Study by transmission and scanning electron microscopy of the morphogenesis of three types of lingual papillae in the albino rat (Rattus rattus)

El‐Bakry, A.M. 2010. Study by transmission and scanning electron microscopy of the morphogenesis of three types of lingual papillae in the albino rat (Rattus rattus).—Acta Zoologica (Stockholm) 91 : 267–278 Tongues were removed from albino rat foetus on days 12 (E12) and 16 (E16) of gestation and from newborns (P0) and from juvenile rats on days 7 (P7), 14 (P14) and 21 (P21) postnatally for investigation by light, scanning, and transmission electron microscopy. Significant changes appeared during the morphogenesis of the papillae. At E12, two rows of rudiments of fungiform papillae were extended bilaterally on the anterior half of the tongue. At E16, the rudiments of fungiform papillae were regularly arranged in a lattice‐like pattern. A rudiment of circumvallate papillae could be recognized. No rudiment of filiform papillae was visible. No evidence of keratinization was recognizable. At P0, rudiments of filiform papillae were visible but had a more rounded appearance, with keratinization. The fungiform and circumvallate papillae were large and their outlines were somewhat irregular as that found in the adult rat. At P7, the filiform papillae were large and slender. The fungiform papillae became large and the shape of circumvallate papillae was almost similar to that observed in the adult. At P14 and P21, the shape and structure of the three types of papillae were irregular as those found in the adult. In conclusion, the rudiments of the fungiform and circumvallate papillae were visible earlier than those of the filiform papillae. The morphogenesis of filiform papillae advanced in a parallel manner with the keratinization of the lingual epithelium, in the period from just before birth to a few weeks after birth.     

Spacing patterns on tongue surface-gustatory papilla

The dorsal surface of the mammalian tongue is covered with four kinds of papillae, fungiform, circumvallate, foliate and filiform papillae. With the exception of the filiform papillae, these types of papillae contain taste buds and are known as the gustatory papillae. The gustatory papillae are distributed over the tongue surface in a distinct spatial pattern. The circumvallate and foliate papillae are positioned in the central and lateral regions respectively and the fungiform papillae are distributed on the anterior part of the tongue in a stereotyped array. The patterned distribution and developmental processes of the fungiform papillae indicate some similarity between the fungiform papillae and the other epithelial appendages, including the teeth, feathers and hair. This is because 1) prior to the morphological changes, the signaling molecules are expressed in the fungiform papillae forming area with a stereotyped pattern; 2) the morphogenesis of the fungiform papillae showed specific structures in early development, such as epithelial thickening and mesenchymal condensation and 3) the fungiform papillae develop through reciprocal interactions between the epithelium and mesenchymal tissue. These results led us to examine whether or not the early organogenesis of the fungiform papillae is a good model system for understanding both the spacing pattern and the epithelial-mesenchymal interaction during embryogenesis.     

Immunohistochemical localization of neuron-specific enolase and calcitonin gene-related peptide in pig taste papillae.

Immunoreactivity to neuron-specific enolase (NSE), a specific neuronal marker, and calcitonin gene-related peptide (CGRP) was localized in lingual taste papillae in the pigs. Sequential staining for NSE and CGRP by an elution technique allowed the identification of neuronal subpopulations. NSE-staining revealed a large neuronal network within the subepithelial layer of all taste papillae. NSE-positive fibers then penetrated the epithelium as isolated fibers, primarily in the foliate and circumvallate papillae, or as brush-shaped units formed by a multitude of fibers, especially in the fungiform papillae and in the apical epithelium of the circumvallate papilla. Taste buds of any type of taste papillae were found to express a dense subgemmal/intragemmal NSE-positive neuronal network. CGRP-positive nerve fibers were numerous in the subepithelial layer of all three types of taste papillae. In the foliate and circumvallate papillae, these fibers penetrated the epithelium to form extragemmal and intragemmal fibers, while in the fungiforms, they concentrated almost exclusively in the taste buds as intragemmal nerve fibers. Intragemmal NSE- and CGRP-positive fiber populations were not readily distinguishable by typical neural swellings as previously observed in the rat. The NSE-positive neuronal extragemmal brushes never expressed any CGRP-like immunoreactivity. Even more surprising, fungiform taste buds, whether richly innervated by or devoid of NSE-positive intragemmal fibers, always harboured numerous intragemmal CGRP-positive fibers. Consequently, NSE is not a general neuronal marker in porcine taste papillae. Our observations also suggest that subgemmal/intragemmal NSE-positive fibers are actively involved in synaptogenesis within taste buds. NSE-positive taste bud cells were found in all three types of taste papillae. CGRP-positive taste bud cells were never observed.     

Neuro-histochemical study of the tongue of Gallus domesticus (White Leghorn)

An investigation was undertaken to study the neuro-histochemistry of tonue of Gallus domesticus (white Leghorn) by cholinesterase technique under maintained pH of 5.2, temperature 40 degrees C and incubation period 20 h, and Ungewitter's silver impregnation method. Fungiform (8--16 in number), filiform (5--16 in number), circumvallate and foliate (numerous) were recorded profusely innervated in the various regions of the tongue. Neuro-muscular spindles related either with the non-myelinated, myelinated nerve fibres or with the ganglia via pre- or postganglionic nerve fibre showed cholinesterase (AChE) positive activity in the form of dark brown patches. Neural network over the glands, formed the basket envelope likestructures. Glands showed positive cholinesterase (AChE) reaction on the periphery. Dot-like nerve endings were connected with the neural terminal network in the foliate papillae region and motor end plate like endings on blood vessel, were occasionally recorded. Parasympathetic ganglia of various shape and sizes, arranged in chain-like fashion were observed in the connective tissue space between the fungiform and circumvallate papillae and foliate papillae region. The pre- and postganglionic fibres of these ganglia were related with the anteriorly and posteriorly located papillae via the neural terminal network. Cholinesterase (AChE) activity was marked either in the centre, of the fungiform or on the periphery of the filiform, circumvallate and foliate papillae.     

Three-dimensional architecture of the connective tissue core and surface structures of the lingual papillae in the rabbit

Three-dimensional characteristics of the epithelial cell layer and connective tissue interface of the tongue were studied using scanning electron microscopy. In this study, the fragments of tongue were fixed in modified Karnovsky's fixative solution. Subsequently, the specimens were treated with 10% NaOH solution for 4-7 days at room temperature and postfixed in 1% OsO4 in 0.1 M phosphate buffer (pH 7.4) for 2 hours at 4 degrees C. They were dehydrated through a graded ethanol series, and critical-point dried with CO2. The specimens were coated with gold and observed in a scanning electron microscope, JEOL JSM-6100. The results showed numerous papillae on the dorsal surface of the tongue divided into four groups (filiform, fungiform, foliate and vallate papillae). Filiform papillae are conically shaped; fungiform papillae have an irregular round surface; foliate papillae are oval in shape and have some parallel projections; and vallate papillae are located in the posterior part of the tongue and have a depression around the center. After the treatment with 10% NaOH solution, the original arrangements of connective papillae could be seen. This characteristic three-dimensional distribution of the collagen fiber bundles is typical for each superficial papillae depending on whether it is filiform, fungiform, foliate or vallate.     

Anatomical, neurohistological and histochemical observations on the tongue of Francolinus pondicerianus (grey partridge or safed teeter).

The staining and demonstration of neural elements in relation with other associated structures was most satisfactory at pH 5.2, incubation period 20 h, and temperature 40degreesC. Ganglia of various shape and size were observed on or near the fungiform, filiform and foliate papillae. In the region of the fungiform papillae, a chain of ganglia (parasympathetic) was observed. The nerve cells of these ganglia showed cholinesterase (ChE) activity. The neural networks of the foliate papillae were connected with the neural terminal network and occasionally to the ganglia. The innervation of blood vessels (artery and vein) was profuse. At times ganglia were noticed on the periphery of the artery. Neuromuscular spindle-like structures were noticed occasionally. Dot-like free nerve endings were also observed. The ChE activity was very marked in the muscle fibres lying in the region of the circumvallate papillae but was less marked in the muscle fibres lying in the region of the filiform and fungiform papillae.     

Anatomical and neurohistological observations on the tongue of the India goat, Capra aegagrus.

The anatomy and neurohistology of the tongue of the Indian goat, Capra aegagrus, have been described. The apex linguae is notched in the centre. The foramen caecum is found to be absent. The sublingua could not be traced. The filiform papillae are the most common and divided into three types: the simple, giant, and true filiform papillae. The true filiform papillae are the most developed of the three types. The foliate papillae are absent. There are 13--14 circumvallate papillae arranged in two rows in a V-shaped pattern. The fungiform papillae are large and could easily be seen with the naked eye. They are scattered over the entire dorsum, being in abundance at the tip. The tongue of the goat is richly innervated. On the dorsum, the lamina propria is innervated by thick nerve fibres. In the fungiform papillae quite a large number of nerve fibres could be seen. The circumvallate papillae are also abundantly provided with nerves. A few ganglion cells are found below the circumvallate papillae. Thick nerve fibres are seen across the numerous glands and their ducts. Muscle fibres and connective tissue are also richly innervated.     

Localization of type II collagen in the lingual mucosa of rats during the morphogenesis of circumvallate papillae

Iwasaki, S., Aoyagi, H. and Yoshizawa, H. 2011. Localization of type II collagen in the lingual mucosa of rats during the morphogenesis of circumvallate papillae. —Acta Zoologica (Stockholm) 92 : 67–74. Immunoreactivity specific for type II collagen was recognized first in the mesenchymal connective tissue just beneath the circumvallate papilla placode in fetuses on E13. At this stage, most of the lingual epithelium was pseudostratified epithelium composed of one or two layers of cuboidal cells. However, the epithelium of the circumvallate papilla placode was composed of several layers of cuboidal cells. Immunoreactivity specific for type II collagen was detected mainly on the lamina propria just beneath the lingual epithelium of the rudiment of the circumvallate papilla in fetuses on E15 and on E17, and slight immunostaining was detected on the lamina propria around the rudiment. In fetuses on E19, immunoreactivity specific for type II collagen was widely and densely distributed on the connective tissue around the developing circumvallate papillae and on the connective tissue that surrounded the lingual muscle. Immunoreactivity specific for type II collagen was sparsely distributed on the lamina propria of central bulge. After birth, morphogenesis of the circumvallate papillae advanced gradually with the increase in size of the tongue. Immunoreactivity specific for type II collagen was distinctively distributed in the lamina propria around circumvallate papilla, in the central bulge, and in the connective tissue that surrounded the lingual muscle.     

Scanning electron microscopic study on the structure of the lingual papillae of the Saanen goat

The morphology of lingual papillae of the ten male mature Saanen goats (11 months old, approximately 42 kg in weight and of a known pedigree) was examined by scanning electron microscopy. Tissues were taken from the dorsal and ventral surfaces of the apex, body and root of the tongue, and were prepared accordingly and observed under the scanning electron microscope. On the dorsal and ventro-lateral surfaces of the lingual mucosa, three types of mechanical papillae (filiform, lenticular, and conical) and two types of gustatory papillae (vallate and fungiform) were observed. The structure and density of the filiform papillae differentiated on the anterior, posterior and ventro-lateral aspects of the tongue. Two types of lenticular papillae, both possessing a prominent surrounding papillary groove, were determined. The pyramidal-shaped type I lenticular papilla had a pointed apex while the round-shaped type II lenticular papilla possessed a blunt apex. Certain number of the type I lenticular papillae had double apices. The larger conical papillae were hollow structures, differing structurally from the filiform papillae with their larger size, a tip without projections and lack of the secondary papillae. The vallate papillae were present on both rims of the torus linguae, were encircled by a prominent gustatory furrow which was also surrounded by a thick annular fold. The fungiform papillae were scattered among the filiform papillae in the anterior two-thirds of the dorsal and lateral surfaces, and each of them was highly protected by surrounding filiform papillae, yet encircled by a papillary groove. Our findings indicate that Saanen goat have profuse distribution of papillae on the tongue displaying morphological features characteristic of mechanical function.     

Ultrastructural study of the precursor to fungiform papillae prior to the arrival of sensory nerves in the fetal rat.

The structure of precursors to fungiform papillae without taste buds, prior to the arrival of sensory nerve fibers at the papillae, was examined in the fetal rat on embryonic day 13 (E13) and 16 (E16) by light and transmission electron microscopy in an attempt to clarify the mechanism of morphogenesis of these papillae. At E13, a row of rudiments of fungiform papillae was arranged along both sides of the median sulcus of the lingual dorsal surface, and each row consisted of about 10 rudiments. There was no apparent direct contact between papillae rudiments and sensory nerves at this time. Bilaterally towards the lateral side of the tongue, adjacent to these first rudiments of fungiform papillae, a series of cord-like invaginations of the dorsal epithelium of the tongue into the underlying connective tissue, representing additional papillary primordia parallel to the first row, was observed. The basal end of each invagination was enlarged as a round bulge, indented at its tip by a mound of fibroblasts protruding into the bulge. At E16 there was still no apparent direct contact between rudiments of fungiform papillae and sensory nerves. Each rudiment apically contained a spherical core of aggregating cells, which consisted of a dense assembly of large, oval cells unlike those in other areas of the lingual dorsal epithelium. The differentiation of these aggregated cells was unclear. The basal lamina was clearly recognizable between the epithelium of the rudiment of fungiform papillae and the underlying connective tissue. Spherical structures, which appeared to be sections of the cord-like invaginations of the lingual epithelium that appeared on E13, were observed within the connective tissue separated from the dorsal lingual epithelium. Transverse sections of such structures revealed four concentric layers of cells: a central core, an inner shell, an outer shell, and a layer of large cells. Bundles of fibers were arranged in the central core, and the diameters of bundles varied somewhat depending on the depth of the primordia within the connective tissue and their distance from the median sulcus. Ultrastructural features of cells in the outer shell differed significantly in rudiments close to the lingual epithelium as compared to those in deeper areas of connective tissue. Around the outer shell there was a large-cell layer consisting of one to three layers of radially elongated, oval cells that contained many variously sized, electron-dense, round granules. Large numbers of fibroblasts formed dense aggregates around each spherical rudiment, and were separated by the basal lamina from the large-cell epithelial layer. Progressing from deep-lying levels of the rudiments of the papillae to levels close to the lingual surface epithelium, the central core, inner shell, and outer shell gradually disappeared from the invaginated papillary cords.     

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.     

Fine structure of the dorsal lingual epithelium of the Japanese monkey Macaca fuscata fuscata.

Filiform papillae, which were densely distributed all over the dorsal surface of the lingual body, were crown-shaped, with a central, circular area that sloped in the anterior direction and several branches that surrounded it in a semicircle from the back of the central area. Dome-shaped, fungiform papillae were scattered among these filiform papillae. At the posterior end of the lingual body, there were four circumvallate papillae. Prominent microridges and elevated intercellular borders were widely distributed in the central area of the filiform papillae and the interpapillar region. On the surface of the branches of the filiform papillae, microridges were rarely seen. On the surface of the fungiform papillae, indistinct microridges were observed. Histologically, the dorsal lingual epithelium revealed three different regions: the epithelium on the anterior side of the filiform papillae, the epithelium on the posterior side of the filiform papillae and the interpapillar epithelium. Whereas the basal and suprabasal cells are similar throughout, differences characterize the intermediate and surface layers. Keratohyalin granules appear predominantly in the intermediate layer in the epithelium on the anterior side of filiform papillae. In the epithelium on the posterior side of the filiform papillae, no keratohyalin granules occur and, instead, tonofibrils are prominent. The cells become significantly flattened. In the interpapillar epithelium, no keratohyalin granules are visible, and the tonofilaments occupy almost the entire cytoplasm of most cells in the intermediate and surface layers. The cells are larger in volume in these layers.     

Comparative scanning electron-microscopic study of the lingual papillae in two species of domestic mammals (Equus caballus and Bos taurus). 1. Gustatory Papillae

The morphological characteristics of bovine and equine gustatory lingual papillae are compared by scanning electron microscopy. The fungiform papillae in the cow have a shape that corresponds to their name, while in the horse, they almost do not emerge from the surface of the tongue. These papillae show taste pores in both species. The vallate papillae, four times larger in the horse than in the cow, show a complex organization of papillae and secondary grooves in the horse. In the cow, they occur single and are surrounded by a thick annular pad of lingual mucosa. Taste pores have been observed in the vallate papillae of both species, whereas in the foliate papillae, they are present only in the horse. A characteristic distribution of stratified scales and channeled tracts is observed on the surface of all gustatory papillae in both species. The possible functional importance of each type of gustatory papilla is discussed on the basis of their morphostructural features.     

Location of taste buds in intact taste papillae by a selective staining method

Summary Taste buds were found to stain strongly and selectively in intact papillae with highly acidic dyes such as ponceau S. In intact tongues the taste buds in the fungiform, circumvallate and foliate papillae of the cynomolgus monkey and in the fungiform papillae of the rat as well as the taste discs in the fungiform papillae of the frog could be visualized. This method enables a rapid location and counting of taste buds in taste papillae without preparing histological sections. In cynomolgus tongue material fixed in formalin, the dyes penetrate into the buds. In fresh tongues only the taste pore region of the buds stains, which suggests that in vivo taste buds are impenetrable underneath the pore.     

Taste profiles from single circumvallate papillae: comparison with fungiform profiles

Two experiments were conducted to investigate the psychophysicalresponse characteristics of single circumvallate papillae. InExperiment 1, 12 circumvallate papillae in four subjects werechemically stimulated to assess identification of taste qualities.Single circumvallate papillae were found to mediate multipletaste qualities, and the taste profiles obtained from differentpapillae were similar within the same subject. Moreover, sucrose,quinine monohydrochloride and citric acid elicited unitary andcharacteristic quality responding in these papillae from allsubjects, whereas NaCl elicited predominantly sour and/or bitterresponses from three of the four subjects. Experiment 2 directly compared responses obtained from singlecircumvallate papillae with those obtained from fungiform regionsof the tongue. Data for 10 subjects showed significantly greatersour responses to citric acid and NaCl in circumvallate papillaeand significantly greater salty responses to these compoundson the anterior tongue. In addition, the taste profiles forcitric acid and NaCl were distinct for circumvallate papillae,while those from the anterior tongue were similar. These datasuggest that the bitterness and sweetness of quinine and sugar,respectively, can be identified on the basis of sensory informationarising from either circumvallate or fungiform regions, butthat differentiation of the tastes of salts and acids may dependon a comparison of the input from both regions and/or additionalinformation arising from foliate regions.     

Location of taste buds in intact taste papillae by a selective staining method.

Taste buds were found to stain strongly and selectively in intact papillae with highly acidic dyes such as ponceau S. In intact tongues the taste buds in the fungiform, circumvallate and foliate papillae of the cynomolgus monkey and in the fungiform papillae of the rat as well as the taste discs in the fungiform papillae of the frog could be visualized. This method enables a rapid location and counting of taste buds in taste papillae without preparing histological sections. In cynomolgus tongue material fixed in formalin, the dyes penetrate into the buds. In fresh tongues only the taste pore region of the buds stains, which suggests that in vivo taste buds are impenetrable underneath the pore.     

Morphogenesis of rat lingual filiform papillae.

The morphogenesis of filiform papillae on rat tongue was investigated with the electron microscope. Tongue rudiments were first seen on the 12th day of gestation. At 15-17 days, dermal papillae had formed and were arranged in hexagonal array on the dorsal lingual surface. Capping each dermal papilla was a two-layered epithelium that protruded slightly above the lingual surface, thus forming the early filiform papilla. In the next stage of development, at 18-19 days of gestation, the epithelium lining the papilla had differentiated into two cell populations, one producing hard keratin, the other producing soft keratin. Some of the keratinized epithelial cells assumed a position at an acute angle to the tongue surface and extended deep into the epithelium. In the next stage, 20-21 days, a cleft appeared within these angularly oriented cells. This resulted in the division of the epithelium into keraatin-lined individual filiform papillae. Finally, the individual papillae increased in size to the adult form.