroplasticity of sympathetic and afferent neurons during postnatal ontogenesis, the distribution of neuronal NO synthase (nNOS) immunoreactivity was studied in the sympathetic para- and prevertebral, nodose ganglion (NG) and Th2 and L4 dorsal root ganglia (DRG) from female Wistar rats of different ages (newborn, 10-day-old, 20-day-old, 30-day-old, 2-month-old, 6-month-old, 1-year-old, and 3-year-old). nNOS-positive neurons were revealed in all sensory ganglia but not in sympathetic ones from birth onward. The percentage of nNOS-immunoreactive (IR) neurons increased during first 10 days of life from 41.3 to 57.6 in Th2 DRG, from 40.9 to 59.1 in L4 DRG and from 31.6 to 38.5 in NG. The percentage of nNOS-IR neurons did not change in the NG later during development and senescence. However, in Th2 and L4 DRG the proportion of nNOS-IR neurons was high in animals between 10 and 30days of life and decreased up to the second month of life. In 2-month-old rats, the percentage of nNOS-IR neurons was 52.9 in Th2 DRG and 51.3 in L4 DRG. We did not find statistically significant differences in the percentage of nNOS-IR neurons between Th2 and L4 DRG and between young and aged rats. In NG and DRG of 10-day-old and older rats, a high proportion of nNOS-IR neurons binds isolectin B4. In newborn animals, only 41.3%, 45.3% and 28.4% of nNOS neuron profiles bind to IB4 in Th2, L4 DRG and NG, respectively. In 10-day-old and older rats, the number of sensory nNOS-IR neurons binding IB4 reached more than 90% in DRG and more than 80% in NG. Only a small number of nNOS-positive cells showed immunoreactivity to calcitonin gene-related peptide, neurofilament 200, calretinin. The information provided here will also serve as a basis for future studies investigating mechanisms of the development of sensory neurons., (Copyright © 2013 IBRO. Published by Elsevier Ltd. All rights reserved.)

Previous studies indicated that intracerebroventricular administration of nerve growth factor (NGF) leads to massive Schwann cell hyperplasia surrounding the medulla oblongata and spinal cord. This study was designed to characterize the proliferation of peripheral glial cells, that is, Schwann and satellite cells, in the trigeminal ganglia and dorsal root ganglia (DRG) of adult rats during two weeks of NGF infusion using bromodeoxyuridine (BrdU) to label dividing cells. The trigeminal ganglia as well as the cervical and lumbar DRG were analyzed. Along the entire neuraxis a small number of dividing cells were observed within these regions under physiological condition. NGF infusion has dramatically increased the generation of new cells in the neuronal soma and axonal compartments of sensory ganglia and along the dorsal root and the dorsal root entry zone. Quantification of BrdU positive cells within sensory ganglia revealed a 2.3- to 3-fold increase in glial cells compared to controls with a similar response to NGF for the different peripheral ganglia examined. Immunofluorescent labeling with S100β revealed that Schwann and satellite cells underwent mitosis after NGF administration. These data indicate that intracerebroventricular NGF infusion significantly induces gliogenesis in trigeminal ganglia and the spinal sensory ganglia and along the dorsal root entry zone as well as the dorsal root.

Afferent neurons containing substance P (SP) were studied immunohistochemically in the sensory ganglion of the spinal nerve in 30 rats aged 10-90 days. The results obtained indicated that SP-immunoreactive neurons are present in thesel ganglia from the moment of birth. During the development, the percentage of SP-containing neurons decreased till day 10. SP-immunoreactive neurons were represented by the cells of very small or small size.

Phosphatase and tensin homolog (PTEN) is a tumor suppressor gene that controls neural stem cell renewal and differentiation and is a potential target for regeneration in the optic nerve. Here we show that it has a critical pattern of expression in the mammalian developing auditory system. PTEN was expressed in the cochlear-vestibular ganglion at embryonic day 10.5 and then progressively in hair cells as they differentiated from the base to the apex of the cochlea. By postnatal day 7, PTEN was downregulated in hair cells and subsequently in the neurons. This very specific, transient expression pattern suggests that PTEN plays a crucial role in the differentiation of the sensory neurons and hair cells and that it is a potential therapeutic target for hearing regeneration.

Stress-induced arrest of ventilatory motor pattern generation is tightly correlated with an abrupt increase in extracellular potassium concentration ([K+]o) within the metathoracic neuropil of the locust, Locusta migratoria. Na+/K+-ATPase inhibition with ouabain elicits repetitive surges of [K+]o that coincide with arrest and recovery of motor activity. Here we show that ouabain induces repetitive [K+]o events in a concentration-dependent manner. 10(-5)M, 10(-4)M, and 10(-3)M ouabain was bath-applied in semi-intact locust preparations. 10(-4)M and 10(-3)M ouabain reliably induced repetitive [K+]o events whereas 10(-5)M ouabain had no significant effect. In comparison to 10(-4)M ouabain, 10(-3)M ouabain increased the number and hastened the time to onset of repetitive [K+]o waves, prolonged [K+]o event duration, increased resting [K+]o, and diminished the absolute value of [K+]o waves. Recovery of motor patterning following [K+]o events was less likely in 10(-3)M ouabain. In addition, we show that K+ channel inhibition using TEA suppressed the onset and decreased the amplitude of ouabain-induced repetitive [K+]o waves. Our results demonstrate that ventilatory circuit function in the locust CNS is dependent on the balance between mechanisms of [K+] accumulation and [K+] clearance. We suggest that with an imbalance in favour of accumulation the system tends towards a bistable state with transitions mediated by positive feedback involving voltage-dependent K+ channels.

Recent evidence indicates that endomorphins, endogenous mu-opioid receptor (MOR) agonists, modulate synaptic transmission in both somatic and visceral sensory pathways. Here we show that endomorphin-2 (END-2) is expressed in newborn rat dorsal root ganglion (DRG) and nodose-petrosal ganglion complex (NPG) neurons, and rarely co-localizes with brain-derived neurotrophic factor (BDNF). In order to examine activity-dependent release of END-2 from neurons, we established a model using dispersed cultures of DRG and NPG cells activated by patterned electrical field stimulation. To detect release of END-2, we developed a novel rapid capture enzyme-linked immunosorbent assay (ELISA), in which END-2 capture antibody was added to neuronal cultures shortly before their electrical stimulation. The conventional assay was effective at reliably detecting END-2 only when the cells were stimulated in the presence of CTAP, a MOR-selective antagonist. This suggests that the strength of the novel assay is related primarily to rapid capture of released END-2 before it binds to endogenous MORs. Using the rapid capture ELISA, we found that stimulation protocols known to induce plastic changes at sensory synapses were highly effective at releasing END-2. Removal of extracellular calcium or blocking voltage-activated calcium channels significantly reduced the release. Together, our data provide the first evidence that END-2 is expressed by newborn DRG neurons of all sizes found in this age group, and can be released from these, as well as from NPG neurons, in an activity-dependent manner. These results point to END-2 as a likely mediator of activity-dependent plasticity in sensory pathways.

This paper studies the molecular organization, neuronal distribution and cellular differentiation dynamics of the giant fibrillar centers (GFCs) of nucleoli in rat sensory ganglia neurons. The GFC appeared as a round nucleolar domain (1-2 microm in diameter) partially surrounded by the dense fibrillar component and accompanied by numerous small FCs. By immunocytochemistry, the GFC concentrated the upstream binding factor, which may serve as a marker of this structure, and also contain RNA polymerase I, DNA topoisomerase I, SUMO-1 and Ubc9. However, they lack ubiquitin-proteasome conjugates and 20S proteasome. Transcription assay with 5'-fluorouridine incorporation revealed the presence of nascent RNA on the dense fibrillar component of the neuronal nucleolus, but not within the low electron-density area of the GFC. The formation of GFCs is neuronal size dependent: they were found in 58%, 30% and 0% of the large, medium and small neurons, respectively. GFCs first appeared during the postnatal period, concomitantly with a stage of neuronal growth, myelination and bioelectrical maturation. GFCs were not observed in segregated nucleoli induced by severe inhibition of RNA synthesis. We suggest that the formation of GFCs is associated with a high rate of ribosome biogenesis of the transcriptionally more active large-size neurons.

Hyperoxia, during development in rats, results in hypoxic chemosensitivity ablation, carotid body hypoplasia, and reduced chemoafferents. We hypothesized that hyperoxia increases reactive oxygen species (ROS) in cell bodies of chemoafferents. Organotypic slices of petrosal-nodose ganglia from rats at day of life (DOL) 5-6 and 17-18 were exposed to 8%, 21%, or 95% O(2) for 4 h in the presence or absence of the ROS-sensitive fluorescent indicator, CM-H(2)DCFDA, and propidium iodide was used to determine the relationship between cell death and oxygen tension. In tissue slices from DOL 5-6 rats, fluorescence intensity was 182.5 +/- 2.9 for hypoxia, 217.5 +/- 3.3 for normoxia, and 336.6 +/- 3.8 for hyperoxia, (mean +/- SEM, p < 0.001, ANOVA). Normoxia increased ROS levels by 19.2% from hypoxia (p < 0.01) with a further increase of 54.8% from normoxia to hyperoxia (p < 0.001). In tissue slices from DOL 17-18 rats, ROS levels increased with increasing oxygen tension but were less than in younger animals (p < 0.01, ANOVA). The antioxidants, NAC and TEMPO-9-AC, attenuated ROS levels and cell death. Electron microscopy demonstrated that hyperoxia damages the ultrastructure within petrosal ganglion neurons. Hyperoxic-induced increased levels of ROS in petrosal ganglion neurons may contribute to loss of hypoxic chemosensitivity during early postnatal development.

To study the role of different members of the bHLH gene family for sensory organ development we have generated NSCL-1 and NeuroD compound-mutant mice. Double homozygous animals were characterized by a more severe reduction of the petrosal and vestibulocochlear ganglia than NeuroD-knockout mice. The more severe reduction of the petrosal and vestibulocochlear ganglia in double-knockout mice indicates overlapping functions of the two genes during neuronal development. Interestingly, we also found that the two genes are jointly regulated by thyroid hormone during sensory hair cell development. We further present a detailed expression analysis of NSCL-1 and NSCL-2 during sensory neuron development. NSCL-1 expression was detected in all developing cranial ganglia including the petrosal and vestibulocochlear ganglion, in inner and outer hair cells of the organ of Corti and in hair cells of the vestibular system. Expression domains in other sensory structures include the retina, Merkel cells of the developing skin and sensory cells of the tongue. The expression of NSCL-2 was restricted to developing cranial ganglia, the retina and the vestibular nerve. Both NSCL-1 and NSCL-2 genes are active only in postmitotic neurons, indicating a role for neuronal cell migration and/or differentiation within the sensory system.

Early dietary sodium restriction has profound influences on the organization of the gustatory brainstem. However, the anatomical relationships among multiple gustatory nerve inputs have not been examined. Through the use of triple-fluorescence labeling and confocal laser microscopy, terminal fields of the greater superficial petrosal (GSP), chorda tympani (CT), and glossopharyngeal (IX) nerves were visualized concurrently in the nucleus of the solitary tract (NTS) of developmentally sodium-restricted and control rats. Dietary sodium restriction during pre- and postnatal development resulted in a twofold increase in the volume of both the CT and the IX nerve terminal fields but did not affect the volume of the GSP terminal field. In controls, these nerve terminal fields overlapped considerably. The dietary manipulation significantly increased the overlapping zones among terminal fields, resulting in an extension of CT and IX fields past their normal boundaries. The differences in terminal field volumes were exaggerated when expressed relative to the respective NTS volumes. Furthermore, increased terminal field volumes could not be attributed to an increase in the number of afferents because ganglion cell counts did not differ between groups. Taken together, selective increases in terminal field volume and ensuing overlap among terminal fields suggest an increased convergence of these gustatory nerve terminals onto neurons in the NTS. The genesis of such convergence is likely related to disruption of cellular and molecular mechanisms during the development of individual terminal fields, the consequences of which have implications for corresponding functional and behavioral alterations., (Copyright 2006 Wiley-Liss, Inc.)

Inhibition of the proteasome by lactacystin, a specific blocker of the catalytic beta-subunits, results in transient neurite outgrowth by neuronal cell lines. Vice versa, as demonstrated in this study, treatment of pheochromocytoma (PC12) cells with nerve growth factor (NGF) or other differentiating agents reduces proteasomal activity. This is accompanied by an increase in mRNA and protein levels of the catalytically active subunits beta1, beta2 and beta5, but not of their inducible counterparts, indicating changes in subunit composition of the proteasome during neuronal differentiation. In contrast to neuronal cell lines, however, pre-treatment of primary neurons with proteasome inhibitors completely prevents axon formation, and lower concentrations of lactacystin (0.5-5 microm) significantly reduce axonal elongation and branching in vitro. Furthermore, established axonal networks degenerate rapidly and long-term survival of peripheral neurons is impaired in the presence of proteasome inhibitors. Axonal pathology is reminiscent of the morphological changes observed in neurodegenerative disorders and supports a crucial role of the constitutive catalytic subunits in axon initiation, maintenance and regeneration.

NGF, a neurotrophic factor best known for its role in promoting cell survival, regulates many neurodevelopmental processes, including synaptic plasticity, neurite outgrowth and programmed cell death. Although there is a large amount of data regarding NGF in the developing nervous system of many species, there is little known about its regulation and role in the frog, Xenopus laevis. In this report, immunocytochemistry was used to characterize NGF protein expression in developing tadpoles. Protein expression was analyzed in tadpoles from stage 44/45 through stage 50, a period of development characterized by extensive neurite outgrowth, neuronal differentiation and an initial period of programmed cell death. Similar to other species, NGF was expressed in sensory cells and tissues, including the inner ear, eye, olfactory system, lateral line organs, papillae in the oral cavity, and gills tufts. In addition, NGF was expressed in specific cells in the central nervous system, cranial and dorsal root ganglia, spinal sensory and motoneurons, and muscle tissues in the tail and body cavity. In the mammalian nervous system, the cytokine, interleukin-1beta (IL-1beta) induces expression of NGF. In this report, double-label immunocytochemistry was used to determine the relationship between NGF and IL-1beta. Results showed most cell types and/or tissues that expressed NGF also expressed IL-1beta. However, NGF was typically associated with cellular and nuclear membranes, whereas IL-1beta appeared in the cytoplasm and nucleolus. The nuclear localization of IL-1beta supports the idea that it regulates gene transcription in the frog. The appearance of NGF and IL-1beta in the same cells suggests they may interact to influence neural development.

First described in 1973, the Grueneberg ganglion (GG) is an arrow-shaped neuronal structure at the anterior end of the nasal cavity. It lines both sides of the nasal septum, within the nasal vestibule, close to the opening of the naris. The functions of the GG and the pattern of projections to the brain are not known. Here, we report that neurons of the mouse GG express olfactory marker protein, which is normally expressed in mature olfactory or vomeronasal sensory neurons. The approx. 500 cells in each GG are arranged in several densely packed cell clusters. Individual cells give rise to single axons, which fasciculate to form a nerve bundle that projects caudally. The axons terminate in glomeruli of the olfactory bulb, one or two large glomeruli associated with a semicircle of up to 10 smaller, somewhat diffusely organized glomeruli that surround the most anterior part of the accessory olfactory bulb. Development of the GG starts around embryonic day 16 and appears to be completed at birth; cell numbers then undergo a minor decrease during postnatal development. The strategic location of the GG, expression of olfactory marker protein, axonal projections to glomeruli at particular locations in the olfactory bulb and early development suggest that this neuronal structure performs specific chemosensory functions at neonatal stages.

The formation of the eight independent endorgan compartments (sacculus, utricle, horizontal canal, anterior canal, posterior canal, lagena, amphibian papilla, and basilar papilla) of the Xenopus laevis inner ear is illustrated as the otic vesicle develops into a complex labyrinthine structure. The morphology of transverse sections and whole-mounts of the inner ear was assessed in seven developmental stages (28, 31, 37, 42, 45, 47, 50) using brightfield and laser scanning confocal microscopy. The presence of mechanosensory hair cells in the sensory epithelia was determined by identification of stereociliary bundles in cryosectioned tissue and whole-mounts of the inner ear labeled with the fluorescent F-actin probe Alexa-488 phalloidin. Between stages 28 and 45, the otic vesicle grows in size, stereociliary bundles appear and increase in number, and the pars inferior and pars superior become visible. The initial formation of vestibular compartments with their nascent stereociliary bundles is seen by larval stage 47, and all eight vestibular and auditory compartments with their characteristic sensory fields are present by larval stage 50. Thus, in Xenopus, inner ear compartments are established between stages 45 and 50, a 2-week period during which the ear quadruples in length in the anteroposterior dimension. The anatomical images presented here demonstrate the morphological changes that occur as the otic vesicle forms the auditory and vestibular endorgans of the inner ear. These images provide a resource for investigations of gene expression patterns in Xenopus during inner ear compartmentalization and morphogenesis., (Developmental Dynamics 234:791-801, 2005. (c) 2005 Wiley-Liss, Inc.)

Retrograde axonal transport of horseradish peroxidase was used in this study to determine the location and basic morphological parameters of neurons innervating the trachea in newborn, 10-, 20-, 30-day-old and 2-month-old kittens. Labeled neurons were detected in all animals in the nodose ganglion of the vagus nerve and in the spinal ganglia (C1-C7 and T1-T6 after injection of tracer into the cervical trachea, C5-C7 and T1-T8 with injection into the thoracic part of the trachea) from both sides. The content of vagal and spinal afferent neurons innervating the cervical part of trachea declined during development. The number of spinal afferent neurons with connections to the thoracic trachea did not change but the quantity of cells in nodose ganglion supplying the thoracic trachea increased from the moment of birth till 10 and 20 days and decreased later in postnatal development. In newborn, 10-day-old and 20-day-old animals, the largest number of afferent cells was connected with the cervical part of the trachea in comparison with the thoracic one, whereas in 2-month-old kittens the relation was opposite. We suggest that afferent innervation of the trachea is not morphologically complete at the moment of birth and does not become mature until the second month of life.

This study follows the histogenesis of the oropharyngeal cavity taste buds, along with the development of the relevant neural centers and gustatory nerves, in two cichlid species: the substrate-brooding Cichlasoma cyanoguttatum and the mouth-brooding Astatotilapia flavijosephi, from fertilization to 20-day-old juveniles, grown at a temperature of 26 degrees C. Significant differences in pace of development were shown between the two social types: Substrate-brooders complete embryogenesis and hatch 48 h after fertilization (HAF) and begin to swim 120 HAF, with the yolk sac disappearing 160 HAF, whereas mouth-brooders hatch 84 HAF and begin to swim 196 HAF, with the yolk sac disappearing 360 HAF. Histogenesis of primordial taste buds occurs 75 HAF and 160 HAF in C. cyanoguttatum and A. flavijosephi, respectively. Accordingly, the related sensory ganglia and nerves (VII, IX, and X) develop much earlier in the substrate-brooded larvae and postlarvae. Nerve and brain development in juvenile A. flavijosephi of 13 mm total length (TL) closely resemble those of 8-mm-TL C. cyanoguttatum. These differences in development continue throughout the early stages of growth. Similar differences are observed in the ripening and increase in number of taste buds and dentition on the jaws and pharyngeal bones. The possible triggers and causes of such differences in development, as well as the inductors of taste bud development, are discussed.

The expression of the P2X(3) nucleotide receptor in embryonic day 14-18, postnatal day 1-14 and adult mouse sensory ganglia was examined using immunohistochemistry. Nearly all sensory neurons in dorsal root ganglia, trigeminal ganglia and nodose ganglia in embryos at embryonic day 14 expressed P2X(3) receptors, but after birth there was a gradual decline to about 50% of neurons showing positive immunostaining for P2X(3). In embryos there were only small neurons, while from postnatal day 7 both large and small neurons were present. Isolectin B(4) (IB(4))-positive neurons in dorsal, trigeminal and nodose ganglia did not appear until birth, but the numbers increased to about 50% by postnatal day 14 when a high proportion of IB(4)-positive neurons were also positively labelled for the P2X(3) receptor. About 10% of neurons in dorsal, trigeminal and nodose ganglia were positive for calcitonin gene-related peptide in embryos, nearly all of which stained for P2X(3) receptors. This increased postnatally to about 35-40% in adults, although only a few colocalised with P2X(3) receptors. Neurofilament 200 was expressed in about 50% of neurons in trigeminal ganglia in the embryo, and this level persisted postnatally. All neurofilament 200-positive neurons stained for P2X(3) in embryonic dorsal root ganglia, trigeminal ganglia and nodose ganglia, but by adulthood this was significantly reduced. The neurons that were positive for calbindin in embryonic dorsal, trigeminal and nodose ganglia showed colocalisation with P2X(3) receptors, but few showed colocalisation postnatally.

The mammalian trkB locus undergoes alternative splicing to produce two different types of brain-derived neurotrophic factor receptors. The first type is the full-length receptor tyrosine kinase (TrkB(Tk+); the second type is a truncated receptor lacking the intracellular tyrosine kinase domain (TrkB(Tk-)). To investigate the function of both types of TrkB receptor in vivo, we have generated knockout mice lacking all isoforms of the TrkB receptor (trkB-/-) and compared sensory neuron survival in these mice to that in the previously described TrkB kinase domain knockout mice (trkB(k)-/-). We observed that the presence of truncated TrkB receptors in trkB(k)-/- mice results in more severe sensory neuron losses. Increased neuron losses associated with the presence of truncated TrkB were most severe in regions where neuron survival is most dependent on brain-derived neurotrophic factor and neurotrophin-3. Our data suggest that truncated TrkB receptors negatively influence neuron survival by interfering with the function of catalytic TrkB receptors.

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., (Copyright 2001 Wiley-Liss, Inc.)

The end-artery retinal vasculature of a marsupial, the fat-tailed dunnart, was defined by India ink injection and studied in wholemounts. In the adult, the vitreal vasculature supplying the ganglion-cell layer has major paired-vessels in a horizontal H shape. These vessels skirt the area centralis and visual streak that are supplied by fine end-loops. A second vascular layer of uniformly distributed endloops arises from the superficial vessels and lies at the inner nuclear/outer plexiform border. During development, vessels enter the eye via the optic nerve head to form the upper vasculature, assuming an essentially mature arrangement prior to the formation of the area centralis and visual streak. Vessels then descend to form the lower bed. Unlike the cat, the dunnart has retinal vessels that are patent throughout development, their growth is interstitial and reductive remodelling is not seen. A retinal end-artery system may have evolved in marsupials because their precocity requires a vasculature that is functional from early stages of development.

The sensitivity of peripheral arterial chemoreceptors in the carotid body to hypoxia increases with postnatal maturation. Carotid sinus nerve activity is augmented by adenosine binding to A(2a)-adenosine receptors and attenuated by dopamine binding to D(2)-dopamine receptors. In this study, we used in situ hybridization histochemistry to determine the change in the levels of mRNA expression for A(2a) and A(1)-adenosine receptors and D(2)-dopamine receptors in the rat carotid body. We also investigated the cellular distribution and possible colocalization of these receptor mRNAs and tyrosine hydroxylase (TH) mRNAs during the first 2 weeks of postnatal development. By using immunohistocytochemistry, we detected A(2a)-adenosine receptor protein in the carotid body and petrosal ganglion. We found that A(2a)-adenosine receptor mRNA and protein are expressed in the carotid body in animals at 0, 3, 6 and 14 postnatal days. The level of A(2a)-adenosine receptor mRNA expression significantly decreased by 14 postnatal days (P<0.02 vs. day 0) while D(2)-dopamine receptor mRNA levels significantly increased by day 3 and remained greater than D(2)-dopamine receptor mRNA levels at day 0 (P<0.001 all ages vs. day 0). TH mRNA was colocalized in cells in the carotid body with A(2a) adenosine receptor and D(2)-dopamine receptor mRNAs. A(1)-adenosine receptor mRNA was not expressed in the carotid body at any of the ages examined. In the petrosal ganglion, A(1)-adenosine receptor mRNA was abundantly expressed in numerous cells, A(2a)-adenosine receptor mRNA was expressed in a moderate number of cells while D(2)-dopamine receptor mRNA was seen in a few cells in the rostral petrosal ganglion. In conclusion, using in situ hybridization histochemistry, we have shown that mRNA for both the excitatory, A(2a)-adenosine receptor, and the inhibitory, D(2)-dopamine receptor, is developmentally regulated in presumably type I cells in the carotid body which may contribute to the maturation of hypoxic chemosensitivity. Furthermore, the presence A(1)-adenosine receptor mRNAs in cell bodies of the petrosal ganglion suggests that adenosine might also have an inhibitory role in hypoxic chemotransmission.

Object: The results of previous clinical trials have indicated that intraventricular infusion of nerve growth factor (NGF) in patients with Alzheimer's disease is frustrated by the appearance of weight loss and diffuse back pain. The present study tested whether NGF induces sympathetic sprouting in sensory ganglia. Such sprouting has been implicated in previous studies as a possible mechanism of sympathetically maintained pain in neuropathic animals., Methods: Nineteen Long-Evans rats underwent intraventricular infusion of either artificial cerebrospinal fluid (ACSF; seven animals) or NGF (12 animals). After 14 days of infusion, the sensory ganglia of the trigeminal nerve and the C-2, C-8, T-1, L-4, and L-5 dorsal roots were examined for sympathetic sprouting by using tyrosine hydroxylase immunohistochemical analysis., Conclusions: In the animals receiving NGF, 52 of 144 ganglia showed sympathetic fiber sprouting. In the control animals receiving ACSF, only two of 72 ganglia showed minor sympathetic fiber sprouting. A preferential sprouting of sympathetic fibers was demonstrated at lower lumbar ganglia compared with the cervical and thoracic ganglia. The data presented here demonstrate that in the rat intraventricular NGF infusion caused sympathetic sprouting in dorsal root ganglia (p < 0.01). These findings may have importance both for the treatment of Alzheimer's disease and the understanding of neuropathic pain.

The endorgan-specific distribution of vestibular ganglion cells was studied in neonatal and postnatal rats and mice using indocarbocyanine dye (DiI) and dextran amines for retrograde and anterograde labeling. Retrograde DiI tracing from the anterior vertical canal labeled neurons scattered throughout the whole superior vestibular ganglion, with denser labeling at the dorsal and central regions. Horizontal canal neurons were scattered along the dorsoventral axis with more clustering toward the dorsal and ventral poles of this axis. Utricular ganglion cells occupied predominantly the central region of the superior vestibular ganglion. This utricular population overlapped with both the anterior vertical and horizontal canals' ganglion cells. Posterior vertical canal neurons were clustered in the posterior part of the inferior vestibular ganglion. The saccular neurons were distributed in the two parts of the vestibular ganglion, the superior and inferior ganglia. Within the inferior ganglion, the saccular neurons were clustered in the anterior part. In the superior ganglion, the saccular neurons were widely scattered throughout the whole ganglion with more numerous neurons at the posterior half. Small and large neurons were labeled from all endorgans. Examination of the fiber trajectory within the superior division of the vestibular nerve showed no clear lamination of the fibers innervating the different endorgans. These results demonstrate an overlapping pattern between the different populations within the superior ganglion, while in the inferior ganglion, the posterior canal and saccular neurons show tighter clustering but incomplete segregation. This distribution implies that the ganglion cells are assigned for their target during development in a stochastic rather than topographical fashion.

Neuropeptide Y, a 36 amino acid peptide, mediates its biological effects by activating the Y1, Y2, Y5 and Y6 receptors, which are also receptors for the structurally related peptide YY. Different classes of receptors have been suggested to be involved in different neuropeptide Y functions. In this report, we have characterized the developmental regulation and compared the cellular localization of these receptors in the developing and in the adult central and peripheral nervous systems of the mouse. RNase protection assays revealed that Y1, Y2 and Y5 messenger RNAs were expressed very early in spinal cord, brain, cerebellum and dorsal root ganglion development and were often down-regulated at times corresponding to their acquirement of the adult function in neurotransmission. In situ hybridization of the adult brain showed that Y1 was widely expressed, Y2 displayed a more restricted pattern, Y5 was expressed at very low levels and only in a few brain nuclei and Y6 was not expressed. Virtually all areas containing neurons positive for Y5 also expressed Y1, whereas many Y1-positive cells clearly did not express Y5. In contrast, Y2 was not expressed by the neurons expressing Y1 or Y5. These findings suggest that neuropeptide Y signaling in the brain could be mediated by simultaneous Y1 and Y5 activation. Similar results were also obtained in peripheral sensory neurons. Furthermore, our results suggest that neuropeptide Y/peptide YY receptors play an important role in nervous system development and that selective receptor combinations are responsible for signaling the different effects of neuropeptide Y in the peripheral and central nervous systems.

1. To define the role of environmental oxygen in regulating postnatal maturation of the carotid body afferent pathway, light and electron microscopic methods were used to compare chemoafferent neurone survival and carotid body development in newborn rats reared from birth in normoxia (21 % O2) or chronic hyperoxia (60 % O2). 2. Four weeks of chronic hyperoxia resulted in a significant 41 % decrease in the number of unmyelinated axons in the carotid sinus nerve, compared with age-matched normoxic controls. In contrast, the number of myelinated axons was unaffected by hyperoxic exposure. 3. Chemoafferent neurones, located in the glossopharyngeal petrosal ganglion, already exhibited degenerative changes following 1 week of hyperoxia from birth, indicating that even a relatively short hyperoxic exposure was sufficient to derange normal chemoafferent development. In contrast, no such changes were observed in the vagal nodose ganglion, demonstrating that the effect of high oxygen levels was specific to sensory neurones in the carotid body afferent pathway. Moreover, petrosal ganglion neurones were sensitive to hyperoxic exposure only during the early postnatal period. 4. Chemoafferent degeneration in chronically hyperoxic animals was accompanied by marked hypoplasia of the carotid body. In view of previous findings from our laboratory that chemoafferent neurones require trophic support from the carotid body for survival after birth, we propose that chemoafferent degeneration following chronic hyperoxia is due specifically to the loss of target tissue in the carotid body.

Triple label immunohistochemistry was used to study the coexistence of the catecholamine-synthesising enzymes dopamine beta-hydroxylase (DBH) and tyrosine hydroxylase (TH) and several neuropeptides including neuropeptide Y (NPY), vasoactive intestinal polypeptide (VIP), substance P (SP), calcitonin gene-related peptide (CGRP), somatostatin (SOM) and galanin (GAL) as well as nitric oxide synthase (NOS) in developing pelvic paraganglion cells in a series of human male fetal, neonatal and infant specimens ranging in age from 13 wk of gestation to 3 y postnatal. 13-20 wk old fetal specimens possessed large clusters of paraganglion cells lying lateral to the urinary bladder and prostate gland which were intensely DBH-immunoreactive (-IR) but lacked TH, NOS and the neuropeptides investigated. With increasing fetal age small clusters of paraganglion cells were observed in the muscle coat of the urinary bladder. At 23 wk of gestation occasional paraganglion cells were NOS or NPY-IR while at 26 wk of gestation the majority of paraganglion cells were TH-IR and a few were SOM or GAL-IR. Some postnatal paraganglia within the bladder musculature contained cells which were all VIP, SP or CGRP-IR while others displayed coexistence of NOS and NPY, SP and CGRP, or NPY and VIP. The presence of NOS in certain paraganglion cells indicates their capacity to generate nitric oxide (NO). These results show that human paraganglion cells develop different phenotypes possibly dependent upon their location within the bladder wall. A delicate plexus of branching varicose nerves was observed in the fetal paraganglia which increased in density with increasing gestational age. The majority of these nerves were VIP-IR while others were CGRP, SP, NPY, NOS or GAL-IR. The presence of nerve terminals adjacent to the paraganglion cells implies a neural influence on the functional activity of the paraganglia. Some paraganglia in the late fetal and early postnatal specimens contained Timofeew's sensory corpuscles, resembling pacinian corpuscles in their morphology. The central nerve fibre of these corpuscles displayed immunoreactivity for SP, CGRP and NOS, the latter indicating a possible role for NO in afferent transmission from the urinary bladder. In addition, a few corpuscles were penetrated by a noradrenergic nerve fibre immunoreactive for NPY and TH, which may have a modulatory role on the sensory receptor.

The ARIA (acetylcholine receptor inducing activity) polypeptide is a member of the neuregulin gene family. It was originally purified on the basis of its ability to induce skeletal muscle nicotinic acetylcholine receptors (nAChRs). ARIA mRNA is expressed in ventral horn motor neurons and brain cholinergic neurons. We report here that ARIA mRNA is heavily expressed in the embryonic, developing, and adult peripheral auditory and vestibular ganglia, the spiral ganglion and Scarpa's ganglion. Neither ganglion is cholinergic, but both express mRNAs for nicotinic and muscarinic receptors. The expression of ARIA in these ganglia may be related to the regulation of cholinergic receptors or a more general role for ARIA in growth and development.

Analysis of mice carrying targeted mutations in genes encoding neurotrophins and their signalling Trk receptors has provided critical information regarding the role that these molecules play in the mammalian nervous system. In this study we generated mice defective in both TrkB and TrkC tyrosine kinase receptors to determine the biological effects of these receptors in the absence of compensatory mechanisms. trkB(-/-);trkC(-/-) double-mutant mice were born at the expected frequency, indicating that TrkB and TrkC signalling are not required for embryonic survival. However, these double-mutant mice had a significantly shorter lifespan and displayed more severe sensory defects than their single-mutant trkB(-/-) and trkC(-/-) littermates. The most dramatic sensory deficit observed in trkB(-/-);trkC(-/-) mutant mice was the absence of vestibular and cochlear ganglia. Interestingly, these mice developed inner ear sensory epithelia in spite of the complete absence of sensory innervation. Analysis of the CNS in trkB(-/-);trkC(-/-) mutant mice revealed a well formed hippocampus, cortex and thalamus. Moreover, the pattern of expression of several neuronal markers appeared normal in these animals. These observations suggest that neurotrophin signalling through TrkB and TrkC receptors is essential for the development of sensory ganglia; however, it does not play a major role in the differentiation and survival of CNS neurons during embryonic development.

Members of the collapsin/semaphorin family play an important role in creating the complex pattern of neuronal connectivity. Inhibition of growth cone motility by chick collapsin is mediated by the intraneuronal protein CRMP-62. We have now isolated four rat sequences that are highly related to chick CRMP-62. All four genes are expressed exclusively in the nervous system and primarily during development. Rat CRMP-2/TOAD-64 is most closely related to chick CRMP-62 and is the most widely expressed CRMP within the nervous system. Rat CRMP-1 and CRMP-4/rUlip are expressed during discrete periods of neuronal development and are not found in the adult nervous system. Rat CRMP-3 has a distinct distribution, being expressed transiently in developing spinal cord and selectively in the postnatal cerebellum. The differential expression of these genes suggests that CRMPs may transduce signals from different semaphorins and that semaphorins may regulate the plasticity of the adult nervous system.

Auxiliary structures of the cutaneous sensory nerve formations (SNF) are dependent on sensory innervation during their critical period of development. Denervation of mature cutaneous corpuscles results in survival of the terminal Schwann cells and the capsular structures which are probably responsible for successful reinnervation of the cutaneous SNF. In addition, the basal lamina tubes of Schwann cells are connected with the terminal Schwann cells and play an important role in the guidance of regrowing axons to their original targets. Long-lasting denervation causes atrophic changes of the terminal Schwann cells and alterations of their molecular equipment. These atrophic changes in the terminal Schwann cells may be responsible for erroneous reinnervation of cutaneous SNF. A population of the cutaneous Merkel cells surviving denervation may also serve as targets for regrowing sensory axons. The basal laminae of terminal Schwann cells are produced under control of the sensory terminals during maturation of cutaneous SNF. In adult animals, the basal laminae are capable of stimulating differentiation of migrated Schwann cells to the terminal Schwann cells without the presence of the sensory terminals. Nonspecific cholinesterase (nChE) is secreted by the terminal Schwann cells and is attached to their extracellular matrix. The synthesis of these molecules in adult animals is not influenced by the sensory terminals. However, the presence of nChE molecules is associated with living terminal Schwann cells. Fetal orthotopically grafted dorsal root ganglion (DRG) neurons have the ability to reinnervate cutaneous SNF of adult hosts. When cutaneous areas are denervated, axons from adjacent sensory nerves may extend collateral branches into this area. The capacity for such extension is dependent on: (1) type of sensory nerve ending, C and A delta fibers having significantly greater capacity than sensory axons of larger caliber; (2) age of the animal, immature animals generally showing a greater capacity for collateral sprouting; (3) the state of the adjacent axons, those already in a growth mode being more capable than "resting" ones; and (4) the regional and mechanical conditions at the site of denervation, hindpaw skin being much less extensively reinnervated by collateral fibers than that of the trunk.

Aims/background: Recent observations have shown that the glial scar resulting from a surgical lesion of the immature retina differs from elsewhere in the central nervous system, in that it permits the through growth and reconnection of regenerating axons. This study in the opossum examines in detail the development and reaction to injury of retinal glia at different developmental stages, and specifically examines the distribution of the gliosis related inhibitory molecule, chondroitin sulphate proteoglycan (CSPG), making comparisons with a control site of gliosis in the cerebral cortex., Methods: A linear slit was cut into the retina or cortex with a fine tungsten probe. After a variable time delay, immunocytochemistry of the resulting gliosis was employed to detect astrocytes with glial fibrillary acidic protein (GFAP), Müller cells with vimentin, and CSPG with CS-56 antibodies. GFAP was also used at different ages to examine the normal development of astrocytes in the retina of this species., Results: Astrocytes entered the retina 12 days after birth (P12), closely associated with blood vessels in the nerve fibre layer. In experiments at all ages studied, cellular continuity was re-established across the lesioned retina, which did not result in a significant astrocyte proliferation or CSPG expression. In contrast, cortical injury led to the development of a cystic cavity surrounded by astrocytes and CSPG. Müller cells expressed GFAP but not CSPG in the lesioned retina., Conclusion: Successful regrowth of ganglion cells through a retinal lesion may be partly the result of the scarcity of astrocytes in the retina, which results in minimal gliosis, or of their apparent inability to express inhibitory molecules.

Neuronal growth cones, the motile tips of elongating axons and dendrites, respond very precisely to cues encountered during pathfinding. During the past year, our knowledge about their sensory function, their integrative properties, and their motor function has advanced significantly. In particular, growth cone filopodia are currently being recognized for their prominent roles as sensors, transducers, and autonomous motor structures important for growth cone steering.

Repulsive guidance cues can steer neuronal growth cones during development and prevent mature axons from regenerating. We have identified a 100 kd glycoprotein in the chick brain that is a good candidate for a repulsive cue. Since it induces the collapse and paralysis of neuronal growth cones in vitro, we have named it collapsin. It is effective at concentrations of approximately 10 pM. The C-terminal half of collapsin contains a single immunoglobulin-like domain and an additional highly basic region. The N-terminal half of collapsin shares significant homology with fasciclin IV, a growth cone guidance protein in grasshopper. Recombinant collapsin causes sensory ganglion growth cones to collapse but not retinal ganglion cell growth cones. We propose that collapsin could serve as a ligand that guides specific growth cones by a motility-inhibiting mechanism.