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PubMed2005

Expression and characterization of delayed rectifying K+ channels in anterior rat taste buds.

Delayed rectifying K+ (DRK) channels in taste cells have been implicated in the regulation of cell excitability and as potential targets for direct and indirect modulation by taste stimuli. In the present study, we have used patch-clamp recording to determine the biophysical properties and pharmacological sensitivity of DRK channels in isolated rat fungiform taste buds. Molecular biological assays at the taste bud and single-cell levels are consistent with the interpretation that taste cells express a variety of DRK channels, including members from each of the three major subfamilies: KCNA, KCNB, and KCNC. Real-time PCR assays were used to quantify expression of the nine DRK channel subtypes. While taste cells express a number of DRK channels, the electrophysiological and molecular biological assays indicate that the Shaker Kv1.5 channel (KCNA5) is the major functional DRK channel expressed in the anterior rat tongue.

Animals↗

PubMed2003

Expression of BDNF and TrkB in mouse taste buds after denervation and in circumvallate papillae during development.

BDNF (brain-derived neurotrophic factor) is a member of the neurotrophin family which affects the proliferation and survival of neurons. Using an immunocytochemical method, we examined the expression of BDNF and its receptor, TrkB, in the taste bud cells of the circumvallate papillae of normal mice and of mice after transection of the glossopharyngeal nerves. We additionally observed the expression of BDNF and TrkB in the developing circumvallate papillae of late prenatal and early postnatal mice. In normal untreated mice, BDNF was expressed in most of the taste bud cells; TrkB was detected in the plasma membrane of taste bud cells and in the nerve fibers. Double-labeling studies showed that BDNF and NCAM (neural cell adhesion molecule) or TrkB and NCAM colocalized in some of the taste bud cells, but that most taste bud cells were immunopositive for only BDNF or TrkB. NCAM-immunoreactive cells are known to be type-III cells, which have afferent synaptic contacts with the nerve terminals. Five days after denervation, the number of taste buds and nerve fibers markedly decreased; however, the remaining taste bud cells still expressed BDNF and TrkB. By 10 days after denervation, most of the taste buds had disappeared, and there were a few TrkB-immunoreactive nerve fibers in the connective tissue core. By 4 weeks after denervation, numerous TrkB-immunoreactive nerve fibers had invaded the papillae, and a few taste buds expressing BDNF and TrkB had regenerated. At E (embryonic day) 15 during development, the circumvallate papillae appeared, and then TrkB-immunoreactive nerve fibers entered the connective tissue core, and some of these fibers further invaded among the dorsal epithelial cells of the papillae. TrkB-immunoreactive oval-shaped cells were occasionally found in the dorsal epithelium. Such TrkB-immunoreactive nerve fibers and cells were also observed at E16-18. However, BDNF was not expressed in the papillae through the late prenatal days of E15 to E18. At P (postnatal day) 0, a cluster of BDNF-and TrkB-immunoreactive cells appeared in the dorsal epithelium of the papillae, and was presumed to be primitive taste buds. We conclude that TrkB-immunoreactive nerve fibers are necessary for papillary and taste bud formation during development and for the regeneration of taste buds after denervation. BDNF in the taste bud cells may act as a neurotrophic factor for innervating sensory neurons--through TrkB receptors of the axons of those neurons, and also may exert autocrine and paracrine trophic actions on neighboring taste bud cells by binding to their TrkB receptors.

Animals↗

PubMed2000

Induction of apoptosis by colchicine in taste bud and epithelial cells of the mouse circumvallate papillae.

Apoptotic cells in the taste buds and epithelia of mouse circumvallate papillae after colchicine treatment were examined by the methods of in situ DNA nick-end labeling, immunocytochemistry, and electron microscopy. After colchicine treatment, numerous positive cells appeared in the taste buds by DNA nick-end labeling, and some epithelial cells in the basal and suprabasal layers in and around the circumvallate papillae also revealed positive staining. Condensed and fragmented nuclei with a high density were occasionally found in the taste bud cells and in the basal and suprabasal layer epithelial cells by electron-microscopic observation. An immunocytochemical reaction for tubulin revealed weak staining in taste bud cells, because of the depolymerization of microtubules, and a decrease of the microtubules in the taste bud cells was observed by electron microscopy. These results indicate that colchicine treatment of mice induces the apoptosis of taste bud and epithelial cells in the circumvallate papillae and dorsal epithelial cells around the circumvallate papillae.

Animals↗

PubMed1976

Comparative ultrastructure of vallate, foliate and fungiform taste buds of golden Syrian hamster.

A fine-structure study of the hamster fungiform, foliate and vallate taste buds was undertaken for comparative purposes. All three taste bud types shared in common composition of the dark cells, light cells, basal cells, nerve fibers and nerve endings and undifferentiated peripheral cells, but morphological difference existed among them. The foliate and vallate taste buds were quite similar in their ultrastructural morphology. Their dark cells displayed long apical necks, long apical microvilli, apical osmiophilic secretory granules and an abundant rough endoplasmic reticulum. The dark cells of the fungiform taste buds, however, showed no neck formation and lacked apical osmiophilic granules. They had short apical microvilli and relatively scant rough endoplasmic reticulum. There was no difference in the fine structure features of the light cells, basal cells and neural elements of different types of taste buds. Both light and dark cells were much more readily distinguishable in foliate and vallate buds than in fungiform buds at both light-and electron-microscopic levels. Foliate and vallate buds demonstrated homogeneous dense substance within the taste pores while fungiform pores were frequently empty. It is speculated that the differences in taste bud morphology may be due to their different lingual locations and/or may be a reflection of the differences in the inductive influences from different nerves. Furthermore, structural differences may be responsible for varying thresholds to different taste modalities.

Animals↗

PubMed1985

Immunohistochemical observation on substance P in regenerating taste buds of the rat.

The present study was performed to investigate the relationship between substance P-positive (SP-positive) nerve fibers and regeneration of taste buds in the foliate papillae of the rat by means of immunohistochemistry. It was confirmed by neurotomy that taste buds in the foliate papillae of the rat were innervated mostly (90%) by the glossopharyngeal (IXth) nerve and partly (10%) by the chorda tympani. In this experiment, the IXth nerve was sectioned distal to the petrosal ganglion. The rats were sacrificed at various intervals from 20 days to 80 days after the operation. In the course of degeneration and disappearance of taste buds, both SP-positive fibers and taste buds disappeared completely from the posterior folds of the foliate papillae 7 days after the operation. Within 22 days, regenerated SP-positive fibers began to appear in the lamina propria, and following the penetration of the fibers into the epithelium, taste bud anlagen reappeared at the bottom of the trench, and in the posterior folds at 24 days. The process of new taste bud formation extended toward the apex of the trench and to the anterior folds, which seemed to follow the regeneration in the nerve trunk. Quantitative data showed a gradual increase in the number of taste buds and taste buds containing SP-positive fibers. These findings indicate that SP might have a role in regeneration of taste buds.

Animals↗

PubMed1998

Possible role of serotonin in Merkel-like basal cells of the taste buds of the frog, Rana nigromaculata.

Merkel-like basal cells in the taste buds of the frog were examined by fluorescence histochemistry, immunohistochemistry and electron microscopy. There were about 16-20 basal cells arranged in a radial fashion at the base of each taste bud. These cells were strongly immunopositive for serotonin antiserum. They were characterised by the presence of numerous dense-cored granules in the cytoplasm ranging from 80 to 120 nm in diameter, and of microvilli protruding from the cell surface. For 4 mo after sensory denervation by cutting the gustatory nerves, all cell types of the taste bud were well preserved and maintained their fine structure. Even at 4 mo after denervation, the basal cells exhibited a strong immunoreaction with serotonin antiserum. To investigate the function of serotonin in the basal cells in taste bud function, serotonin deficiency was induced by administration of p-chlorophenylalanine (PCPA), an inhibitor of tryptophan hydroxylase, and of p-chloroamphetamine (PCA), a depletor of serotonin. After administration of these agents to normal and denervated frogs for 2 wk, a marked decrease, or complete absence, of immunoreactivity for serotonin was observed in the basal cells. Ultrastructurally, degenerative changes were observed in both types of frog; numerous lysosome-like myelin bodies were found in all cell types of the taste buds. The number of dense-cored granules in the basal cells also was greatly decreased by treatment with these drugs. Serotonin in Merkel-like basal cells appears to have a trophic role in maintenance of the morphological integrity of frog taste bud cells.

Animals↗

PubMed1984

Thyroid hormone inhibits purified taste bud membrane adenosine 3',5'-monophosphate phosphodiesterase activity.

Thyroid hormone inhibited purified taste bud membrane adenosine 3'5'-monophosphate (cAMP) phosphodiesterase (PDE) activity in a dose-dependent manner. Taste bud membrane cAMP PDE was inhibited most effectively by thyroxine (T4) followed in order by triiodothyionine (T3), diiodotyrosine (DIT) and monoiodotyrosine (MIT). Concentrations required for 50% inhibition (IC50) of enzyme activity were about 1 x 10(-6), 1 x 10(-5), 6 x 10(-4) M and greater than 1 x 10(-3) M for T4, T3, DIT and MIT, respectively. Addition of zinc at physiological concentrations found in serum greatly augmented the inhibitory effects of T4 and T3 at lower concentrations (10(-7) and 10(-6) M, respectively) resulting in further inhibition of cAMP PDE by 40-50%. Inhibition of cAMP PDE by T4 appears to be relatively tissue selective as indicated by the IC50 of 1 x 10(-6) M for the taste bud but only 7 x 10(-6) M, 3 x 10(-5) M, and 4 x 10(-5) M, for heart, kidney and brain cAMP PDE, respectively. Inhibition of taste bud membrane cAMP PDE by T4 was competitive with substrate cAMP with a Ki of 4 microM. These results suggest that inhibition of cAMP PDE, which increases taste bud membrane intracellular cAMP, may participate in the action of thyroid hormone in the taste process.

3',5'-Cyclic-AMP Phosphodiesterases↗

PubMed1991

HVEM serial-section analysis of rabbit foliate taste buds: I. Type III cells and their synapses.

Serially sectioned rabbit foliate taste buds were examined with high voltage electron microscopy (HVEM) and computer-assisted, three-dimensional reconstruction. This report focuses on the ultrastructure of the type III cells and their synapses with sensory nerve fibers. Type III cells have previously been proposed to be the primary gustatory receptor cells in taste buds of rabbits and other mammals. Within rabbit foliate taste buds, type III cells constitute a well-defined, easily recognizable class and are the only taste bud cells observed to form synapses with intragemmal nerve fibers. Among 18 type III cells reconstructed from serial sections, 11 formed from 1 to 6 synapses each with nerve fibers; 7 reconstructed type III cells formed no synapses. Examples of both convergence and divergence of synaptic input from type III cells onto nerve fibers were observed. The sizes of the active zones of the synapses and numbers of vesicles associated with the presynaptic membrane specializations were highly variable. Dense-cored vesicles 80-140 nm in diameter were often found among the 40-60 nm clear vesicles clustered at presynaptic sites. At some synapses, these large dense-cored vesicles appeared to be the predominant vesicle type. This observation suggests that there may be functionally different types of synapses in taste buds, distinguished by the prevalence of either clear or dense-cored vesicles. Previous investigations have indicated that the dense-cored vesicles in type III cells may be storage sites for biogenic amines.

Animals↗

PubMed2006

Taste bud contains both short-lived and long-lived cell populations.

Taste bud cells undergo continual turnover even in adulthood, and their average lifespan has been estimated as approximately 10 days. However, it is not clear whether this figure can be applied to all the different cell types contained in a taste bud. Here, we describe the age and life cycle of taste bud cells in rat circumvallate papillae, and indicate that the lifespan is heterogeneous, ranging from 2 days to over 3 weeks. Taste bud cells were incorporated from the basal proliferative layer in 1-2 days after birth. After incorporation, approximately half of the cells were eliminated within 2-3 days, and the remaining half were maintained with gradual decrease, suggesting that there are at least two types of cells; short-lived cells and long-lived cells. Moreover, above 10% of the incorporated cells were maintained at 3 weeks. In order to gain information about the relationship between the cell functions and the cell age, we carried out double-labeling experiments using 5-bromo-2'-deoxyuridine and each of two markers for in situ hybridization: mammalian achaete-scute homolog 1 (Mash1) and phospholipase C beta 2 (PLCbeta2) as markers of early differentiation and functional taste signaling, respectively. Mash1 expression began immediately after the incorporation and reached a maximum at 5-6 days after birth. Fewer but distinct Mash1-positive cells were still observed after 3 weeks. PLCbeta2 expression was observed from day 5, reached a maximum at day 12, and continued over 3 weeks. Taken together, a taste bud contains both short-lived and long-lived cells: the short-lived cells are eliminated in a time course similar to the surrounding epithelial cells, and the long-lived cells including taste receptor cells have a lifespan longer than the previous estimation.

Analysis of Variance↗

PubMed2000

Distribution pattern of taste buds along hyoidal barbels of Mullus barbatus and M. surmuletus.

The external taste buds of bony fishes are differentiated and grouped throughout the body epithelium, concentrating in the buccopharyngeal cavity or in certain external organs such as the lips, barbels or fins. The family Mullidae (goatfishes) is characterized by having barbels covered with taste buds (TBs). However, It is not known how the TBs are distributed and how their characteristics vary along the barbels. To obtain answers to these questions, the morphological variability in taste bud density (deltaTB), pore area (A(P)) and taste bud area (A(TB)) was studied in Mullus barbatus (L., 1758) (red mullet) and M. surmuletus (L., 1758) (striped red mullet), for fish ranging in size from 58 to 227 and from 56 to 287 mm in total length (L(T)), respectively. Assuming that larger fish are older, in both species the pore area (A(P)) and taste bud area (A(TB)) increased with ontogenic development. Also, in both species clear differences were observed along the barbel. The delta(TB) increased from the basal area to the apical area, whereas both the A(P) and A(TB) had an inverse pattern.

Animals↗

PubMed2004

Cell contact-dependent mechanisms specify taste bud pattern during a critical period early in embryonic development.

After gastrulation, the pharyngeal endoderm is specified to give rise to taste receptor organs without further signaling from other embryonic tissues. We hypothesized that intercellular signaling might be responsible for the specification of taste buds. To test if and when this signaling was occurring, intercellular contacts were transiently disrupted in cultures of pharyngeal endoderm from axolotl embryos, and the number, size, and distribution of taste buds analyzed. Disruption of cell contacts at progressive time points, from neurula to late tail bud stages, revealed a critical period, during mid-tail bud stages, when disruption of cell contacts resulted in a significant increase in taste bud number and size. The spatial distribution of taste buds was also altered; taste buds were more clustered in explants disrupted during the critical period. These effects were not due to general alterations in mitosis and apoptosis. Rather, at least three aspects of taste bud patterning, i.e., number, size, and distribution, are governed by mechanisms dependent on normal cell contacts during a concise time window. Furthermore, our findings are consistent with specification of taste buds by means of lateral inhibitory signaling, which we hypothesize results from cell contact-dependent or short-range diffusible signals.

Ambystoma↗

PubMed2006

Espin cytoskeletal proteins in the sensory cells of rodent taste buds.

Espins are multifunctional actin-bundling proteins that are highly enriched in the microvilli of certain chemosensory and mechanosensory cells, where they are believed to regulate the integrity and/or dimensions of the parallel-actin-bundle cytoskeletal scaffold. We have determined that, in rats and mice, affinity purified espin antibody intensely labels the lingual and palatal taste buds of the oral cavity and taste buds in the pharyngo-laryngeal region. Intense immunolabeling was observed in the apical, microvillar region of taste buds, while the level of cytoplasmic labeling in taste bud cells was considerably lower. Taste buds contain tightly packed collections of sensory cells (light, or type II plus type III) and supporting cells (dark, or type I), which can be distinguished by microscopic features and cell type-specific markers. On the basis of results obtained using an antigen-retrieval method in conjunction with double immunofluorescence for espin and sensory taste cell-specific markers, we propose that espins are expressed predominantly in the sensory cells of taste buds. In confocal images of rat circumvallate taste buds, we counted 21.5 +/- 0.3 espin-positive cells/taste bud, in agreement with a previous report showing 20.7 +/- 1.3 light cells/taste bud when counted at the ultrastructural level. The espin antibody labeled spindle-shaped cells with round nuclei and showed 100% colocalization with cell-specific markers recognizing all type II [inositol 1,4,5-trisphosphate receptor type III (IP(3)R(3))(,) alpha-gustducin, protein-specific gene product 9.5 (PGP9.5)] and a subpopulation of type III (IP(3)R(3), PGP9.5) taste cells. On average, 72%, 50%, and 32% of the espin-positive taste cells were labeled with antibodies to IP(3)R(3), alpha-gustducin, and PGP9.5, respectively. Upon sectional analysis, the taste buds of rat circumvallate papillae commonly revealed a multi-tiered, espin-positive apical cytoskeletal apparatus. One espin-positive zone, a collection of approximately 3 mum-long microvilli occupying the taste pore, was separated by an espin-depleted zone from a second espin-positive zone situated lower within the taste pit. This latter zone included espin-positive rod-like structures that occasionally extended basally to a depth of 10-12 mum into the cytoplasm of taste cells. We propose that the espin-positive zone in the taste pit coincides with actin bundles in association with the microvilli of type II taste cells, whereas the espin-positive microvilli in the taste pore are the single microvilli of type III taste cells.

Adaptor Proteins, Vesicular Transport↗

PubMed1995

Taste bud cell generation in the perihatching chick.

Chick taste bud primordia initially appear in late gestation on embryonic day 17 (E17), 4 days before hatching. To track DNA synthesis and subsequent taste bud cell proliferation between E17 and the second day post-hatching (H2), single 25 muCi injections of tritiated thymidine (specific activity = 72.5 Ci/mmol) were administered in ovo during E15, E16, E17 or E18. Anterior mandibular oral epithelium was processed for light microscopic autoradiography. Sections through each taste bud's center were analysed for label (> or = 6 silver grains/gemmal cell nucleus), and bud diameter. Results indicated a major part of gemmal cell DNA synthesis does not occur until after E19 irrespective of the day of thymidine injection, suggesting postmitotic or quiescent (decycled) cells assemble to form the early bud primordium (E17-19) based on local tissue interactions. All buds examined from E20-H2 contained labelled cells. The day of injection was important since 5-day survival cases after E16 injection yielded about 25% the number of labelled cells/bud as compared with equivalent survival cases following E17-18 injections. These results are discussed with respect to parallel changes in bud shape and increasing bud diameter, and cell proliferation in possible extra- and intragemmal sources of bud cells.

Animals↗

PubMed1993

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.

Animals↗

PubMed2000

Localization of the glutamate-aspartate transporter, GLAST, in rat taste buds.

A number of putative neurotransmitter substances have been found in vertebrate taste buds. Amongst these glutamate has been localized in fibres innervating the buds and uptake of glutamate has been shown to occur into receptor cells. It is therefore possible that, in common with other sensory systems, glutamate is a neurotransmitter in taste buds. In the inner ear and retina of mammals, the membranes of supporting cells have been shown to contain the glial glutamate transporter GLAST. In the brain, this protein is involved in glutamate re-uptake into glial cells where the glutamate is converted into glutamine for recycling into glutamatergic terminals. In this study, the presence of GLAST has been investigated in taste buds in the rat vallate papilla and its distribution compared with that of glutamine to determine whether there are cells in this system that play a glia-like role in glutamate handling. Immunofluorescent labelling showed that a subset of cells in the taste bud contains GLAST. Immunogold labelling indicated that it occurs in the plasma membranes of supporting cells, especially on the fine cytoplasmic processes of dark cells towards the basal region of the bud. A protein of molecular mass similar to that of cerebellar GLAST was detected in immunoblots of excised papillae. Double labelling and semiquantitative analysis of glutamine and GLAST immunoreactivity showed that the GLAST-positive cells have a higher level of cytoplasmic glutamine than the adjacent cells. It is proposed that these GLAST-positive cells play a glia-like role in the uptake of glutamate following its release at synapses within the taste bud although the precise location of the latter remains uncertain. The GLAST-positive cells may also be involved in its subsequent conversion to glutamine in a glutamate/glutamine cycle similar to that described in the brain.

ATP-Binding Cassette Transporters↗

PubMed1980

Human taste: response and taste bud number in fungiform papillae.

The number of basic taste qualities registered by single human fungiform papillae is correlated with the number of taste buds borne on these papillae. Multiple sensitivity was demonstrated both in single fungiform papillae and in single taste buds, with response to all four of the basic taste qualities occuring in a single taste bud.

Adolescent↗

PubMed1983

Morphology of taste buds on the gill arches of the mullet Mugil cephalus, and the killifish Fundulus heteroclitus.

The morphology of taste buds on the gill arches of two euryhaline teleosts, the mullet Mugil cephalus, and the killifish Fundulus heteroclitus, were investigated using light microscopic and scanning and transmission electron microscopic techniques. On the mullet gill arches, taste buds were limited to the pharyngeal surfaces of the smooth-surfaced gill rakers. On the killifish gill arches, taste buds were located on the pharyngeal surfaces of all gill rakers and on the gill arch itself at the bases of the gill rakers. Despite dramatic differences in gill-raker structure between these two species, the taste buds themselves were similar ultrastructurally and closely resembled those described in other fishes. Cells within the taste buds included spindle-shaped dark and light cells and basal cells. Ultrastructural features of both the light and dark cells could support either receptor or transport functions. Tufts of microvilli, including one thick microvillus per light cell and numerous thin microvilli per dark cell, protruded at the apex of each taste bud between the ridged surface epithelial cells. Light cells contained numerous tubular membrane elements some of which appeared to open onto the apical surface of the taste bud. Dark cells contained numerous microtubules and apical, electron-lucent vesicles possibly involved in transport.

Animals↗

PubMed1987

Fixation and occurrence of dark and light cells in taste buds of fungiform papillae.

The fine structure of taste buds of the fungiform papillae of the Old World monkey Cynomolgus (Macaca fascicularis) was found to vary with the primary fixative solution used. Standard primary fixative solutions of glutaraldehyde as well as fixatives containing both formaldehyde and glutaraldehyde gave non-reproducible and usually poor results. Osmium tetroxide in distilled water gave reproducible fixation of acceptable quality. Further, when fixed in osmium tetroxide in distilled water, taste buds of the fungiform papillae did not show the dark and light cells well known from other studies. It is therefore suggested that the terms dark and light cells not be used as synonyms for receptor or support cells, since the appearance of taste bud cells is affected by fixation.

Animals↗