Occurrence of nitric oxide synthase in taste buds of the rat vallate papilla

Nitric oxide (NO) is generated by some types of cells as a membrane-permeant, short-acting paracrine signal. Its effects include activation of ion channels as well as formation of cGMP in the NO-generating and/or neighbouring cells. We have explored the possible involvement of NO in taste transduction by searching for NO synthase with histochemical and immunohistochemical methods. In taste buds of the rat vallate and foliate papilla, we found NADPH-diaphorase activity under stringent conditions that suppress the reactions of non-NO synthase enzymes. Furthermore, an antibody against neuronal NO synthase (NOS-I) labelled the basal and apical parts of taste cells, while an antibody against endothelial NO synthase (NOS-III) labelled taste buds and lingual epithelium more uniformly. The inducible macrophage enzyme NOS-II did not show immunoreactivity in taste buds. The results provide a first suggestion that NO may play a role in taste transduction. © 1998 Chapman & Hall     

MDR1 in taste buds of rat vallate papilla: functional, immunohistochemical, and biochemical evidence

Multidrug resistance P-glycoprotein (MDR1) is a membrane proteinof 150-170 kDa that catalyzes the ATP-driven efflux of hydrophobic xenobiotics, including fluorescent dyes, from cells. Expressed in manyepithelial tissues and in the endothelia of the blood-brain barrier,the MDR1 protein provides major routes of detoxification. We found thattaste cells of the rat vallate papilla (VP; posterior tongue) had onlya slow increase in fluorescence due to uptake of the hydrophobic dyecalcein acetoxymethyl ester. However, the development of fluorescencewas accelerated two- to threefold by substrates and/orinhibitors of MDR1, such as verapamil, tamoxifen, and cyclosporin A,and by addition of the transport-blocking antibody to MDR1, UIC2.Western blots of vallate tissue rich in taste buds withthe MDR1-specific monoclonal antibodies C219 and C494 revealed animmunoreactive protein at ~170 kDa. In contrast, the lingual epithelium surrounding the VP showed a much weaker band with these antibodies. Furthermore, using the antibodies C494 and UIC2 with tissuesections, MDR1-like immunoreactivity was found in taste cells. Theseresults show that MDR1 is present and functional in vallate taste cellsof the rat. MDR1-related transport may achieve active elimination ofxenobiotics from the sensory cells and thereby protect the peripheraltaste organs from potentially harmful molecules contained in ananimal's food.

     

Histogenesis of the taste buds of the vallate papilla in the in the rat in the postnatal stages of development

The developing taste buds of vallate papillae were studied with electron microscope in rats during the first 7 days after birth. Two types of cells--light and dark--are identified in the taste buds of a one day old animal. The apical parts of dark cells are characterized by numerous dark granules. A distinguishing feature of light cells is the presence of synaptic contacts with afferent intragemmal nerves. On the 4th day of development on the top of the apical parts of the cell, a microvillar apparatus is seen to form, which does not yet communicate with the oral cavity. On the 7th day, basal cells appear in the taste buds. Some of these cells are seen mitotically dividing. The differentiated microvillar apparatus now communicates with oral cavity. The structure of the taste buds is getting similar to that in the adults. The structural and functional peculiarities of the developing taste buds are discussed in association with the period of ontogenesis under consideration.     

The timing of alpha-gustducin expression during cell renewal in rat vallate taste buds

The G protein subunit alpha-gustducin is expressed in a subset of light (Type II) but not in dark (Type I) cells in rat vallate taste buds. The thymidine analogue 5-bromo-2'-deoxyuridine (BrdU) is incorporated into DNA during the S-phase of the cell cycle and can be used to determine the time of origin of a cell. In this study, 31 rats were injected with BrdU (50 mg/kg i.p.) and perfused at various times, from 2.5 to 10.5 days, following BrdU administration. Vallate papillae were embedded in polyester wax, cut into 4 microm transverse sections, and characterized with antibodies to BrdU and alpha-gustducin. Sections were processed for indirect immunofluorescence or with an immunoperoxidase procedure. From immunoperoxidase material on 21 rats, counts of alpha-gustducin- and BrdU-labeled cells were obtained from 300-800 taste bud profiles at each survival time; a total of 4122 taste bud profiles were examined. Cells with nuclei immunoreactive for BrdU occurred within the taste buds at 2.5 days and double-labeled cells were clearly evident at 3.5 days; a small number of double-labeled cells were seen as early as 2.5 days. Double-labeled cells reached a peak at 6.5 days and did not decline significantly by 10.5 days. Cells labeled for BrdU but not alpha-gustducin peaked at 5.5 days and showed a significant decline by 8.5 days. These latter cells included light cells not expressing alpha- gustducin and dark cells, which have previously been shown to have a shorter life span than light cells. These data suggest that expression of alpha-gustducin appears very early in a cell's life span and that these cells are longer lived than many of the cells that do not express this G protein.      

Solitary chemosensory cells in the developing chemoreceptorial epithelium of the vallate papilla

Summary The solitary chemosensory cells are considered typical of aquatic vertebrates and have never been described in the oral cavity of terrestrial vertebrates. We describe elements with ultrastructural characteristics of the solitary chemosensory cell in the gustatory epithelium of rats in the first week of extrauterine life. These elements appeared isolated, located among keratinocytes, and wrapped by glial-like elements. They showed a bipolar shape with a round cell body, a thin apical process, and a thicker basal one. Nerve fibers contacted the cell body and the processes. The cells showed small dense granules mainly located near nerve contacts. Small bundles of tonofilaments were visible in the perinuclear cytoplasm. Similar elements were not found in rats after the first week of extrauterine life. The present study demonstrates in a mammal that the development of taste buds is preceded by the presence of epithelial elements with ultrastructural characteristics of the solitary chemosensory cells described in lower vertebrates. This finding suggests that the oral chemoreception in the suckling rats may be mediated by three different patways (i.e., gustatory system, common chemical sense, and solitary chemosensory cell system).     

Apoptosis in mouse taste buds after denervation

Apoptotic cells in the taste buds of mouse circumvallate papillae after the sectioning of bilateral glossopharyngeal nerves were examined by the method of DNA nick-end labeling (TUNEL), together with standard electron microscopy. The taste buds decreased in number and size 3–11 days after denervation and disappeared at 11 days. The TUNEL method revealed only a few positively stained nuclei in normal taste buds but, in those of mice 1–5 days after denervation, the number of positive nuclei had increased to 3–5 times that of taste buds from normal mice. Electron-microscopic observation after denervation demonstrated taste bud cells containing condensed and fragmentary nuclei in a cytoplasm with increased density. The results show that taste bud cells under normal conditions die by apoptosis at the end of their life span, and that gustatory nerve sectioning causes apoptosis of taste bud cells with taste buds decreasing in number and ultimately disappearing. Received: 20 November 1995 / Accepted: 15 May 1996     

Number and distribution of ganglion cells in the vallate papilla of adult human

We examined the number and distribution of ganglion cells within a vallate papilla from a healthy human adult. The vast majority of cell bodies were located near the base of the papilla, clustered in the central portion of its core immediately above underlying muscles and von Ebner's glands. Neurons appeared to be of one morphological type with ovoid cell bodies and spherical nuclei. The possible functional significance of their presence in invaginated or trench-containing gustatory papillae is discussed.     

Regeneration of the vallate papilla in the rat with special reference to the origin of the taste bud-bearing epithelium

Summary The regeneration of the vallate papilla in the rat was studied by both light and electron microscopy. The papillae were excised and regeneration was studied at time intervals of 3 to 45 days. It was found that the vallate papilla is capable of regeneration after both partial and total papillectomies. The regenerated papillae were asymmetrical in shape. Several invaginations, independent of one another, were the equivalent of the original sulcus. Regeneration of the gustatory system occurred at circumscribed portions of the associated Von Ebner glands.The amounts of newly formed taste buds were proportional to the amount of nerve fibers in the subepithelial plexus. The regenerated taste buds showed normal histochemical and fine-structural characteristics. The results support the contention of some degree of specificity concerning the epithelium where taste bud regeneration occurs.Supported by CAICT Grant no 1776     

Effect of monoamines on the taste buds in the mouse

Summary Mouse taste buds were investigated following administration of monoamines and their precursors by fluorescence and electron microscopy. The appearance of fluorescent cells within the taste bud and the ultrastructural changes of vesicles in the gustatory cells were due to the treatment of 5-hydroxytryptophan. Small dense-cored vesicles (30–60 nm in diameter) appeared throughout the cytoplasm and accumulated especially at the presynaptic membranes of afferent synapses. Large dense-cored vesicles (80–100 nm) increased twice in number, and electron densities of their cores became more dense as compared with untreated mice. Fluorescent cells appeared in the taste bud of l-DOPA treated mice, whereas no ultrastructural changes were observed. These results suggest that the gustatory cells of the taste bud are capable of taking up and storing monoamines, which might act as neurotransmitters from the gustatory cells to the nerves.     

Network model of chemical-sensing system inspired by mouse taste buds

Taste buds endure extreme changes in temperature, pH, osmolarity, so on. Even though taste bud cells are replaced in a short span, they contribute to consistent taste reception. Each taste bud consists of about 50 cells whose networks are assumed to process taste information, at least preliminarily. In this article, we describe a neural network model inspired by the taste bud cells of mice. It consists of two layers. In the first layer, the chemical stimulus is transduced into an irregular spike train. The synchronization of the output impulses is induced by the irregular spike train at the second layer. These results show that the intensity of the chemical stimulus is encoded as the degree of the synchronization of output impulses. The present algorithms for signal processing result in a robust chemical-sensing system.     

Alpha-gustducin-immunoreactive solitary chemosensory cells in the developing chemoreceptorial epithelium of the rat vallate papilla

The presence of solitary chemosensory cells was studied in rat vallate papillae during the first week of post-natal life by alpha-gustducin immunocytochemistry. In 1- to 3-day-old rats, isolated alpha-gustducin-immunoreactive cells were found within the epithelium of the vallate papilla. These cells, mainly located in the basal layer, were scattered among keratocytes and wrapped in alpha-gustducin-negative epithelial cells in a glia-like fashion. The alpha-gustducin-immunoreactive cells were usually round and some of them gave rise to short, large processes directed towards the lumen of the oral cavity or the basal lamina. Rarely, some cells showed an evident bipolar shape. Small taste buds containing either alpha-gustducin-immunoreactive or alpha-gustducin-negative cells appeared in the vallate papillae of 4-day-old rats in which isolated, bipolar-shaped alpha-gustducin-immunoreactive cells were also found. After the first week of post-natal life, the taste buds appeared basically similar to those of adult animals. In conclusion, the present study demonstrates that the presence of epithelial cells with characteristics of solitary chemosensory cells precedes the development of the taste buds.     

Secretions of von Ebner's glands influence responses from taste buds in rat circumvallate papilla

The influence of secretions from von Ebner's lingual salivaryglands on gustatory function was studied in the rat. Neurophysiologicaltaste responses elicited by chemical stimulation of the circumvallatepapilla were recorded from the glossopharyngeal nerve whileinitiating salivary secretion in the same papilla. Salivarysecretion from von Ebner's glands significantly reduced tasteresponses to stimulation of the circumvallate papilla with variouschemicals. However, the magnitude of the reduction in responsediffered depending on the taste stimulus used. The reductionin response due to salivary secretion was blocked by prior administrationof the parasympathetic antagonist, atropine. These results demonstratea direct effect of salivary secretion on taste responses andillustrate the close relationship between taste function andthe secretion of von Ebner's glands.     

Effects of colchicine on the ultrastructure of mouse taste buds

Summary Effect of colchicine on the ultrastructure of taste bud cells was studied in the mouse. In untreated mice microtubules were abundant throughout the entire cytoplasm of type-III cells, but only in the apical cytoplasm of type-I cells. After 2 h of colchicine treatment, no microtubules were observed in any taste bud cells; dense secretory granules in the apical cytoplasm of type-I cells mostly disappeared, and instead, numerous phagosomes appeared. It is suggested that colchicine causes an interruption of the transport of the secretory granules in type-I cells from the Golgi apparatus to the membrane of the apical surface, from which release occurs. In type-III cells, after 4 or 5 h of treatment, dense-cored vesicles scattered throughout the cytoplasm tended to increase in number; they were often observed to accumulate in the vicinity of the Golgi apparatus. Five hours after treatment with 5-hydroxy-l-tryptophan (5-HTP) following colchicine pretreatment, monoamine specific fluorescent cells and vesicles with highly electron-dense cores of type-III cells were still present. On the other hand, 5 h after 5-HTP treatment alone both fluorescent cells and vesicles with highly electron-dense cores had already disappeared. These observations suggest that the treatment with colchicine interrupts the transport of densecored vesicles of type-III cells to synaptic areas, in which those vesicles are presumed to discharge the neurotransmitter substance.     

The effects of beta-bungarotoxin on the morphogenesis of taste papillae and taste buds in the mouse

Although it has been long accepted that innervation by a tastenerve is essential for maintenance of taste buds, it is notclear what role, if any, innervation plays in the morphogenesis oftaste papillae and taste bud development. The following studywas undertaken to determine what effects lack of sensory innervationhave on the development of taste papillae and the formationof taste buds in the mouse. Timed-pregnant female mice (n =3) at gestational day 12 (gd12) were anesthetized and a 1 µlsolution (1 µg/µl) of ß-bungarotoxin (ß-BTX),a neurotoxin that disrupts sensory and motor neuron development,was injected into the amniotic cavity of two embryos per dam.Two shams were injected with PBS. Fetuses were harvested atgd18, 1 day before birth, and four ß-BTX-injected embryos,two shams and two controls were fixed in buffered paraformaldehyde.Serial sections were examined for the presence and morphologyof taste papillae and taste buds. No nerve profiles were observedin ß-BTX-injected tongues. Although circumvallate papillaewere present on ß-BTX tongues, only five fungiform papillaecould be identified. Taste buds were present on a large percentageof fungiform papillae profiles (24% and on circumvallate papillaein sham and control fetuses; in contrast, no taste buds wereassociated with taste papillae in ß-BTX fetuses. Theseresults implicate a significant role for innervation in tastepapillae and taste bud morphogenesis.