Lingual Epithelium Research Articles

Cloning and expression of a mouse member of the PLUNC protein family exclusively expressed in tongue epithelium

Palate, lung, and nasal epithelium clone (Plunc, now renamed Splunc1) is a small secreted protein expressed in the oropharynx and upper airways of humans, mice, rats, and cows. This protein is structurally homologous to known mediators of host defense against gram-negative bacteria. We have characterized the genomic sequence and expression of a novel but closely related gene from rodents, which we call splunc5. Mouse Splunc5 sequence is 60% identical to mouse Splunc1. The genes also share highly conserved genomic elements including intron–exon structure and intronic sequence. Strikingly, splunc5 is expressed exclusively in the interpapillary epithelium of the tongue's dorsal surface. By comparing the expression profiles of splunc5, splunc1, and a third related sequence, lplunc1, in mice, we show that these genes are expressed in unique domains along a continuous corridor of oral, lingual, pharyngeal, and respiratory epithelia. This expression pattern is consistent with the hypothesis that these proteins protect epithelial surfaces colonized by potentially pathogenic microorganisms.

Na+-H+ exchange activity in taste receptor cells.

mRNA for two Na(+)-H(+)-exchanger isoforms 1 and 3 (NHE-1 and NHE-3) was detected by RT-PCR in fungiform and circumvallate taste receptor cells (TRCs). Anti-NHE-1 antibody binding was localized to the basolateral membranes, and the anti-NHE-3 antibody was localized in the apical membranes of fungiform and circumvallate TRCs. In a subset of TRCs, NHE-3 immunoreactivity was also detected in the intracellular compartment. For functional studies, an isolated lingual epithelium containing a single fungiform papilla was mounted with apical and basolateral sides isolated and perfused with nominally CO(2)/HCO(3)(-)-free physiological media (pH 7.4). The TRCs were monitored for changes in intracellular pH (pH(i)) and Na(+) ([Na(+)](i)) using fluorescence ratio imaging. At constant external pH, 1) removal of basolateral Na(+) reversibly decreased pH(i) and [Na(+)](i); 2) HOE642, a specific blocker, and amiloride, a nonspecific blocker of basolateral NHE-1, attenuated the decrease in pH(i) and [Na(+)](i); 3) exposure of TRCs to basolateral NH(4)Cl or sodium acetate pulses induced transient decreases in pH(i) that recovered spontaneously to baseline; 4) pH(i) recovery was inhibited by basolateral amiloride, 5-(N-methyl-N-isobutyl)-amiloride (MIA), 5-(N-ethyl-N-isopropyl)-amiloride (EIPA), HOE642, and by Na(+) removal; 5) HOE642, MIA, EIPA, and amiloride inhibited pH(i) recovery with K(i) values of 0.23, 0.46, 0.84, and 29 microM, respectively; and 6) a decrease in apical or basolateral pH acidified TRC pH(i) and inhibited spontaneous pH(i) recovery. The results indicate the presence of a functional NHE-1 in the basolateral membranes of TRCs. We hypothesize that NHE-1 is involved in sour taste transduction since its activity is modulated during acid stimulation.

Gustatory neurons derived from epibranchial placodes are attracted to, and trophically supported by, taste bud-bearing endoderm in vitro

Taste buds are multicellular receptor organs innervated by the VIIth, IXth, and Xth cranial nerves. In most vertebrates, taste buds differentiate after nerve fibers have reached the lingual epithelium, suggesting that nerves induce taste buds. However, under experimental conditions, taste buds of amphibians develop independently of innervation. Thus, rather than being induced by nerves, the developing taste periphery likely regulates ingrowing nerve fibers. To test this idea, we devised a culture approach using axolotl embryos. Gustatory neurons were generated from cultured epibranchial placodes, and when cultured alone, axon outgrowth was random over 4 days, a time period coincident with axon growth to the periphery in vivo. In contrast, cocultures of placodal neurons with oropharyngeal endoderm (OPE), the normal taste bud-containing target for these neurons, resulted in neurite growth toward the target tissue. Unexpectedly, placodal neurons also grew toward flank ectoderm (FE), which these neurons do not encounter in vivo. To compare further the impact of OPE and FE explants on gustatory neurons, cocultures were extended and examined at 6, 8, and 10 days, when, in vivo, placodal fibers have innervated the epithelium but prior to taste bud formation, when taste buds have differentiated and are innervated, and when the mouth has opened and larvae have begun to feed, respectively. The behavior of placodal axons with respect to target type did not differ between OPE and FE cocultures at 6 days. However, by 8 days, differences in axonal outgrowth were observed with respect to target type, and these differences were enhanced by 10 days in vitro. Most clearly, exuberant placodal fibers grew in 10-day OPE cocultures, and numerous neurites had invaded OPE explants by this time, whereas gustatory neurites were sparse in FE cocultures, and rarely approached and almost never contacted FE explants. Thus, embryonic endoderm destined to give rise to taste buds specifically attracts its innervation early in development, as placodal neurons send out axons. Later, when gustatory axons synapse with differentiated taste buds in vivo, the OPE provides trophic support for cultured gustatory neurons.

Immunohistochemical detection of the expression of keratin 14 in the lingual epithelium of rats during the morphogenesis of filiform papillae

An immunofluorescence study of the expression of keratin 14 (K14) during the formation of filiform papillae was performed and the progress of keratinization of the epithelium of the rat tongue was monitored on semi-ultrathin sections by laser-scanning microscopy. Differential interference contrast (DIC) images were also examined to provide details of histology and cell morphology. No cells with immunoreactivity specific for K14 were detected on the lingual epithelium of foetuses on embryonic days 12 and 16 (E12 and E16), when the lingual epithelium was composed of a single layer or several layers of cuboidal cells. Immunoreactivity specific for K14 was detected first on basal and suprabasal keratinocytes of the dorsal epithelium of the tongue of new-borns on postnatal day 0 (P0) and was conspicuous in juveniles on P14. The immunoreactivity was particularly strong on the basal and suprabasal keratinocytes along the connective tissue papillae. The immunoreactivity extended over the entire cytoplasm but was not detected in the nucleus. The lingual epithelium was composed of stratified squamous cells and the rounded rudiments of filiform papillae were compactly arranged at equal intervals, for the most part, and the spaces between them were narrow and indistinct. Immunostaining of K14 was distinct on basal and suprabasal keratinocytes of the filiform papillar area of tongues of juveniles on P21, when the filiform papillae were conical. The spaces between them were relatively wide and, as a result, interpapillar cell columns were clearly visible. Immunoreactivity specific for K14 in the basal and suprabasal keratinocytes of the interpapillar cell columns was recognizable but was weaker than that in cells of papillar cell columns. The thickness of the epithelium in papillar and interpapillar areas increased gradually with the development of filiform papillae. However, sizes of basal and suprabasal keratinocytes remained almost unchanged during this process. These results suggest that the basal and suprabasal keratinocytes of the filiform papillar area proliferate with the initiation of the morphogenesis of filiform papillae and the keratinization of the epithelium. In addition, it appears that, after P14, the basal and suprabasal keratinocytes of the interpapillar area proliferate to supply the keratinocytes of the expanding interpapillar regions.

Adenylyl cyclase expression and modulation of cAMP in rat taste cells.

cAMP is a second messenger implicated in sensory transduction for taste. The identity of adenylyl cyclase (AC) in taste cells has not been explored. We have employed RT-PCR to identify the AC isoforms present in taste cells and found that AC 4, 6, and 8 are expressed as mRNAs in taste tissue. These proteins are also expressed in a subset of taste cells as revealed by immunohistochemistry. Alterations of cAMP concentrations are associated with transduction of taste stimuli of several classes. The involvement of particular ACs in this modulation has not been investigated. We demonstrate that glutamate, which is a potent stimulus eliciting a taste quality termed umami, causes a decrease in cAMP in forskolin-treated taste cells. The potentiation of this response by inosine monophosphate, the lack of response to d-glutamate, and the lack of response to umami stimuli in nonsensory lingual epithelium all suggest that the cAMP modulation represents umami taste transduction. Because cAMP downregulation via ACs can be mediated through Galpha(i) proteins, we examined the colocalization of the detected ACs with Galpha(i) proteins and found that 66% of AC8 immunopositive taste cells are also positive for gustducin, a taste-specific Galpha(i) protein. Whether AC8 is directly involved in signal transduction of umami taste remains to be established.

The epithelia of the protrusible tongue of Eurycea longicauda guttolineata (Hoolbrook 1838) (Urodela: Plethodontidae).

In this study the lingual and sublingual glands, the lingual stem and the epithelial surface of the protrusible secondary tongue were investigated by light, scanning and transmission electron microscopy. The quality of the secretions of the epithelia was characterized histochemically. The lingual epithelium is formed by superficial (pavement) and goblet cells and at the margin of the tongue pad are also regions covered by ciliated cells. On the dorsal part of the tongue there are goblet cells of type A with mainly acidic secretions and of type B containing neutral secretions. Most of the goblet cells on the ventral side of the tongue (hypoglottis) show a strong alcian blue/PAS positive reaction (type I) and some produce neutral secretions (type II). The glandular cells of the lingual gland react positively to alcian blue and PAS in the apical region of the gland. In contrast there is only alcian blue-positive staining in the basal part of the gland. The size and complexity of the inclusion bodies of the secretory granules increase in a basal direction. In addition, there are ciliated cells in the glandular epithelium. Although the epithelium of the lingual stem is thin, it is double-layered. The cell types observed in this region are identical to those of the ventral part of the protrusible tongue. At the margin of the sublingual gland are trough-like structures. In the center, tubular parts are observed. The cells of this gland are stain strongly with alcian blue (pH 1.0) mainly in the basal part of the gland. The results of this are compared to the tongue pad and the lingual gland of Salamandra salamandra and Ambystoma mexicanum.

Cyclopamine and jervine in embryonic rat tongue cultures demonstrate a role for Shh signaling in taste papilla development and patterning: fungiform papillae double in number and form in novel locations in dorsal lingual epithelium

From time of embryonic emergence, the gustatory papilla types on the mammalian tongue have stereotypic anterior and posterior tongue locations. Furthermore, on anterior tongue, the fungiform papillae are patterned in rows. Among the many molecules that have potential roles in regulating papilla location and pattern, Sonic hedgehog (Shh) has been localized within early tongue and developing papillae. We used an embryonic, tongue organ culture system that retains temporal, spatial, and molecular characteristics of in vivo taste papilla morphogenesis and patterning to study the role of Shh in taste papilla development. Tongues from gestational day 14 rat embryos, when papillae are just beginning to emerge on dorsal tongue, were maintained in organ culture for 2 days. The steroidal alkaloids, cyclopamine and jervine, that specifically disrupt the Shh signaling pathway, or a Shh-blocking antibody were added to the standard culture medium. Controls included tongues cultured in the standard medium alone, and with addition of solanidine, an alkaloid that resembles cyclopamine structurally but that does not disrupt Shh signaling. In cultures with cyclopamine, jervine, or blocking antibody, fungiform papilla numbers doubled on the dorsal tongue with a distribution that essentially eliminated inter-papilla regions, compared with tongues in standard medium or solanidine. In addition, fungiform papillae developed on posterior oral tongue, just in front of and beside the single circumvallate papilla, regions where fungiform papillae do not typically develop. The Shh protein was in all fungiform papillae in embryonic tongues, and tongue cultures with standard medium or cyclopamine, and was conspicuously localized in the basement membrane region of the papillae. Ptc protein had a similar distribution to Shh, although the immunoproduct was more diffuse. Fungiform papillae did not develop on pharyngeal or ventral tongue in cyclopamine and jervine cultures, or in the tongue midline furrow, nor was development of the single circumvallate papilla altered. The results demonstrate a prominent role for Shh in fungiform papilla induction and patterning and indicate differences in morphogenetic control of fungiform and circumvallate papilla development and numbers. Furthermore, a previously unknown, broad competence of dorsal lingual epithelium to form fungiform papillae on both anterior and posterior oral tongue is revealed.

Histochemical and ultrastructural features of the lingual epithelium of the rat snake (Elaphe climacophora)

The histological, ultrastructural and histochemical characteristics of the lingual epithelium of the rat snake (Elaphe climacophora) were investigated by light and transmission electron microscopy. The cells in the beta-layer of the epithelium of the bifurcated apex were filled with beta-keratin fibers and an amorphous matrix. Round projections covering the surface of the epithelial cells, namely, microfacets which contained pale granules, were clearly visible on the outer faces of Oberhäutchen cells on the epithelium, and they were identified as fine granules filled with lipid. These granules might play an important role as a coating on the surface of the bifurcated lingual apex. The lipid on the surface of the lingual apex might also serve to trap and retain odorant molecules. Keratohyalin-like granules were distributed within the a-layer of the epithelium of the bifurcated apex of the tongue in the resting phase and cellular interdigitation was well developed in this region. Evidence of a shedding line was apparent under the light microscope in the cleft between outer and inner epithelial generations. The epithelial surface of the body of the tongue appeared suitable for retention of odorant and other molecules.

Cathelicidin antimicrobial peptides are expressed in salivary glands and saliva.

The expression of antimicrobial peptides at epithelial surfaces such as skin, lung, and intestine is thought to provide protection against infection. Cathelicidin antimicrobial peptides are essential for the protection of skin against invasive bacterial infection. To determine if cathelicidins are also present in the oral cavity, we examined the expression of both mRNA and protein in mice and human saliva. The murine cathelicidin (CRAMP) was detected in the adult by reverse-transcription/polymerase chain-reaction (RT-PCR), and in embryonic, newborn, and adult tissues by in situ hybridization and immunohistochemistry. CRAMP mRNA and protein were localized to the salivary glands, specifically in acinar cells of the submandibular gland and palatine minor glands, as well as in lingual epithelium and palatal mucosa. In man, the human cathelicidin LL-37 was detected in human saliva by Western blotting. These results indicate that cathelicidins are present in the salivary system, in some oral epithelia, and in saliva, contributing to broad-spectrum defense of the oral cavity.

Comparative Analysis of Telomere Lengths and Erosion with Age in Human Epidermis and Lingual Epithelium

We investigated progressive telomere shortening in normal human epidermis and lingual epithelium during aging, and attempted, in particular, to ascertain whether the telomere shortening that accompanies aging occurs at the same rate in different tissues. We studied telomeric DNA integrity, and estimated annual telomere loss, in 52 specimens of epidermis and 48 specimens of lingual epithelium collected at autopsy from subjects who had died at ages between 0 and 101 y. Most of the DNA samples were measured twice by southern blot hybridization. In addition, the correlation between telomere lengths in the two types of tissues was examined. The telomere reduction rates in epidermis and lingual epithelium were 36 bp and 30 bp per y, respectively, and these were significantly different. The rates obtained by the second measurements in epidermis and lingual epithelium were 39 and 32 bp per y, respectively, and these were also significantly different. The mean telomere lengths in the epidermis of eight neonates and the lingual epithelium of seven neonates were 13.2+/-1.0 and 13.8+/-1.0 kb, respectively. Comparison of telomere lengths in the two tissues for 41 paired samples showed that the mean telomere length in the epidermis (10.7+/-2.3 kb) was less than that in the lingual epithelium (12.4+/-2.5 kb); however, statistical analysis revealed a very significant relationship between epidermal and lingual epithelial telomere length (r=0.842, p<0.0001). These results indicate that the telomeres in epidermis and lingual epithelium are characterized by tissue-specific loss rates.

Adrenergic signalling between rat taste receptor cells.

In taste buds, synaptic transmission is traditionally thought to occur from taste receptor cells to the afferent nerve. This communication reports the novel observation that taste receptor cells respond to adrenergic stimulation. Noradrenaline application inhibited outward potassium currents in a dose-dependent manner. This inhibition was mimicked by the beta agonist isoproterenol and blocked by the beta antagonist propranolol. The alpha agonists clonidine and phenylephrine both inhibited the potassium currents and elevated intracellular calcium levels. Inwardly rectifying potassium currents were unaffected by adrenergic stimulation. Experiments using the RT-PCR technique demonstrate that lingual epithelium expresses multiple alpha (alpha1a, alpha1b, alpha1c, alpha1d, alpha2a, alpha2b, alpha2c) and beta (beta1, beta2) subtypes of adrenergic receptors, and immunocytochemistry localized noradrenaline to a subset of taste receptor cells. Collectively, these data imply strongly that adrenergic transmission within the taste bud may play a paracrine role in taste physiology.

Novel Expression of Vascular Cell Adhesion Molecule-1 (CD106) by Squamous Epithelium in Experimental Acute Graft- versus-Host Disease

Vascular cell adhesion molecule-1 (VCAM-1; CD106), the receptor for VLA-4, is an important mediator of adhesive and co-stimulatory interactions that govern cutaneous immune responses. Initial studies designed to elucidate temporal aspects of endothelial adhesion molecule induction in murine acute graft-versus-host disease (aGVHD) revealed unexpected and novel VCAM-1 expression by cutaneous and mucosal epithelial cells. Immunohistochemical techniques confirmed VCAM-1 staining as early as 7 days after transplantation in a distinctive subpopulation of squamous epithelial cells that normally occupy focal domains within the epidermal basal cell layer, the follicular infundibulum, and the dorsal lingual epithelium. Specifically, VCAM-1 expression was intimately associated with rete ridge-like prominences in footpad epidermis and in dorsal lingual epithelium. VCAM-1, as evaluated by serial section-labeling techniques, was preferentially expressed at sites of early epithelial infiltration by CD4(+) T cells. Western blot analysis confirmed expression of the 110-kd isoform of VCAM-1 in epithelium isolated from aGVHD animals, and immunoelectron microscopy demonstrated VCAM-1 reactivity restricted exclusively to epithelial cell plasma membranes. It is concluded that VCAM-1 is selectively expressed by discrete squamous epithelial subpopulations in murine aGVHD. As such, VCAM-1 may play a previously unrecognized role in mediating interactions between donor effector T lymphocytes and host epithelial cell targets.

Morphological study of lingual papillae in the fetal cat at mid-gestation, as revealed by light and scanning electron microscopy.

Tongues were removed from five cat fetuses, after approximately 1 month of gestation, for examination by light and scanning electron microscopy. Rudiments of filiform papillae were visible over the entire dorsal surface of the tongue. The epithelium of the dorsal surface of the tongue was of the stratified cuboidal type. No evidence of keratinization was detected anywhere in the entire dorsal lingual epithelium. Rudiments of fungiform papillae were recognizable only at the lingual apex; none were recognizable on other parts of the dorsal surface of the tongue at this stage. By contrast, rudiments of circumvallate and foliate papillae were already distinguishable from the filiform papillae. Differences between these results and those obtained previously in rats and mice are discussed.

Developmental patterning of the circumvallate papilla.

Organogenesis is regulated by the sequential and reciprocal interactions between epithelial and mesenchymal tissues. Many molecules, including growth factors, transcription factors, extracellular matrices, cell surface receptors, and matrix degrading enzymes, have been found to be involved in this process. To investigate the molecular mechanism responsible for morphogenesis of the circumvallate papilla/von Ebners' gland complex, we examined the expression patterns of selected cell adhesion molecules, extracellular matrix molecules, innervation and cell division in the circumvallate papilla of mouse embryos from embryonic day 11.5 (E11.5) to E14. At E11.5-E13.5, the lingual epithelium, the site where the circumvallate papilla will develop, is negative for BrdU labeling. At E14-E15, we detected cell division in the papillary area, especially in the epithelial invagination where von Ebners' minor salivary gland will form. The basement membrane component, laminin, is expressed as a continuous thin line separating the epithelia from the underlying mesenchyme, but it is broadly and strongly expressed in the area wherethe nervefibers penetrate into the circumvallate papilla. At the E12-E12.5 stage of development, the epithelial thickening shows intense E-cadherin staining in the superficial and basal layers, but weak E-cadherin staining in the suprabasal layer. E-cadherin is strongly expressed, but appears dispersed among the basal layer of lingual epithelium, the site where nerve fibers will innervate. At E13, nerve fibers reach the circumvallate papilla. These nerve fibers penetrate into and split the epithelial cell mass into two stalks which will later differentiate to form the von Ebners' gland. These results suggest that 1) the formation of the circumvallate papilla does not initially depend on cell division, 2) cell migration likely plays a major role during circumvallate placode formation, 3) E-cadherin and laminin may play a role in nerve guidance and 4) innervation impacts the final morphogenesis of the circumvallate papilla.

Evolution of the structure and function of the vertebrate tongue.

Studies of the comparative morphology of the tongues of living vertebrates have revealed how variations in the morphology and function of the organ might be related to evolutional events. The tongue, which plays a very important role in food intake by vertebrates, exhibits significant morphological variations that appear to represent adaptation to the current environmental conditions of each respective habitat. This review examines the fundamental importance of morphology in the evolution of the vertebrate tongue, focusing on the origin of the tongue and on the relationship between morphology and environmental conditions. Tongues of various extant vertebrates, including those of amphibians, reptiles, birds and mammals, were analysed in terms of gross anatomy and microanatomy by light microscopy and by scanning and transmission electron microscopy. Comparisons of tongue morphology revealed a relationship between changes in the appearance of the tongue and changes in habitat, from a freshwater environment to a terrestrial environment, as well as a relationship between the extent of keratinization of the lingual epithelium and the transition from a moist or wet environment to a dry environment. The lingual epithelium of amphibians is devoid of keratinization while that of reptilians is keratinized to different extents. Reptiles live in a variety of habitats, from seawater to regions of high temperature and very high or very low humidity. Keratinization of the lingual epithelium is considered to have been acquired concomitantly with the evolution of amniotes. The variations in the extent of keratinization of the lingual epithelium, which is observed between various amniotes, appear to be secondary, reflecting the environmental conditions of different species.

Electrophysiological properties of the tongue epithelium of the toad Bufo marinus.

The dorsal lingual epithelium from the tongue of the toad Bufo marinus was mounted in an Ussing-type chamber, and the short-circuit current (I(sc)) was measured using a low-noise voltage clamp. With NaCl Ringer bathing the mucosal and serosal surfaces of the isolated tissue, an outwardly directed (mucosa-positive) I(sc) was measured that averaged -10.71+/-0.82 microA cm(-2) (mean +/- S.E.M., N=24) with a resistance of 615+/-152 Omega cm(2) (mean +/- S.E.M., N=10). Substitution of chloride with sulfate as the anion produced no significant change in I(sc). Fluctuation analysis with either NaCl or Na(2)SO(4) Ringer bathing both sides of the tissue revealed a spontaneous Lorentzian component, suggesting that the I(sc) was the result of K(+) secretion through spontaneously fluctuating channels in the apical membrane of the epithelium. This hypothesis was supported by the reversible inhibition of I(sc) by Ba(2+) added to the mucosal Ringer. Analysis of the kinetics of Ba(2+) inhibition of I(sc) indicates that there might be more than one type of K(+) channel carrying the I(sc). This hypothesis was supported by power spectra obtained with a serosa-to-mucosa K(+) gradient, which could be fitted to two Lorentzian components. At present, the K(+) secretory current cannot be localized to taste cells or other cells that might be associated with the secretion of saliva or mucus. Nonetheless, the resulting increase in [K(+)] in fluid bathing the mucosal surface of the tongue could presumably affect the sensitivity of the taste cells. These results contrast with those from the mammalian tongue, in which a mucosa-negative I(sc) results from amiloride-sensitive Na(+) transport.

Les troubles du goût

Purpose. – This review focuses on the present state of the question about taste disorders with reference to their associated factors, diagnostic methods, and potential effects. Current knowledge and key points. – Taste disorders may be induced by many drugs and are associated to a number of acute or chronic diseases. Patients may be asked about their taste complaints, and taste thresholds may be determined by electrogustometry or chemical gustometry. Taste impairment may provide a good indicator to the course of some diseases such as diabetes mellitus in which hypogeusia predicts occurrence of degenerative complications. Dysgeusia may induce nutritional disorders and contribute to wasting in chronic liver disease, cancer, or human immunodeficiency virus infected patients. Mechanisms involved in dysgeusia are more than one in a patient. Taste disturbance may be secondary to a variety of causes that include zinc deficiency, lesions of the lingual epithelium, neurological impairment, and a pharmacological effect. Future prospects and projects. – A better understanding of the transduction mechanisms of the gustatory signal and the main pathogenic factors involved in dysgeusia may possibly improve the follow-up of the concerned patients notably in terms of nutritional status.

Ganglion cells and topographically related nerves in the vallate papilla/von Ebner gland complex.

Ganglion cells and topographically related nerves in the vallate papilla/von Ebner gland complex were investigated in rat tongue by cytochemical, immunocytochemical, and ultrastructural methods to evaluate the possible presence of different neuronal subpopulations. Immunostaining for neurofilaments and protein gene product 9.5 revealed ganglionic cell bodies and nerve fibers. A large part of the neurons were positive at immunostaining for neuronal nitric oxide synthase (NOS), vesicular acetylcholine transporter (VAChT), or vasoactive intestinal peptide (VIP). A small subset of nerve fibers revealed immunoreactivity for cholecystokinin. Axons traveling under the lingual epithelium were evidenced by their content of calcitonin gene-related peptide (CGRP) or substance P (SP). Cell bodies positive for SP or CGRP were not detected. Using methods of co-localization, three different neuronal classes were detected. The main population was composed of AChE/NADPH-diaphorase (NADPHd)-positive cells. Small groups of acetylcholine esterase (AChE)-positive/NADPHd-negative cells were visible. Isolated neurons were AChE-negative/NADPHd-positive. The results of co-localization experiments for VAChT/NOS were consistent with those obtained by cytochemical co-localization of AChE and NADPHd. Experiments of co-localization for peptidergic and nitrergic structures revealed CGRP- and SP-immunoreactive fibers in the vallate papilla/von Ebner gland ganglion. In conclusion, the results demonstrated in the VP/VEG complex peptidergic, cholinergic, and nitrergic neurons. The presence of different neuronal subclasses suggests that a certain degree of functional specialization may exist.

Expression of NeuroD in the mouse taste buds.

NeuroD, a basic helix-loop-helix transcription factor, has been shown to play a role in the differentiation of neurons, olfactory cells, and neuroendocrine tissues. Since the taste buds have characteristics of typical paraneurons, we examined the expression of NeuroD in the taste buds of mice. By RT-PCR analysis, NeuroD mRNA was found to be expressed in the epithelium of circumvallate papillae-containing taste buds, but not in the lingual epithelium lacking them. NeuroD immunoreactivity was detected in a subset of taste bud cells in the circumvallate, foliate, and fungiform papillae and in the soft palate. NeuroD-expressing cells had a spindle-like shape, first appeared at postnatal day 3, and increased in number during postnatal development. After bisection of the glossopharyngeal nerves, NeuroD-expressing cells decreased in number at day 4 and disappeared from the trench wall of the circumvallate papillae by day 14. A few NeuroD-expressing taste buds reappeared at postoperative day 28. Denervation and regeneration experiments showed that expression of NeuroD in the taste buds was dependent upon gustatory innervation. Double immunolabeling with gustducin or with neural cell adhesion molecule (NCAM) showed that NeuroD-expressing cells did not express NCAM, but did express gustducin. These results suggest that NeuroD is expressed in a mature cell type, type-II cells, but not in type-III cells.

Ultrastructure of the lingual epithelium of adult scincine lizard Chalcides ocellatus

In the present investigation, the histological and histochemical characteristics, as well as the ultrastructure of the lingual epithelium of the adult lizard Chalcides ocellatus, were investigated by light microscopy (LM) and scanning electron microscopy (SEM). Histological examination revealed taste buds and sensory papillae on the tongue tip and on the foretongue. The papillae were covered by stratified squamous epithelium and the core of the papillae is penetrated by connective tissues, pigment cells and striated muscle fibers. The acinar lingual glands appeared between the lateral sides of the papillae with increased number posteriorly. These glands stained purple with AB-PAS dorsally, and pink, with some purple cells, ventrally. SEM observations revealed three different types of lingual papillae, which were widely distributed over most dorsal surface of the tongue. The epithelial cells covering the surface of each papilla include smooth surface epithelial cells at the distal part of the papillae (free border) and cells with microridges interspersed with micropores at the intermediate portion of the papillae. Cells with densely distributed tall microvilli bearing extracellular mucus were noticed at the proximal portion of the papillae (near the base). Most of the surface of the bifurcated part of the tongue was relatively smooth with pitted and wrinkled surfaces (characteristic marks of the keratinized epithelium). Also the correlations between the structure and function were discussed in details.