Distinct expression patterns of Hedgehog signaling components in mouse gustatory system during postnatal tongue development and adult homeostasis.

The Hedgehog (HH) pathway regulates embryonic development of anterior tongue taste fungiform papilla (FP) and the posterior circumvallate (CVP) and foliate (FOP) taste papillae. HH signaling also mediates taste organ maintenance and regeneration in adults. However, there are knowledge gaps in HH pathway component expression during postnatal taste organ differentiation and maturation. Importantly, the HH transcriptional effectors GLI1, GLI2 and GLI3 have not been investigated in early postnatal stages; the HH receptors PTCH1, GAS1, CDON and HHIP, required to either drive HH pathway activation or antagonism, also remain unexplored. Using lacZ reporter mouse models, we mapped expression of the HH ligand SHH, HH receptors, and GLI transcription factors in FP, CVP and FOP in early and late postnatal and adult stages. In adults we also studied the soft palate, and the geniculate and trigeminal ganglia, which extend afferent fibers to the anterior tongue. Shh and Gas1 are the only components that were consistently expressed within taste buds of all three papillae and the soft palate. In the first postnatal week, we observed broad expression of HH signaling components in FP and adjacent, non-taste filiform (FILIF) papillae in epithelium or stroma and tongue muscles. Notably, we observed elimination of Gli1 in FILIF and Gas1 in muscles, and downregulation of Ptch1 in lingual epithelium and of Cdon, Gas1 and Hhip in stroma from late postnatal stages. Further, HH receptor expression patterns in CVP and FOP epithelium differed from anterior FP. Among all the components, only known positive regulators of HH signaling, SHH, Ptch1, Gli1 and Gli2, were expressed in the ganglia. Our studies emphasize differential regulation of HH signaling in distinct postnatal developmental periods and in anterior versus posterior taste organs, and lay the foundation for functional studies to understand the roles of numerous HH signaling components in postnatal tongue development.

Polycomb Repressive Complex 1 Controls Maintenance of Fungiform Papillae by Repressing Sonic Hedgehog Expression.

How tissue patterns are formed and maintained are fundamental questions. The murine tongue epithelium, a paradigm for tissue patterning, consists of an array of specialized fungiform papillae structures that harbor taste cells. The formation of fungiform papillae is preceded by pronounced spatial changes in gene expression, in which taste cell genes such as Shh, initially diffused in lingual epithelial progenitors, become restricted to taste cells when their specification progresses. However, the requirement of spatial restriction of taste cell gene expression for patterning and formation of fungiform papillae is unknown. Here, we show that a chromatin regulator, Polycomb repressive complex (PRC) 1, is required for proper maintenance of fungiform papillae by repressing Shh and preventing ectopic SHH signaling in non-taste cells. Ablation of SHH signaling in PRC1-null non-taste cells rescues the maintenance of taste cells. Altogether, our studies exemplify how epigenetic regulation establishes spatial gene expression patterns necessary for specialized niche structures.

Abundant proliferating cells within early chicken taste buds indicate a potentially "built-in" progenitor system for taste bud growth during maturation in hatchlings.

Like other epithelial cells, taste bud cells have a short life span and undergo continuous turnover. An active stem or progenitor cell niche is essential for taste bud formation and maintenance. Early taste bud cells have a life span of ~4 days on average in chicken hatchlings when taste buds grow rapidly and undergo maturation. The average life span is shorter than that of mature taste bud cells of rodents (~10-12 days on average). To better understand the mechanism underlying taste bud growth and homeostasis in chickens, we analyzed the distribution of proliferating cells in different tissue compartments, including taste buds, the surrounding epithelium and the underlying connective tissue in P1-3 hatchlings and P45 chickens. Unlike rodents, which lack proliferating cells within both early and mature taste buds, chickens possessed abundant proliferating cells within early taste buds. Further, at post-hatch day 45, when taste buds are mature and undergo continuous cell renewal, taste buds also contained proliferating cells, though to a lesser extent. These proliferating cells in early taste buds, indicated by PCNA⁺ and BrdU⁺ cells, primarily localized to the basal region of taste buds and were largely unlabeled by the two known molecular markers for chicken taste bud cells (Vimentin and α-Gustducin), suggesting their undifferentiated status. Our data indicate that early chicken taste buds have "built-in" progenitors in order to grow to and maintain their large size and rapid cell turnover in hatchlings.

Neuronal delivery of Hedgehog directs spatial patterning of taste organ regeneration.

How organs maintain and restore functional integrity during ordinary tissue turnover or following injury represents a central biological problem. The maintenance of taste sensory organs in the tongue was shown 140 years ago to depend on innervation from distant ganglion neurons, but the underlying mechanism has remained unknown. Here, we show that Sonic hedgehog (Shh), which encodes a secreted protein signal, is expressed in these sensory neurons, and that experimental ablation of neuronal Shh expression causes loss of taste receptor cells (TRCs). TRCs are also lost upon pharmacologic blockade of Hedgehog pathway response, accounting for the loss of taste sensation experienced by cancer patients undergoing Hedgehog inhibitor treatment. We find that TRC regeneration following such pharmacologic ablation requires neuronal expression of Shh and can be substantially enhanced by pharmacologic activation of Hedgehog response. Such pharmacologic enhancement of Hedgehog response, however, results in additional TRC formation at many ectopic sites, unlike the site-restricted regeneration specified by the projection pattern of Shh-expressing neurons. Stable regeneration of TRCs thus requires neuronal Shh, illustrating the principle that neuronal delivery of cues such as the Shh signal can pattern distant cellular responses to assure functional integrity during tissue maintenance and regeneration.

Maintenance of Taste Organs Is Strictly Dependent on Epithelial Hedgehog/GLI Signaling.

For homeostasis, lingual taste papilla organs require regulation of epithelial cell survival and renewal, with sustained innervation and stromal interactions. To investigate a role for Hedgehog/GLI signaling in adult taste organs we used a panel of conditional mouse models to manipulate GLI activity within epithelial cells of the fungiform and circumvallate papillae. Hedgehog signaling suppression rapidly led to taste bud loss, papilla disruption, and decreased proliferation in domains of papilla epithelium that contribute to taste cells. Hedgehog responding cells were eliminated from the epithelium but retained in the papilla stromal core. Despite papilla disruption and loss of taste buds that are a major source of Hedgehog ligand, innervation to taste papillae was maintained, and not misdirected, even after prolonged GLI blockade. Further, vimentin-positive fibroblasts remained in the papilla core. However, retained innervation and stromal cells were not sufficient to maintain taste bud cells in the context of compromised epithelial Hedgehog signaling. Importantly taste organ disruption after GLI blockade was reversible in papillae that retained some taste bud cell remnants where reactivation of Hedgehog signaling led to regeneration of papilla epithelium and taste buds. Therefore, taste bud progenitors were either retained during epithelial GLI blockade or readily repopulated during recovery, and were poised to regenerate taste buds once Hedgehog signaling was restored, with innervation and papilla connective tissue elements in place. Our data argue that Hedgehog signaling is essential for adult tongue tissue maintenance and that taste papilla epithelial cells represent the key targets for physiologic Hedgehog-dependent regulation of taste organ homeostasis. Because disruption of GLI transcriptional activity in taste papilla epithelium is sufficient to drive taste organ loss, similar to pharmacologic Hedgehog pathway inhibition, the findings suggest that taste alterations in cancer patients using systemic Hedgehog pathway inhibitors result principally from interruption of signaling activity in taste papillae.

ß-Catenin Signaling Biases Multipotent Lingual Epithelial Progenitors to Differentiate and Acquire Specific Taste Cell Fates.

Continuous taste bud cell renewal is essential to maintain taste function in adults; however, the molecular mechanisms that regulate taste cell turnover are unknown. Using inducible Cre-lox technology, we show that activation of ß-catenin signaling in multipotent lingual epithelial progenitors outside of taste buds diverts daughter cells from a general epithelial to a taste bud fate. Moreover, while taste buds comprise 3 morphological types, ß-catenin activation drives overproduction of primarily glial-like Type I taste cells in both anterior fungiform (FF) and posterior circumvallate (CV) taste buds, with a small increase in Type II receptor cells for sweet, bitter and umami, but does not alter Type III sour detector cells. Beta-catenin activation in post-mitotic taste bud precursors likewise regulates cell differentiation; forced activation of ß-catenin in these Shh+ cells promotes Type I cell fate in both FF and CV taste buds, but likely does so non-cell autonomously. Our data are consistent with a model where ß-catenin signaling levels within lingual epithelial progenitors dictate cell fate prior to or during entry of new cells into taste buds; high signaling induces Type I cells, intermediate levels drive Type II cell differentiation, while low levels may drive differentiation of Type III cells.

Histomorfometria do epitélio ruminal de cabritos 1/2 sangue Boer submetidos a dietas com tortas oriundas da produção do biodiesel / Histomorphometrics of the ruminal epithelium of 1/2 blood Bôer goats submitted to diets with pies from the production of biodiesel

Objetivou-se determinar o melhor nível de inclusão e de substituição das tortas de dendê e amendoim, respectivamente, em dietas para cabritos 1/2 sangue Boer, por meio da avaliação histomorfométrica das papilas ruminais. Foram utilizados 40 cabritos 1/2 sangue Boer, para cada período experimental, machos, não castrados, com idade aproximada de três meses e com peso inicial de 15,01±1,76kg. Os períodos experimentais constaram de 75 dias e 72 dias. As dietas consistiram de volumoso feno de Tifton-85 e de ração em mistura completa, contendo níveis de inclusão da torta de dendê nas proporções de 0,0; 7,0; 14,0; e 21,0% com base na matéria seca; e de mistura completa, contendo níveis de substituição do farelo de soja pela torta de amendoim nas proporções de 0,0; 33,0; 66,0; e 100%, constituindo-se os tratamentos. No epitélio do rúmen, foram avaliadas altura e largura das papilas, assim como densidade papilar e espessura da parede muscular do rúmen. Entre estas, a altura das papilas, no experimento com torta de amendoim, foi a única a sofrer efeito linear decrescente (P<0,0014); as demais variáveis, tanto dos animais alimentados com torta de amendoim quanto daqueles alimentados com torta de dendê, não foram afetadas pela dieta. As médias de altura encontradas foram de 2,6 e 2,3mm para as tortas de amendoim e dendê, respectivamente. Portanto, a inclusão de até 21% de torta de dendê e a substituição de 100% de torta de amendoim no concentrado de cabritos não alteraram a morfometria das papilas ruminais.

The aim of this study was to determine the best level of pies and replacement of palm oil and peanut, respectively, in diets of 1/2 blood Bôer goats from the histomorphometry of the rumen papillae. A total of 40 1/2 blood male, unneutered, aged approximately three months and initial weight of 15.01±1.76kg Bôer goats were used in each experimental period. The experimental periods consisted of 75 days and 72 days. The diets consisted of roughage hay Tifton-85 and total mixed ration containing levels of inclusion of palm kernel cake in the proportions of 0.0, 7.0, 14.0 and 21.0% based on dry matter; and the second experiment consisted of a complete mixture containing substitution levels of soybean meal by groundnut cake in the proportions of 0.0, 33.0, 66.0 and 100%. Epithelial cells were evaluated in the rumen height and width of the papillae, and density and thickness of the papillary muscle of the rumen. Among these, the height of the papillae in the experiment with peanut butter pie was the only one to suffer a negative linear effect (P<0.0014), the other variables, both in animals fed groundnut cake and palm oil, were not affected by the diet. The average heights found were 2.6 and 2.3, for pies, peanut and palm oil, respectively. Therefore, the inclusion of up to 21% palm kernel cake and replacement of 100% peanut cake in the concentrate of kids did not alter the morphology of the rumen papillae.

The glossopharyngeal nerve controls epithelial expression of Sprr2a and Krt13 around taste buds in the circumvallate papilla.

Tastants reach the tip of taste bud cells through taste pores which are openings in the epithelium. We found Sprr2a is selectively expressed in the upper layer of the epithelium surrounding taste buds in the circumvallate papilla (CV) where the epithelium is organized into taste pores. Sprr2a is a member of a small proline-rich protein family, which is suggested to be involved in the restitution/migration phase of epithelial wound healing. The expression of Sprr2a was restricted to the upper layer and largely segregated with Ptch1 expression that is restricted to the basal side of the epithelium around the taste buds. Denervation resulted in the gradual loss of Sprr2a-expressing cells over 10 days similarly to that of taste bud cells which is in contrast to the rapid loss of Ptch1 expression. We also found that denervation caused an increase of Keratin (Krt)13 expression around taste buds that corresponded with the disappearance of Sprr2a and Ptch1 expression. Taste buds were surrounded by Krt13-negative cells in the CV in control mice. However, at 6 days post-denervation, taste buds were tightly surrounded by Krt13-positive cells. During taste bud development, taste bud cells emerged together with Krt13-negtive cells, and Sprr2a expression was increased along with the progress of taste bud development. These results demonstrate that regional gene expression surrounding taste buds is associated with taste bud formation and controlled by the innervating taste nerve.

Morfologia dos pré-estômagos e de papilas ruminais de cordeiras Santa Inês em crescimento submetidas a dois planos nutricionais / Morphology of pre-stomach and ruminal papillae of growing Santa Inês female lambs under two nutritional schemes

Para se avaliar o efeito do plano nutricional e crescimento sobre a massa dos pré-estômagos, morfologia e quantificação de papilas ruminais, trinta e seis cordeiras da raça Santa Inês foram submetidas a dois planos nutricionais (ad libitum ou restrito) sendo abatidas em diferentes pesos vivo (20, 30 ou 40 kg de peso vivo), em um delineamento inteiramente casualizado balanceado em arranjo fatorial 2x3. Feito o abate, as vísceras foram pesadas livres de seu conteúdo em seguida mediu-se o volume de repleção do rúmen e retículo. Amostras do tecido ruminal oriundas dos sacos cranial e ventral foram coletadas para posteriormente serem realizadas com auxílio de lupa estereoscópica as medidas morfométricas das papilas ruminais, altura, largura da base, área, papilas por cm² e área absortiva por cm². Os resultados obtidos foram submetidos a análise de variância e as médias resultantes por tratamento foram comparadas por meio de teste de Student Newmann Keuls. Os diferentes planos nutricionais não influenciaram a massa das vísceras rúmen, retículo e omaso (P>0,05), no entanto, observou-se crescimento dessas vísceras em função do aumento do peso ao abate. O volume dessas vísceras foi afetado pelo peso ao abate, e observou-se menores volumes para animais com alimentação ad libitum (P<0,10). O número de papilas por cm² foi reduzido com o aumento do peso ao abate, sendo que altura e área foram aumentadas quando em pesos maiores. O plano nutricional afetou apenas a área e altura das papilas ruminais oriundas do saco cranial. A área absortiva não foi afetada pelos tratamentos. Plano nutricional e diferentes pesos vivos influenciam a morfologia dos pré-estômagos de cordeiras da raça Santa Inês.

For the evaluation of nutritional schemes and change on the pre-stomach morphology and quantification of rumen papillae, 36 Santa Inês female lambs were submitted to two nutritional schemes (ad libitum or restrict) and slaughtered with different live weights (20, 30 or 40 kg) in a completely randomized factorial design 2x3. After slaughter, the viscera were weighed empty and their volume was measured. Samples of ruminal wall from the cranial and ventral sacs were collected and with a stereomicroscope photographed and analyzed regarding height, basal width, area, papillae per cm² and absorptive area per cm². The results were submitted to analyses of variance and the means were compared by Student Newman Keuls test. The different nutritional schemes did not influence the weight of rumen, reticulum or omasum (P>0.05), although, growth of the viscera was observed by increase in live weight. The viscera volume was affected by live weight, and smaller volume was observed in the animals fed ad libitum diet (P<0.10). The number of papillae per cm² was reduced by the increase in live weight. Height and area of papillae were larger in heavier animals. The nutritional scheme only affected height and area of papillae of the cranial sac. The absorptive area was not affected by the treatments. Different nutritional schemes and live weights affect the pre-stomach morphology of Santa Ines female lambs.

Calcium-activated potassium channel SK1 is widely expressed in the peripheral nervous system and sensory organs of adult zebrafish.

Sensory cells contain ion channels involved in the organ-specific transduction mechanisms that convert different types of stimuli into electric energy. Here we focus on small-conductance calcium-activated potassium channel 1 (SK1) which plays an important role in all excitable cells acting as feedback regulators in after-hyperpolarization. This study was undertaken to analyze the pattern of expression of SK1 in the zebrafish peripheral nervous system and sensory organs using RT-PRC, Westernblot and immunohistochemistry. Expression of SK1 mRNA was observed at all developmental stages analyzed (from 10 to 100 days post fertilization, dpf), and the antibody used identified a protein with a molecular weight of 70kDa, at 100dpf (regarded to be adult). Cell expressing SK1 in adult animals were neurons of dorsal root and cranial nerve sensory ganglia, sympathetic neurons, sensory cells in neuromasts of the lateral line system and taste buds, crypt olfactory neurons and photoreceptors. Present results report for the first time the expression and the distribution of SK1 in the peripheral nervous system and sensory organs of adult zebrafish, and may contribute to set zebrafish as an interesting experimental model for calcium-activated potassium channels research. Moreover these findings are of potential interest because the potential role of SK as targets for the treatment of neurological diseases and sensory disorders.

Neural crest contribution to lingual mesenchyme, epithelium and developing taste papillae and taste buds.

The epithelium of mammalian tongue hosts most of the taste buds that transduce gustatory stimuli into neural signals. In the field of taste biology, taste bud cells have been described as arising from "local epithelium", in distinction from many other receptor organs that are derived from neurogenic ectoderm including neural crest (NC). In fact, contribution of NC to both epithelium and mesenchyme in the developing tongue is not fully understood. In the present study we used two independent, well-characterized mouse lines, Wnt1-Cre and P0-Cre that express Cre recombinase in a NC-specific manner, in combination with two Cre reporter mouse lines, R26R and ZEG, and demonstrate a contribution of NC-derived cells to both tongue mesenchyme and epithelium including taste papillae and taste buds. In tongue mesenchyme, distribution of NC-derived cells is in close association with taste papillae. In tongue epithelium, labeled cells are observed in an initial scattered distribution and progress to a clustered pattern between papillae, and within papillae and early taste buds. This provides evidence for a contribution of NC to lingual epithelium. Together with previous reports for the origin of taste bud cells from local epithelium in postnatal mouse, we propose that NC cells migrate into and reside in the epithelium of the tongue primordium at an early embryonic stage, acquire epithelial cell phenotypes, and undergo cell proliferation and differentiation that is involved in the development of taste papillae and taste buds. Our findings lead to a new concept about derivation of taste bud cells that include a NC origin.

Patterns of immunoreactivity specific for gustducin and for NCAM differ in developing rat circumvallate papillae and their taste buds.

α-Gustducin and neural cell adhesion molecule (NCAM) are molecules previously found to be expressed in different cell types of mammalian taste buds. We examined the expression of α-gustducin and NCAM during the morphogenesis of circumvallate papillae and the formation of their taste buds by immunofluorescence staining and laser-scanning microscopy of semi-ultrathin sections of fetal and juvenile rat tongues. Images obtained by confocal laser scanning microscopy in transmission mode were also examined to provide outlines of histology and cell morphology. Morphogenesis of circumvallate papillae had already started on embryonic day 13 (E13) and was evident as the formation of placode. By contrast, taste buds in the circumvallate papillae started to appear between postnatal day 0 (P0) and P7. Although no cells with immunoreactivity specific for α-gustducin were detected in fetuses from E13 to E19, cells with NCAM-specific immunoreactivity were clearly apparent in the entire epithelium of the circumvallate papillary placode, the rudiment of each circumvallate papilla and the developing circumvallate papilla itself from E13 to E19. However, postnatally, both α-gustducin and NCAM became concentrated within taste cells as the formation of taste buds advanced. After P14, neither NCAM nor α-gustducin was detectable in the epithelium around the taste buds. In conclusion, α-gustducin appeared in the cytoplasm of taste cells during their formation after birth, while NCAM appeared in the epithelium of the circumvallate papilla-forming area. However, these two markers of taste cells were similarly distributed within mature taste cells.

Sox-2 in taste bud and lateral line system of zebrafish during development.

The Sox-2 is a transcription factor involved in adult neurogenesis in different vertebrate species, including fishes. Sox-2 also participates in growth and renewal on sensory cells in neuromasts of the fish lateral line system, and it is essential for development of taste buds in mammals. Using immunohistochemistry and Western blot we have investigated the occurrence and localization of Sox-2 taste buds and neuromast of zebrafish from 10 days post-fertilization to adult stage (1 year). The antibody used identifies two protein bands with estimated molecular weights of 34 and 37kDa which are consistent with those predicted for Sox-2. Sensory cells in taste buds displayed Sox-2 immunoreactivity at all the ages sampled, whereas in the neuromasts Sox-2 expression was restricted to the basal non-sensory cells. Interestingly Sox-2 immunoreactivity was observed in epithelial cells associated with both taste buds and neuromasts. Present results demonstrate that Sox-2 expressed in taste buds and neuromasts of zebrafish during the whole lifespan. Nevertheless, whereas the role of Sox-2 in taste buds of zebrafish remains to be established, the results in neuromast suggest that Sox-2 could participate in cell renewal of the mechanosensory cells.

Volumetry of human taste buds using laser scanning microscopy.

OBJECTIVE: In vivo laser scanning confocal microscopy is a relatively new, non-invasive method for assessment of oral cavity epithelia. The penetration depth of approximately 200-400 microm allows visualisation of fungiform papillae and their taste buds. MATERIALS AND METHODS: This paper describes the technique of in vivo volumetry of human taste buds. Confocal laser scanning microscopy used a diode laser at 670 nm for illumination. Digital laser scanning confocal microscopy equipment consisted of the Heidelberg Retina Tomograph HRTII and the Rostock Cornea Module. Volume scans of fungiform papillae were used for three-dimensional reconstruction of the taste bud. RESULTS: This technique supplied information on taste bud structure and enabled measurement and calculation of taste bud volume. Volumetric data from a 23-year-old man over a nine-day period showed only a small deviation in values. After three to four weeks, phenomenological changes in taste bud structures were found (i.e. a significant increase in volume, followed by disappearance of the taste bud and appearance of a new taste bud). CONCLUSIONS: The data obtained indicate the potential application of this non-invasive imaging modality: to evaluate variation of taste bud volume in human fungiform papillae with ageing; to study the effects of chorda tympani nerve transection on taste bud volume; and to demonstrate recovery of taste buds in patients with a severed chorda tympani nerve who show recovery of gustatory sensibility after surgery.

The morphological observations of some lingual papillae in the prenatal and prepuberal stages of red Sokoto goats (Capra hircus) / Observaciones morfológicas de algunas papilas linguales en las etapas prenatal y prepuberal de cabras rojas de Sokoto (Capra hircus)

This study was carried out to investigate the morphological development of the tongue in the foetal and prepubertal stages of Red Sokoto goats by light microscopy. In foetuses of about 50 days, the tongue tissues showed thickening of the epithelium into about 2-3 layers of cells. In fetuses of about 65 days, mesenchymal tissue was observed under the epithelium.Rudiments of some papillae were observed at this time. Collagenous fibre and blood vessels were scant in the lamina propria. In the 80-day-old foetuses, their was further differentiation of the epithelium rudiments into some papillae and this continued to mature until in foetuses of about 90 and 110 days, were early rudiments of taste buds were observed. Evidence of keratinization was apparent in the prepubertal stages.

El objetivo de este estudio fue investigar el desarrollo morfológico de la lengua en las etapas fetal y prepuberal de la cabra roj a de Sokoto por microscopía de luz. En los fetos de alrededor de 50 días, los tejidos linguales mostraron un engrosamiento del epitelio en cerca de 2-3 capas de células. En los fetos de alrededor de 65 días, se observó tejido mesenquimático bajo el epitelio. Rudimentos de algunas papilas se observaron en esta etapa. Fibras colágenas y vasos sanguíneos fueron observados de manera escasa en la lámina propria. En los 80 días de edad fetal, se observó la mayor diferenciación del epitelio con algunos rudimentos de papilas, lo que continuó hasta la maduración de los fetos, alrededor de los 90 y 110 días, donde fueron observados de manera temprana rudimentos de botones gustativos. Evidencia de queratinización fue evidente en las etapas prepuberales.

Wnt signaling interacts with Shh to regulate taste papilla development.

Wnt and Shh signaling pathways are critical for the development and maturation of many epithelial tissues. Both pathways have roles in stem cell maintenance, tissue development, and tumorigenesis. However, linkage between these pathways in mammalian systems had not been well established. Here, we report that Shh expression in fungiform papillae and formation of normal mature fungiform papillae depend on signaling through Wnt and beta-catenin. We observed that during fungiform papilla formation in mice, Shh and components of the Wnt/beta-catenin signaling pathway are expressed together in the developing placode. The elimination of Wnt/beta-catenin signaling in either Lef1 or Wnt10b knockout mice resulted in down-regulation of Shh expression. In addition, the size and number of fungiform papillae were greatly reduced in Lef1 knockout mice. By examining embryonic mouse tongues in culture we determined that activation of Wnt/beta-catenin signaling up-regulates Shh expression. We observed that blocking Shh signaling in cultured tongue explants enhanced papillae formation and was accompanied by an up-regulation of Wnt/beta-catenin signaling, indicating that Shh inhibits the Wnt/beta-catenin pathway. Exogenously added Shh suppressed expression of endogenous Shh and inhibited Wnt/beta-catenin signaling (assessed in TOPGAL mice), further implicating Shh as an inhibitor of the Wnt/beta-catenin pathway. Our observations indicate that Wnt/beta-catenin signaling and interactions between the Wnt and Shh pathways play essential roles in the development of fungiform papillae.

Modularity and sense organs in the blind cavefish, Astyanax mexicanus.

Mexican tetra (Astyanax mexicanus) exist as two morphs: a sighted (surface) form and a blind (cavefish) form. In the cavefish, some modules are lost, such as the eye and pigment modules, whereas others are expanded, such as the taste bud and cranial neuromast modules. We suggest that modularity can be viewed as being nested in a manner similar to Baupläne so that modules express unique sets of genes, cells, and processes. In terms of evolution, we conclude that natural selection can act on any of these hierarchical levels within modules or on all the sensory modules as a whole. We discuss interactions within and between modules with reference to the blind cavefish from both genetic and developmental perspectives. The cavefish represents an illuminating example of module interaction, uncoupling of modules, and module expansion.

Development of fungiform papillae: patterned lingual gustatory organs.

The fungiform papilla is a gustatory organ that provides a specific tissue residence for taste buds on the anterior tongue. Thus, during development there must be a progressive differentiation to acquire papilla epithelium, then taste cell progenitor epithelium, and finally taste cells within the papilla apex. Arranged in rows, the patterned distribution of fungiform papillae requires molecular regulation not only to induce papillae, but also to suppress papilla formation in the between-papilla tissue. Intact sensory innervation is not required to initiate papilla development or pattern. However, members of several molecular families have now been identified with specific localization in developing papillae. These may participate in papilla development and pattern formation, and subsequently in taste progenitor and taste cell differentiation. This review focuses on development of fungiform papillae in embryonic rat and mouse. Basic morphology, cell biology and molecular phenotypes of developing papillae are reviewed. Regulatory roles for molecules in several families are presented, and a broad schema is proposed for progressive epithelial differentiation to form taste cell progenitors in parallel with the temporal course, and participation of lingual sensory innervation.

Effect of the feeding pattern on rumen wall morphology of cows and sheep

Nutritional manipulation of the rumen wall volatile fatty acid absorption capacity can be a strategy to control ruminal acidosis in dairy cows. Aiming to induce morphological rumen wall variation through diet and to establish efficient papillae morphological markers 2 experiments were performed. In experiment 1, seven rumen-cannulated cows were fed with concentrate 1 or 4 times a day for 19 days followed by fasting for 72 hours. Ruminal papillae were collected on days, 0, 4, 12 and 19 of the treatment period, and 24, 48 and 72 hours after onset of fasting which was able to induce papilar involution. Lower concentrate feeding frequency was associated to insulin increasing over time (P=0,02) and higher (P=0,03) mitotic index (MI), but it did not affect other morphological parameters. In experiment 2, two non-simultaneous trials with 3 rumen-cannulated ovines in each, were conducted and animals were fasted abruptly for 72 hours after feeding. Papillae were collected at the end of the feeding period and at the end of the 72 hour fasting period. MI was higher in the feeding period than the in fasting period (P<0,01), but other morphological parameters were not able to respond to nutritional variation. Among the morphologic markers studied MI seems to be the best variable for evaluation of the rumen epithelium morphologic response to feeding plans. Frequency of concentrate feeding may be used to regulate rumen papillae morphology.

A manipulação nutricional da capacidade de absorção de ácidos graxos voláteis pela parede do rúmen pode ser uma estratégia para controlar acidose em vacas leiteiras. Objetivando induzir variação morfológica da parede do rúmen através da dieta e estabelecer marcadores morfológicos eficientes para epitélio ruminal, dois experimentos foram relizados. No experimento um, sete vacas com cânula ruminal foram alimentadas com concentrado uma ou quatro vezes ao dia por 19 dias seguidos por 72 horas de jejum. Papilas ruminais foram coletadas nos dias zero, quatro, 12 e 19 do período de tratamento e 24, 48 e 72 horas após o início do período de jejum. Baixa freqüência de alimentação concentrada foi associada a um aumento de insulina plasmática através do tempo (P=0,02) e a um maior (P=0,03) índice mitótico (IM), mas não afetou outros parâmetros morfológicos. No experimento dois, foram realizados dois ensaios não-simultâneos com três ovinos canulados no rúmen, os quais foram submetidos abruptamente a 72 horas de jejum. Papilas ruminais foram coletadas no final do período de alimentação e no final do jejum. O IM foi mais alto no período de alimentação do que no período de jejum (P<0,01), mas outros parâmetros morfológicos não foram capazes de responder à variação nutricional. Entres os marcadores morfológicos estudados o IM parece ser a melhor variável para avaliação da resposta morfológica do epitélio ao plano alimentar. A freqüência de alimentação concentrada pode ser usada para regular a morfologia das papilas ruminais.