Development of mucous cells in taste discs of anurans

Differentiation of the lingual taste discs (TDs) was studied in tadpoles of Rana esculenta, Hyla arborea and Pelobatesfuscus by means of LM, TEM, and SEM. The process of differentiation of mucous cells (MCs) within a TD anlage was investigated subsequently from 39th to 46th Gosner's developmental stage. In R. esculenta and P. fuscus, the MCs are distinguishable in TD anlages not earlier than at the 42nd tadpole Gosner's stage. At that stage TDs anlages are covered by a single layer of ordinary epithelial cells. Starting from the stage 44, tips of TDs progressively open outside and free surfaces of MCs can be distinguished as an area with numerous microfolds. At developmental stages 45 and 46, numerous electron-dense secretory granules can be seen in the supranuclear cytoplasm of MCs. The presence of mucopolysaccharides in H. arborea and P. fuscus was observed from the 44th-45th tadpole developmental stage.     

ORAL CHEMORECEPTOR ORGANS OF BULLFROG TADPOLES DURING METAMORPHOSIS*

Degeneration of the premetamorphic papillae and development of the fungiform papillae during metamorphosis of bullfrog tadpoles were investigated by electrophysiological and scanning electron microscopic methods. Premetamorphic papillae were observed during the early metamorphic stages, and these degenerated rapidly at about metamorphic stage 20. The anlage of the tongue appeared at about metamorphic stage 10, but the anlage of the fungiform papillae appeared at about metamorphic stage 18. The microvilli at the apex of the fungiform papillae were observed at about metamorphic stage 21. At metamorphic stage 24 the fungiform papillae had a similar structure to that of adult frogs. Taste responses were recorded from the glossopharyngeal nerve of the tadpole. The responses to 1 M sucrose and 0.01 M quinine hydrochloride could be observed at metamorphic stage 6 or later, though during stage 20 the responses were very weak. The response to 0.02 M ammonium chloride appeared at metamorphic stage 6, but disappeared at stage 20 and did not reappear later. These results indicate that the fungiform papillae become functional as chemo-receptor organs at about metamorphic stage 21 and that, before the fungiform papillae function, the premetamorphic papillae serve as chemoreceptor organs in the tadpole.     

Building sensory receptors on the tongue

Neurotrophins, neurotrophin receptors and sensory neurons are required for the development of lingual sense organs. For example, neurotrophin 3 sustains lingual somatosensory neurons. In the traditional view, sensory axons will terminate where neurotrophin expression is most pronounced. Yet, lingual somatosensory axons characteristically terminate in each filiform papilla and in each somatosensory prominence within a cluster of cells expressing the p75 neurotrophin receptor (p75NTR), rather than terminating among the adjacent cells that secrete neurotrophin 3. The p75NTR on special specialized clusters of epithelial cells may promote axonal arborization in vivo since its over-expression by fibroblasts enhances neurite outgrowth from overlying somatosensory neurons in vitro. Two classical observations have implicated gustatory neurons in the development and maintenance of mammalian taste buds—the early arrival times of embryonic innervation and the loss of taste buds after their denervation in adults. In the modern era more than a dozen experimental studies have used early denervation or neurotrophin gene mutations to evaluate mammalian gustatory organ development. Necessary for taste organ development, brain-derived neurotrophic factor sustains developing gustatory neurons. The cardinal conclusion is readily summarized: taste buds in the palate and tongue are induced by innervation. Taste buds are unstable: the death and birth of taste receptor cells relentlessly remodels synaptic connections. As receptor cells turn over, the sensory code for taste quality is probably stabilized by selective synapse formation between each type of gustatory axon and its matching taste receptor cell. We anticipate important new discoveries of molecular interactions among the epithelium, the underlying mesenchyme and gustatory innervation that build the gustatory papillae, their specialized epithelial cells, and the resulting taste buds.     

Rat gustatory neurons in the geniculate ganglion express glutamate receptor subunits

Taste receptor cells are innervated by primary gustatory neurons that relay sensory information to the central nervous system. The transmitter(s) at synapses between taste receptor cells and primary afferent fibers is (are) not yet known. By analogy with other sensory organs, glutamate might a transmitter in taste buds. We examined the presence of AMPA and NMDA receptor subunits in rat gustatory primary neurons in the ganglion that innervates the anterior tongue (geniculate ganglion). AMPA and NMDA type subunits were immunohistochemically detected with antibodies against GluR1, GluR2, GluR2/3, GluR4 and NR1 subunits. Gustatory neurons were specifically identified by retrograde tracing with fluorogold from injections made into the anterior portion of the tongue. Most gustatory neurons in the geniculate ganglion were strongly immunoreactive for GluR2/3 (68%), GluR4 (78%) or NR1 (71%). GluR1 was seen in few cells (16%). We further examined if glutamate receptors were present in the peripheral terminals of primary gustatory neurons in taste buds. Many axonal varicosities in fungiform and vallate taste buds were immunoreactive for GluR2/3 but not for NR1. We conclude that gustatory neurons express glutamate receptors and that glutamate receptors of the AMPA type are likely targeted to synapses within taste buds.     

Differential anatomical response of Norway spruce stem tissues to sterile and fungus infected inoculations

The anatomical defense responses in stems of Norway spruce (Picea abies) clones of different resistance to pathogenic fungi were characterized over time and distance from small mechanical wounds or wounds inoculated with the root rot fungus Heterobasidion annosum. Common responses for both treatments included division of ray parenchyma and other cells in the cambial zone, accumulation of phenolic inclusions in ray parenchyma cells, activation of phloem parenchyma (PP) cells, and formation of traumatic resin ducts (TDs) in the xylem. TD formation occurred synchronously from a tangential layer of cells, or symplasmic domain, within the zone of xylem mother cells. TD induction is triggered by a signal, which propagates a developmental wave in the axial direction at about 2.5 cm per day. TDs are formed at least 30 cm above single inoculations within 16–36 days after inoculation. The size and number of TDs is attenuated further away from the inoculation site, indicating a dose-dependent activity leading to TD development. Compared to sterile wounding, fungal inoculation gave rise to more and larger TDs in all clones, and multiple rows of TDs in weak clones. Fungal inoculation also induced the formation of more new PP cells, increasing the number of PP cells in the phloem in the year of inoculation up to 100%. TD and PP cell formation was greater in susceptible compared to resistant clones and after fungal versus sterile inoculation. Potential mechanisms responsible for this variable response are discussed.     

Skin epithelial transport and structural relationships in naturally metamorphosing Pelobates syriacus

The onset of active Na(+) transport and activated Cl(-) conductance (G(Cl)) across the skin epithelium of Pelobates syriacus was investigated during natural ontogenetic development. Structural features, including band three and Peanut lectin bindings were tested in parallel and structure-function relationships were attempted. The 22 specimens studied were divided into two tadpole, three juvenile, and two adult stages, corresponding to the Taylor-Kollros standard table, in accordance with external morphology of their developmental stage. Onset of transepithelial electrical potential and drop in conductance occurred abruptly, coinciding with metamorphosis climax of tadpoles into juveniles at about stage XXI of development. Amiloride-sensitive Na(+) transport occurred a little later at stage XXIII, followed by the appearance of activated Cl(-) conductance, G(Cl). Parallel structural examination showed that skin MR cells occurred upon metamorphosis, as the tadpole integument transformed into the adult epithelium and could be associated with the occurrence of activated G(Cl). It was not related temporally with the appearance of band three protein in MR cells. Our findings support the association of G(Cl) with MR cells, whereas band three may only be a corollary of G(Cl) and not necessarily essential for its manifestation.     

Interaction among different sensory units within a single fungiform papilla in the frog tongue

The possible interaction among different sensory units in the frog tongue was studied using several single papillae dually innervated by the medial and lateral branches of the glossopharyngeal (IXth) nerve. The afferent activity in one branch exposed to NaCl stimulation of the papilla revealed marked inhibition after antidromic electrical stimulation (100 Hz, 30 s, and 3 V) of the other branch. The degree of inhibition depended on the number of sensory responses observed in the electrically stimulated branch as well as the nature of the stimulated sensory units. Statistical analysis suggested that antidromic activation of gustatory units conducting the responses to NaCl and quinine and slowly adapting mechanosensitive units produced a large antidromic inhibition amounting to 19-25%, but that of gustatory units conducting the responses to acetic acid and rapidly adapting mechanosensitive units gave rise to only a slight inhibition. To examine the differential effects of these sensory units in antidromic inhibition, antidromic impulses were evoked by chemical stimulation of the adjacent papilla neuronally connected with the dually innervated papilla under study. Antidromic volleys of impulses elicited by NaCl or quinine stimulation produced a large inhibition of the afferent activity in the other branch, as induced by NaCl stimulation of the dually innervated papilla. Plausible mechanisms of synaptic interaction in peripheral gustatory systems are considered.     

Allosteric regulation of Bacillus subtilis threonine deaminase, a biosynthetic threonine deaminase with a single regulatory domain

The enzyme threonine deaminase (TD) is a key regulatory enzyme in the pathway for the biosynthesis of isoleucine. TD is inhibited by its end product, isoleucine, and this effect is countered by valine, the product of a competing biosynthetic pathway. Sequence and structure analyses have revealed that the protomers of many TDs have C-terminal regulatory domains, composed of two ACT-like subdomains, which bind isoleucine and valine, while others have regulatory domains of approximately half the length, composed of only a single ACT-like domain. The regulatory responses of TDs from both long and short sequence varieties appear to have many similarities, but there are significant differences. We describe here the allosteric properties of Bacillus subtilis TD ( bsTD), which belongs to the short variety of TD sequences. We also examine the effects of several mutations in the regulatory domain on the kinetics of the enzyme and its response to effectors. The behavior of bsTD can be analyzed and rationalized using a modified Monod-Wyman-Changeux model. This analysis suggests that isoleucine is a negative effector, and valine is a very weak positive effector, but that at high concentrations valine inhibits activity by competing with threonine for binding to the active site. The behavior of bsTD is contrasted with the allosteric behavior reported for TDs from Escherichia coli and Arabidopsis thaliana, TDs with two subdomains. We suggest a possible evolutionary pathway to the more complex regulatory effects of valine on the activity of TDs of the long sequence variety, e.g., E. coli TD.     

Size distributions of total airborne particles and bioaerosols in a municipal composting facility

Size distributions of total airborne particles and bioaerosols were measured in a full-scale composting facility, using an optical particle counter and an agar-inserted six-stage impactor, respectively. Higher concentrations of total airborne particles and bioaerosols were detected at a sampling location near the screening process preceded by the composting process than at sampling locations in the composting process. At the sampling location near the screening process, the concentrations of total airborne particles were approximately 10(8)particles/m3 at the size of 0.3 microm and 10(5)particles/m3 at 6.2 microm. The concentration of bioaerosols was about 10(4)CFU/m3 in each stage of 7.0 microm (1st stage), 7.0-4.7 microm (2nd), 4.7-3.3 microm (3rd), 3.3-2.1 microm (4th), 2.1-1.1 microm (5th) and 1.1-0.65 microm (6th). Most of submicron particles smaller than 1 microm among the total airborne particles were believed to originate from the ambient air.     

Evolution and development of brain sensory organs in molgulid ascidians

The ascidian tadpole larva has two brain sensory organs containing melanocytes: the otolith, a gravity receptor, and the ocellus, part of a photoreceptor. One or both of these sensory organs are absent in molgulid ascidians. We show here that developmental changes leading to the loss of sensory pigment cells occur by different mechanisms in closely related molgulid species. Sensory pigment cells are formed through a bilateral determination pathway in which two or more precursor cells are specified as an equivalence group on each side of the embryo. The precursor cells subsequently converge at the midline after neurulation and undergo cell interactions that decide the fates of the otolith and ocellus. Molgula occidentalis and M. oculata, which exhibit a tadpole larva with an otolith but lacking an ocellus, have conserved the bilateral pigment cell determination pathway. Programmed cell death (PCD) is superimposed on this pathway late in development to eliminate the ocellus precursor and supernumerary pigment cells, which do not differentiate into either an otolith or ocellus. In contrast to molgulids with tadpole larvae, no pigment cell precursors are specified on either side of the M. occulta embryo, which forms a tailless (anural) larva lacking both sensory organs, suggesting that the bilateral pigment cell determination pathway has been lost. The bilateral pigment cell determination pathway and superimposed PCD can be restored in hybrids obtained by fertilizing M. occulta eggs with M. oculata sperm, indicating control by a zygotic process. We conclude that PCD plays an important role in the evolution and development of brain sensory organs in molgulid ascidians.     

GABAergic modulation of primary gustatory afferent synaptic efficacy

Modulation of synaptic transmission at the primary sensory afferent synapse is well documented for the somatosensory and olfactory systems. The present study was undertaken to test whether GABA impacts on transmission of gustatory information at the primary afferent synapse. In goldfish, the vagal gustatory input terminates in a laminated structure, the vagal lobes, whose sensory layers are homologous to the mammalian nucleus of the solitary tract. We relied on immunoreactivity for the GABA-transporter, GAT-1, to determine the distribution of GABAergic synapses in the vagal lobe. Immunocytochemistry showed dense, punctate GAT-1 immunoreactivity coincident with the layers of termination of primary afferent fibers. The laminar nature and polarized dendritic structure of the vagal lobe make it amenable to an in vitro slice preparation to study early synaptic events in the transmission of gustatory input. Electrical stimulation of the gustatory nerves in vitro produces synaptic field potentials (fEPSPs) predominantly mediated by ionotropic glutamate receptors. Bath application of either the GABA(A) receptor agonist muscimol or the GABA(B) receptor agonist baclofen caused a nearly complete suppression of the primary fEPSP. Coapplication of the appropriate GABA(A) or GABA(B) receptor antagonist bicuculline or CGP-55845 significantly reversed the effects of the agonists. These data indicate that GABAergic terminals situated in proximity to primary gustatory afferent terminals can modulate primary afferent input via both GABA(A) and GABA(B) receptors. The mechanism of action of GABA(B) receptors suggests a presynaptic locus of action for that receptor.     

Pattern and rate of ovary differentiation with reference to somatic development in anuran amphibians

The rate of somatic development of anuran amphibians is only roughly correlated with the rate of gonad differentiation and varies among species. The somatic stage of a tadpole often does not reflect its age, which seems to be crucial for gonad differentiation rate. We compared the morphology and differentiation of developing ovaries at the light and electron microscopy level, with reference to somatic growth and age of a female. Our observations were performed on 12 species of six families (Rana lessonae, R. ridibunda, R. temporaria, R. arvalis, R. pipiens, R. catesbeiana, Bombina bombina, Hyla arborea, Bufo bufo. B. viridis, Xenopus laevis, Pelobates fuscus) and compared with the results obtained by other authors. This allowed us to describe the unified pattern of anuran female gonad differentiation. Ovary differentiation was divided into 10 stages: I-III, undifferentiated gonad; IV, sexual differentiation; V, first nests of meiocytes; VI, first diplotene oocytes; VII-IX, increasing number of diplotene oocytes and decreasing number of oogonia and nests; X, fully developed ovary composed of diplotene oocytes with rudimental patches of oogonia. We distinguished three types of ovary differentiation rate: basic (most species), retarded (genus Bufo), and accelerated (green frogs of the subgenus Pelophylax genus Rana).     

Neuromodulation of Transduction and Signal Processing in the End Organs of Taste

Chemical synapses transmit gustatory signals from taste receptorcells to sensory afferent axons. Chemical (and electrical) synapsesalso provide a lateral pathway for cells within the taste budto communicate. Lateral synaptic pathways may represent someform of signal processing in the peripheral end organs of taste.Efferent synaptic input may also regulate sensory transductionin taste buds. To date, the synaptic neurotransmitter(s) orneuromodulator(s) released at chemical synapses in taste budshave not been identified unambiguously. This paper summarizesthe attempts that have been made over the past 40 years to identifythe neuroactive substances acting at taste bud synapses. Wereview the four traditional criteria for identifying chemicaltransmitters elsewhere in the nervous system—localization,uptake/degradation, release and physiological actions—andapply these criteria to neuroactive substances in taste buds.The most complete evidence to date implicates serotonin as aneuromodulator of taste transduction in the end organs. However,studies also suggest that adrenergic, cholinergic and peptidergicneurotransmission may be involved in taste buds. Chem. Senses21: 353–365, 1996.     

Changes in numbers and dimensions of glomeruli during metamorphosis of Pelobates syriacus (Anura; Pelobatidae)

Changes in glomerular numbers and volume were followed throughout five ontogenetic stages of Pelobates syriacus (Anura, Pelobatidae). The number of glomeruli increased markedly between the 2-legged and the 4-legged tadpole stages. In the post-metamorphic stages the number of glomeruli was about twice their number during the tadpole stages. Glomerular volume peaked during the 2-legged tadpole stage, when the tadpole reached its maximal size, and dropped thereafter. There is some evidence to indicate that the number of glomeruli in post-metamorphic P. syriacus increased during growth thus indicating perhaps an increase in their functional importance. The number was still lower than that reported for some other anurans, more similar to the numbers given for representative urodeles, however, the latter have considerably larger glomeruli. Two main factors appear to be involved in regulating numbers and volume of glomeruli in frogs. The ontogenetic changes during metamorphosis affect both glomerular numbers and volume, whereas post-metamorphic growth affected only the glomerular numbers.     

Ascidian embryos as a model system to analyze expression and function of developmental genes

Ascidians have served as an appropriate experimental system in developmental biology for more than a century. The fertilized egg develops quickly into a tadpole larva, which consists of a small number of organs including epidermis, central nervous system with two sensory organs, endoderm and mesenchyme in the trunk, and notochord and muscle in the tail. This configuration of the ascidian tadpole is thought to represent the most simplified and primitive chordate body plan. Their embryogenesis is simple, and lineage of embryonic cells is well documented. The ascidian genome contains a basic set of genes with less redundancy compared to the vertebrate genome. Cloning and characterization of developmental genes indicate that each gene is expressed under discrete spatio-temporal pattern within their lineage. In addition, the use of various molecular techniques in the ascidian embryo system highlights its advantages as a future experimental system to explore the molecular mechanisms underlying the expression and function of developmental genes as well as genetic circuitry responsible for the establishment of the basic chordate body plan. This review is aimed to highlight the recent advances in ascidian embryology.     

Non-ependymal cilia in the habenulae and the interpeduncular nucleus of the frog tadpole

Summary Cilia of the 9+2 pattern are found electron microscopically in nonependymal cells of the habenulae and the interpeduncular nucleus of the tadpole of Rana esculenta at an early stage of development (8 mm length, head to tip of tail). A comparison is made between these and the ependymal and sensory cilia in the same specimens. The cilia project into the neuropil emerging from a perikaryon rich in free ribosomes and displaying a prominent Golgi apparatus. These perikarya contain dense core vesicles. Synapses with vesicles of the clear spherical type have been observed along the ciliary shaft. On a purely morphologic basis the authors hypothesize that these cilia, at least in this early ontogenetic stage, may extend considerably the conducting surface of the cell and represent a sensory structure which could be stimulated by terminal processes belonging to distantly located cells. In addition, they could also be involved in the trophic exchange of material with the adjacent structures.     

Tarsal sensory complex of the red chicken mite Dermanyssus gallinae (Acari: Dermanyssidae)

The tarsal sensory complex of the red chicken mite Dermanyssus gallinae is situated on dorsal surface of each fore leg near the claw. It comprises 28 sensilla of 5 morphological types: 4 SW-UP (single-wall upper-pore) (gustatory organs), 8 SW-WP (single-wall wall-pore) (olfactory organs), 8 DW-WP (double-wall wall-pore) of two subtypes (thermo-chemoreceptory organs), 6 NP-TB (no pore--tubular body) (tactile organs), and 2 reduced sensilla. No sex or stage dimorphism was revealed. Morphological data point to the fact that tarsal sensory complex of the red chicken mite is mainly an organ detecting temperature changes and olfactory stimula.