Circumvallate Taste Buds Research Articles

The effects of sialoadenectomy and exogenous EGF on taste bud morphology and maintenance.

Taste buds on the dorsal tongue surface are continually bathed in saliva rich in epidermal growth factor (EGF). In the following experiment, taste bud number and morphology were monitored following submandibular and sublingual salivary gland removal (sialoadenectomy), to determine if EGF plays a role in the maintenance and formation of taste buds. Adult male rats were divided into four groups: sialoadenectomized (SX, n = 4); sialoadenectomized with EGF replacement (SX + EGF, n = 5); sham-operated (SH, n = 4); and sham-operated with exogenous EGF (SH + EGF, n = 5). After a 3 week recovery, SX + EGF and SH + EGF animals were given 50 microg/day EGF in their drinking water for 14 days. At day 14, saliva was collected, the animals were killed and the presence of EGF determined by radioligand-binding assay. Tongues were removed and histologically examined for the presence and morphology of taste buds on fungiform and circumvallate papillae, or immunostained for the presence of EGF, TGFalpha (transforming growth factor alpha) and EGFR (EGF receptor). The removal of submandibular and sublingual salivary glands resulted in the loss of fungiform taste buds and normal fungiform papillae morphology. These effects were reversed by EGF supplementation, indicating a role for EGF in fungiform taste bud maintenance. In addition, supplementation of EGF to sham-operated animals increased the size of fungiform taste buds. In contrast, removal of salivary glands had no effect on the size, numbers, or morphology of circumvallate taste buds, suggesting that the formation and maintenance of taste buds in fungiform and circumvallate papillae may involve different and distinct processes. EGF, TGFalpha and EGFR were localized to distinct layers of the dorsal epithelium and to within both fungiform and circumvallate taste buds. Their expression within the epithelium or taste buds was not altered with sialoadenectomy, indicating that the actions of endogenous EGF and TGFalpha are distinct and not regulated by exogenous EGF and TGFalpha supplied in saliva.

Rapid entry of bitter and sweet tastants into liposomes and taste cells: implications for signal transduction

Some amphipathic bitter tastants and non-sugar sweeteners are direct activators of G proteins and stimulate transduction pathways in cells not related to taste. We demonstrate that the amphipathic bitter tastants quinine and cyclo(Leu-Trp) and the non-sugar sweetener saccharin translocate rapidly through multilamellar liposomes. Furthermore, when rat circumvallate (CV) taste buds were incubated with the above tastants for 30 s, their intracellular concentrations increased by 3.5- to 7-fold relative to their extracellular concentrations. The time course of this dramatic accumulation was also monitored in situ in rat single CV taste buds under a confocal laser-scanning microscope. Tastants were clearly localized to the taste cell cytosol. It is proposed that, due to their rapid permeation into taste cells, these amphipathic tastants may be available for activation of signal transduction components (e. g., G proteins) directly within the time course of taste sensation. Such activation may occur in addition to the action of these tastants on putative G protein-coupled receptors. This phenomenon may be related to the slow taste onset and lingering aftertaste typically produced by many bitter tastants and non-sugar sweeteners.

Maturation of taste buds on the soft palate of the postnatal rat

Taste bud distribution on the soft palate and within three types of tongue papillae (fungiform, foliate, and circumvallate) were examined histologically in the rat at different postnatal ages. After paraffin embedding, serial sections (10 μm) were made and stained by HE, and digitized images of each section were examined. The existence of a taste pore was used to identify mature taste buds. At birth, 53% (68 of 127 observed) of the taste buds on the soft palate, but only 14% (14 of 110 observed) within fungiform papillae, contained a taste pore. One week after birth, the number of mature taste buds increased rapidly, resulting in 90% of soft palate taste buds and 80% of fungiform taste buds containing taste pores. In contrast, no taste buds with pores were observed at birth within foliate and circumvallate papillae; however, at two weeks after birth 52% (71 of 132 observed) of the foliate and 68% (180 of 267 observed) of the circumvallate taste buds examined contained taste pores. These results suggest that taste buds within the soft palate play an important role in the detection of nutrients in the neonatal rat.

Efficacy of Exogenous Oral Zinc in Treatment of Patients with Carbonic Anhydrase VI Deficiency

Background: We previously described a disorder in 18 patients with decreased parotid saliva gustin/carbonic anhydrase (CA) VI secretion associated with loss of taste (hypogeusia) and smell (hyposmia) and distorted taste (dysgeusia) and smell (dysosmia). Because gustin/CAVI is a zinc-dependent enzyme we instituted a study of treatment with exogenous zinc to attempt to stimulate synthesis/secretion of gustin/CAVI and thereby attempt to correct the symptoms of this disorder. Methods: Fourteen of the 18 patients with this disorder completed the study. They were treated with 100 mg of exogenous zinc daily for 4 to 6 months, in an open clinical trial. Both before and after treatment, measurements were obtained of parotid saliva gustin/CAVI, parotid saliva, serum and urine zinc, taste and smell function, and, in some patients, examination of circumvallate taste buds by electron microscopy. Results: Treatment success was predicated upon significant increases in parotid saliva gustin/CAVI. This occurred in 10 of the 14 patients who were labeled responders; they also exhibited improvement in taste and smell acuity, a diminution in dysgeusia and dysosmia and increased zinc concentrations in parotid saliva, serum, and urine. Taste bud morphology returned to normal in each responder in whom it was measured. No increase in gustin/CAVI occurred in 4 patients who were labeled nonresponders; they exhibited no improvement in taste or smell acuity and no increases in parotid saliva zinc. However, serum and urine zinc increased to levels similar to those measured in the 10 responders. Two of 4 nonresponders reported diminution in dysgeusia and dysosmia. Taste bud morphology did not change from the abnormal state in the 1 nonresponder in whom it was measured. Conclusions: Zinc treatment is effective in patients in whom this trace metal increases synthesis/secretion of gustin/CAVI and ineffective in those in whom it does not. Increased gustin/CAVI in this disorder is probably associated with zinc stimulation of the gene responsible for the synthesis/secretion of gustin/CAVI. Among nonresponders, zinc was ineffective for several possible reasons, including resistance to zinc and possible sialylation of gustin/CAVI, which may render it functionally ineffective. Results suggest the hypothesis that gustin/CAVI is a trophic factor that promotes growth and development of taste buds through its action on taste bud stem cells.

Expression of Mash1 in basal cells of rat circumvallate taste buds is dependent upon gustatory innervation

Mash1, a mammalian homologue of the Drosophila achaete-scute proneural gene complex, plays an essential role in differentiation of subsets of peripheral neurons. In this study, using RT-PCR and in situ RT-PCR, we investigated if Mash1 gene expression occurs in rat taste buds. Further, we examined dynamics of Mash1 expression in the process of degeneration and regeneration in denervated rat taste buds. In rat tongue epithelium, Mash1 gene expression is confined to circumvallate, foliate, and fungiform papilla epithelia that include taste buds. In taste buds, Mash1-expressing cells are round cells in the basal compartment. In contrast, the mature taste bud cells do not express the Mash1 gene. Denervation and regeneration experiments show that the expression of Mash1 requires gustatory innervation. We conclude that Mash1 is expressed in cells of the taste bud lineage, and that the expression of Mash1 in rat taste buds is dependent upon gustatory innervation.

Molecular and Physiological Evidence for Glutamate (Umami) Taste Transduction via a G Protein‐Coupled Receptora

Recent molecular analyses have demonstrated that a metabotropic glutamate receptor, mGluR4, is expressed in taste buds from rat circumvallate and foliate papillae. Behavioral studies demonstrated that L(+)-2-amino-4-phosphonobutyric acid (L-AP4), an agonist for mGluR4 and related receptors, mimics the taste of monosodium glutamate (MSG) in rats. mGluR4 is known to signal through inhibition of the cyclic adenosine-5',3'-monophosphate (cAMP) cascade. Circumvallate and foliate taste buds exhibit decreases of cAMP levels following stimulation with MSG, and the response is potentiated by 5'-inosine monophosphate, suggesting that it is related to umami taste. Further, experiments on mice with the mGluR4 gene knocked out support the interpretation that mGluR4 is a key component in glutamate taste. Glutamate may also stimulate taste buds through an ionotropic receptor pathway. In patch-clamp studies, glutamate evokes two types of currents, similar to those elicited by N-methyl-D-aspartate (NMDA) and L-AP4. We speculate upon the significance of two glutamate receptor pathways in taste buds.

Occurrence of subtypes of gustatory cells in cat circumvallate taste buds.

Transmission-electron microscopy of cat taste buds confirmed that, as in other mammals, each taste bud comprised four distinct types of cells: Type I, Type II, Type III (gustatory), and Type IV (basal) cells. Gustatory cells made synaptic contacts with nerves to which synaptic vesicles were gathered. The following are the main findings on the cat gustatory cells: 1) The synaptic vesicles of gustatory cells were essentially all dense-cored in type; small clear vesicles, which usually are intermingled in other mammals, could not be found. 2) The vesicles were accumulated not only in the synaptic area but also in the basal cytoplasm. This implies endocrine and paracrine functions. 3) On the basis of the fine structure of the vesicles, two subtypes of gustatory cells were discriminated. A large part of the cells contained vesicles measuring about 180 nm in diameter, while a small part had smaller ones of about 100 nm in diameter. This is the first demonstration of a dual population of gustatory cells in mammals, suggesting different messenger substances utilized.

Distribution and characterization of functional amiloride-sensitive sodium channels in rat tongue.

The role of amiloride-sensitive Na+ channels (ASSCs) in the transduction of salty taste stimuli in rat fungiform taste buds has been well established. Evidence for the involvement of ASSCs in salt transduction in circumvallate and foliate taste buds is, at best, contradictory. In an attempt to resolve this apparent controversy, we have begun to look for functional ASSCs in taste buds isolated from fungiform, foliate, and circumvallate papillae of male Sprague-Dawley rats. By use of a combination of whole-cell and nystatin-perforated patch-clamp recording, cells within the taste bud that exhibited voltage-dependent currents, reflective of taste receptor cells (TRCs), were subsequently tested for amiloride sensitivity. TRCs were held at -70 mV, and steady-state current and input resistance were monitored during superfusion of Na(+)-free saline and salines containing amiloride (0.1 microM to 1 mM). Greater than 90% of all TRCs from each of the papillae responded to Na+ replacement with a decrease in current and an increase in input resistance, reflective of a reduction in electrogenic Na+ movement into the cell. ASSCs were found in two thirds of fungiform and in one third of foliate TRCs, whereas none of the circumvallate TRCs was amiloride sensitive. These findings indicate that the mechanism for Na+ influx differs among taste bud types. All amiloride-sensitive currents had apparent inhibition constants in the submicromolar range. These results agree with afferent nerve recordings and raise the possibility that the extensive labeling of the ASSC protein and mRNA in the circumvallate papillae may reflect a pool of nonfunctional channels or a pool of channels that lacks sensitivity to amiloride.

Changes in IP3 and cytosolic Ca2+ in response to sugars and non-sugar sweeteners in transduction of sweet taste in the rat.

1. The transduction pathways of sweet-sensitive cells in rat circumvallate (CV) taste buds were investigated with assays for inositol 1,4,5-trisphosphate (IP3) and with Ca2+ imaging. Stimulation with the non-sugar sweeteners SC-45647 and saccharin rapidly increased the cellular content of IP3 by 400 pmol (mg protein)-1, while sucrose had a much smaller effect on IP3. As shown previously, sucrose, but not saccharin, increased the content of cyclic adenosine monophosphate (cAMP) of this preparation. 2. Stimulation of isolated CV taste buds with SC-45647 increased the cytosolic Ca2+ concentration ([Ca2+]i) by 56.7 +/- 3.2 nM (n = 181). Due to the non-confocality of the measuring system, these concentrations are underestimates. The increase in [Ca2+]i did not require the presence of extracellular Ca2+, suggesting that the Ca2+ release was from intracellular stores. 3. Individual cells responding to the non-sugar sweeteners with Ca2+ release also responded to sucrose and to forskolin with an increase in [Ca2+]i. Such cells did not respond to the bitter tastant denatonium chloride. 4. Responses to sucrose were abolished by lowering the Ca2+ concentration of the stimulus solution, indicating Ca2+ uptake from the extracellular medium. 5. The responses of sweet-sensitive cells to forskolin were also abolished when Ca2+ ions were omitted from the stimulus solution. They were partially inhibited by the presence of Co2+, Ni2+, D600 (methoxyverapamil) and amiloride, indicating multiple pathways of Ca2+ uptake activated by cAMP. 6. In conclusion, a sweet-sensitive cell of the rat responds to sucrose with an increase in cAMP and Ca2+ uptake, but to non-sugar sweeteners with an increase in IP3 and Ca2+ release. The increase in [Ca2+]i, common to both pathways, is presumably required for synaptic exocytosis and for signal termination.

A possibility of efferent innervation of the gustatory cell in the rat circumvallate taste bud.

A transmission-electron microscope study of the rat circumvallate taste bud revealed that occasionally one and the same gustatory or Type III cell received innervation associated with subsurface cisterns as the Type II cell did, in addition to the ordinary afferent synapse. The subsurface cistern a flattened, smooth-surfaced saccular membrane system, hugged the plasma membrane of the gustatory cell along the boundary against the nerve terminal. The nerve terminal attaching to the cell was occupied with mitochondria and synaptic type vesicles. As these structural features resemble those of efferent synapses in certain other sensory cells including the inner ear hair cells, the gustatory cells dually innervated as demonstrated in the present study are presumably involved not only in the afferent but also in the efferent projection of nerves.

Histological localization of the sweet taste receptor in rat taste buds by the use of gurmarin, a sweet taste-suppressing peptide.

The binding site of gurmarin, a peptide inhibiting the sweet-taste sensation, was studied in taste buds in rat circumvallate papillae by means of a histochemical technique. Frozen sections of tongues were incubated with gurmarin conjugated with biotin and thereafter examined with a light microscope. Positive reactivity for the peptide was localized to the taste hairs, the apical projections of taste bud cells. The reaction appeared in about 10% of the circumvallate taste buds examined. As electrophysiological studies indicate that gurmarin suppresses the sweet-taste sensation at the level of reception, the present study suggests that the receptor for sweet taste is located on the taste hairs, and, furthermore, is present only in a certain, limited number of the taste buds.

Synaptic organization of vertebrate taste buds

The vertebrate taste bud is an end organ specialized to detect and transduce aqueous chemical stimuli. In mammals most taste buds are located on the tongue. Lingual taste buds are typically distributed over three fields or papillae: fungiform, foliate and circumvallate papillae. Fungiform papillae are found on raised eminences near the tip of the tongue. Each fungiform papilla contains from one to several taste buds. Foliate taste buds are located in epithelial folds (foliate papillae) of the posterolateral surfaces of the tongue. In the rear of the tongue circumvallate taste buds line the walls or trenches surrounding the mushroom-shaped circumvallate (= vallate) papillae. In fish, taste buds are more widely distributed, being located on the tongue, lips, barbels, gill rakers, palatal organ and the body surface. A typical vertebrate taste bud comprises 50 to 150 spindle-shaped cells that lie atop the basal lamina of the tongue.In most mammals, the taste bud cells can be classified as dark or light cells, based on the electron-density of their cytoplasm.

Synaptic connectivity in hamster circumvallate taste buds

The hamster is an interesting animal model for studying gustation because of its remarkable responsiveness to sweet stimuli. Previous investigators of hamster circumvallate taste have categorized cells into type I, II, and basal cells. The goals of the present study were to determine the ultrastructure of the various cell types and to analyze synaptic contacts in circumvallate taste buds of the hamster.Adult female Syrian hamsters were prepared for electron microscopy and thick Epon sections (2-4μm) were collected on microscope slides. Selected sections were then reembedded for thin sectioning. Thin sections were sliced and mounted onto Formvar-coated slot grids and poststained in uranyl acetate and lead citrate. The sections were examined with a Hitachi H-7000 transmission electron microscope at 100 kV.The circumvallate (=vallate) papilla of the hamster is located in the midline of the posterior portion of the tongue. Type I (dark cells), type II cells (light cells) and basal cells were identified within these taste buds.

Cellular relations in mouse circumvallate taste buds.

The fine structure of the taste buds of circumvallate papillae of two strains of mice was studied by electron microscopy. Mice anesthetized with ketamine were perfused through the heart with a double aldehyde mixture in cacodylate buffer and the tissues embedded in Epon. Semi-serial sections were employed. The morphology and relationships of cell types are consistent with the majority of descriptions of mammalian taste buds served by the ninth cranial nerve. Cells of type II are particularly well documented, as the stages in their origin, maturation and degeneration could be followed. Significant differences, however, relate to cell type I. These cells contain large dense-cored granules, contrasted with the more irregular and somewhat larger dark granules of the type I cells in the rabbit. These granules do not produce a dense homogenous product for the pore, as seen in the rabbit. Rather the pore substance consists of small, empty vesicles in a diffuse dark matrix. These granules are only moderately larger than the dense-cored vesicles of the type III cells. All features of the type III cell were demonstrated, although no instance of a complete cell was seen in any section. No significant differences were noted between the two strains of mice. Intimate proximity of a nerve to a cell nucleolus, suggestive of a trophic pathway, is illustrated.

HVEM ultrastructural analysis of mouse fungiform taste buds, cell types, and associated synapses

We have used high voltage electron microscopy and computer-generated three-dimensional reconstructions from serial sections to elucidate the structure of taste bud cells and their associated synapses in fungiform taste buds of the mouse. Five fungiform taste buds (two of which were serially sectioned) were examined with the high-voltage electron microscope (HVEM). We identified the synaptic connections from taste cells onto sensory nerve fibers and classified the presynaptic taste cells based on previously established ultrastructural criteria. From those data we have distinguished dark, intermediate, and light cells in murine fungiform taste buds. Synapses in murine fungiform taste buds are fewer in number, but contain many more vesicles than synapses in either foliate or circumvallate taste buds. Synapses in mouse circumvallate and foliate taste buds typically contain a few to several synaptic vesicles per section, whereas fungiform synapses may have in excess of 100 vesicles per profile. The significance of these differences in the numbers of synapses and synaptic structure between fungiform and circumvallate/foliate synapses is not known. Based on the small number of synapses observed in fungiform taste buds, we speculate that fungiform taste buds have only a few cells transducing sensory stimuli at any given time. Alternatively, communication of sensory information from the taste receptor cells to the afferent nerve fibers may be mediated by some other mechanism(s) in addition to classical chemical synapses.

Calbindin-like immunoreactivity in two peripheral chemosensory tissues of the rat: taste buds and the vomeronasal organ

In the rat, calbindin-like immunoreactivity was observed at both the light and electron microscopic levels within the chemoreceptor neurons of the vomeronasal organ (VNO) and both intragemmal cells and associated nerve fibers of the circumvallate taste buds. All VNO neurons were immunoreactive. Only a subset of intragemmal taste cells was immunoreactive; associated immunoreactive nerve fibers were apposed to both labeled and unlabeled cells but no synaptic contacts were observed.

Lectin cytochemistry of the dark granules in the type 1 cells of Syrian hamster circumvallate taste buds

Lectin-gold complexes in the dark granules (DGs) and the dense substance (DS) of vallate taste buds were localized. Both the DGs and the DS were labelled with wheat-germ agglutinin, Ulex europeus agglutinin-I and peanut agglutinin. Their common reaction to these lectins suggested that the DGs contain carbohydrate components similar to those of the DS. The results provide cytochemical evidence that the DS in the taste pit represents, at least in part, the content of the DGs in the type 1 cells.

A histochemical study of APUD ability in the taste buds of experimentally induced zinc‐deficient mice

To explore the relationship between taste acuity and zinc deficiency, a histochemical investigation was made into the taste buds of mice fed a zinc-deficient diet. Nine weeks after the start of the diet, the average serum zinc level of the mice was 45% lower than that of a control group of mice. Moreover, growth was arrested significantly. Two-bottle preference tests revealed that the intake ratio of 10(-5) M quinine hydrochloride solutions had increased markedly in the zinc-deficient mice compared with the controls. The circumvallate taste buds showed no morphological changes. Fluorescent histochemical examination showed an uptake of a monoamine precursor (5-HTP) by the gustatory cells in the zinc-deficient mice after the 5-HTP treatment. Upon immunohistological examination, however, no serotonin immunoreactivity appeared in the gustatory cells of the zinc-deficient mice after the 5-HTP treatment. These results suggest that zinc-deficiency may induce hypogeusia and decrease the ability to transform a monoamine precursor to monoamine in the gustatory cells, albeit the monoamine precursor uptake ability is not affected.

妊娠ラットへのビタミン A の過剰投与が胎仔の舌有郭乳頭ならびに味蕾の発育におよぼす影響に関する神経組織学的研究

妊娠ラットへのビタミン A の過剰投与が胎仔の舌有郭乳頭ならびに味蕾の発育におよぼす影響に関する神経組織学的研究

A morphometrical observation on the nerve fibers in the human taste bud with an electron microscope (author's transl)

The whole thickness of the epithelium of the human circumvallate papillae was dividedtentatively into three portions from its superior surface; upper (UP), middle (MP), and lower portions (LP). The photos of three transverse sections of circumvallate taste bud in one portion, totally nine sections in three portions, were taken with an electron microscope. After making a photomontage connecting nine photos at each level, the number and the area of nerve fibers were observed morphometrically.The results are as follows:1. The numbers of nerve fibers observed were 535 at LP, 272 at MP and 239 at UP. The average numbers in sections were 178.3 at LP, 90.7 at MP and 79.7 at UP. The packing densities were 0.47/μ2 at LP, 0.12/μ2 at MP and 0.10/μ2 at UP.2. The mean values of the area were 0.30μ2 at LP, 0.92μ2 at MP and 1.05μ2 at UP. The size distribution of the area did not distribute even at each portion ranging from 0.01μ2 to 8.23μ2. The areal distribution was not significant under the level of 1 percent by the triserial correlation among LP-MP-UP, but significant under the level of 1 percent by the biserial correlation and critical ratio alnong LP-MP, LP-UP and LP-(MP+UP). Sixty-six percent of the nerve fibers at LP, 29.4 percent at MP and 40.3percent at UP were small than o.20μ2, which indicates characteristic distribution. The result on the significance of the independence of the areal division by χ2-test, when areas were divided into two categories at 0.20μ2 and into four categories at 0.40μ2, 1.80μ2 and 4.00μ2, was the same as that of the areal distribution mentioned above.The nerve process and nerve courses presumed from the results of the present observation are characteristic and support quantitatively the reports previously published by other authors.