The functional role of the T1R family of receptors in sweet taste and feeding

The discovery of the T1R family of Class C G protein-coupled receptors in the peripheral gustatory system a decade ago has been a tremendous advance for taste research, and its conceptual reach has extended to other organ systems. There are three proteins in the family, T1R1, T1R2, and T1R3, encoded by their respective genes, Tas1r1, Tas1r2, and Tas1r3. T1R2 combines with T1R3 to form a heterodimer that binds with sugars and other sweeteners. T1R3 also combines with T1R1 to form a heterodimer that binds with l-amino acids. These proteins are expressed not only in taste bud cells, but one or more of these T1Rs have also been identified in the nasal epithelium, gut, pancreas, liver, kidney, testes and brain in various mammalian species. Here we review current perspectives regarding the functional role of these receptors, concentrating on sweet taste and feeding. We also discuss behavioral findings suggesting that a glucose polymer mixture, Polycose, which rodents avidly prefer, appears to activate a receptor that does not depend on the combined expression of T1R2 and T1R3. In addition, although the T1Rs have been implicated as playing a role in glucose sensing, T1R2 knock-out (KO) and T1R3 KO mice display normal chow and fluid intake as well as normal body weight compared with same-sex littermate wild type (WT) controls. Moreover, regardless of whether they are fasted or not, these KO mice do not differ from their WT counterparts in their Polycose intake across a broad range of concentrations in 30-minute intake tests. The functional implications of these results and those in the literature are considered.     

Chronic restraint stress in rats suppresses sweet and umami taste responses and lingual expression of T1R3 mRNA

Effects of chronic restraint stress on the taste responses to five basic taste qualities were investigated electrophysiologically in the rat chorda tympani. In addition, the mRNA expression for T1R3, the common G-protein-coupled receptor (GPCR) for sweet and umami tastes, was studied quantitatively by RT-PCR after such stress. Rats were restrained in a small cylindrical restrainer made of steel wire for 8h daily for 14 successive days. The integrated responses to sweet and umami tastes, as recorded from the chorda tympani, were significantly suppressed after such stress, but the other three basic taste responses were unaffected. Expression of T1R3 mRNA in the fungiform papillae, as estimated by RT-PCR, was slightly reduced by the stress, and a quantitative real time RT-PCR study revealed a significant suppression of T1R3 mRNA expression in the stress group. These results suggest that the observed stress-induced changes in taste sensation could be caused by a peripheral disorder of the transduction mechanism in taste-receptor cells, involving in particular a stress-induced inhibition of T1R3 expression.     

A candidate taste receptor gene near a sweet taste locus

The mechanisms underlying sweet taste in mammals have been elusive. Although numerous studies have implicated G proteins in sweet taste detection, the expected G protein-coupled receptors have not been found. Here we describe a candidate taste receptor gene, T1r3, that is located at or near the mouse Sac locus, a genetic locus that controls the detection of certain sweet tastants. T1R3 differs in amino acid sequence in mouse strains with different Sac phenotypes ('tasters' versus 'nontasters'). In addition, a perfect correlation exists between two different T1r3 alleles and Sac phenotypes in recombinant inbred mouse strains. The T1r3 gene is expressed in a subset of taste cells in circumvallate, foliate and fungiform taste papillae. In circumvallate and foliate papillae, most T1r3-expressing cells also express a gene encoding a related receptor, T1R2, raising the possibility that these cells recognize more than one ligand, or that the two receptors function as heterodimers.     

Sucrose taste but not Polycose taste conditions flavor preferences in rats

Rats have an inborn preference for sweet taste and learn to prefer flavors associated with sweetness. They are also strongly attracted to the taste of glucose polymers (e.g., Polycose). This "poly" taste differs in quality from the sweet taste of sugar. To determine if poly taste, like sweet taste, conditions flavor preferences rats were trained with a distinctive flavor (CS+) added to 2% Polycose solution and a different flavor (CS-) added to plain water. In a subsequent two-bottle test the rats did not prefer the CS+ to CS- when both flavors were presented in water. In contrast, other rats significantly preferred a CS+ flavor that had been paired with 2% sucrose. Adding saccharin to a flavored Polycose solution did not improve CS+ flavor learning; rather, Polycose appeared to overshadow saccharin-induced conditioning. Flavor conditioning by a 16% Polycose solution was assessed using a sham-feeding procedure to prevent post-oral reinforcement. Although the rats sham-fed substantial amounts of the CS+ flavored Polycose solution, they failed to prefer the CS+ to the CS- flavor. This contrasts with the preference other rats displayed for a CS+ paired with sham-fed sucrose. Why attractive sweet and poly tastes differ in their ability to condition flavor preferences is not certain, although some findings suggest that they differentially activate dopamine and/or serotonin circuits involved in flavor learning.     

Decreased expression of CD36 in circumvallate taste buds of high-fat diet induced obese rats

Mammals spontaneously prefer lipid rich foods. Overconsumption of high-fat diet leads to obesity and related diseases. Recent findings indicate that taste may participate in the orosensory perception of dietary lipids and the fatty taste may contribute to a preference for and excessive consumption of dietary fat. CD36, a trans-membrane glycoprotein, which is located in the taste buds of circumvallate papillae of rodents, appears to be a plausible receptor for this fatty taste. Obese subjects present a stronger preference for fatty foods, though the mechanisms involved are complex and are not fully investigated. Our data from immunofluorescence and real-time RT-PCR showed that the expression levels of CD36 in circumvallate taste buds were significantly lower in high-fat diet induced obese rats as compared with that of control rats fed a normal diet. These results suggest that decreased expression of CD36 in circumvallate taste buds of high-fat diet induced obese rats may be associated with diminished fatty taste sensitivity and in order to compensate the preference for dietary fat, rats consume more fatty foods. Therapeutic strategies designed to alter or manipulate CD36 expression or function in taste buds may have important implications in treating obesity and related diseases.     

云豹味蕾的分布与结构

目的：了解云豹味觉器官味蕾在口腔中的分布和结构。方法：用光镜观察云豹舌的形态结构和味蕾的分布,并用透射电镜观察味蕾的结构。结果：云豹味蕾分布于舌尖及轮廓乳突的上皮层中,主要由明细胞和暗细胞组成。结论：云豹味蕾的分布及舌的形态学特点与它的捕食和吞咽习性相适应。     

Distribution of taste buds on fungiform and circumvallate papillae of bovine tongue

The distribution of taste buds on the fungiform and circumvallate papillae of the cow tongue has been determined. The two tongues studied were from Holstein-Friesian cows four to six years of age; they contained 14,765 and 21,691 taste buds, respectively. The tip of the tongue is well supplied with fungiform papillae, and the posterior portion contains the circumvallate papillae. The midportion of the tongue contains relatively few taste papillae. The fungiform papillae contained 1,580 and 1,838 taste buds on the two tongues, respectively, and the circumvallate papillae were estimated to contain 13,185 and 19,853 taste buds. The highest concentration of taste buds therefore occurs in the circumvallate papillae; these relatively few papillae contain approximately 90% of the taste buds. On a circumvallate papilla, taste buds are found only on the papillary sidewall, with none either on the apical surface of the papilla or on the outer wall of the moat.     

Estrogen increases the taste threshold for sucrose in rats

Anecdotal and empirical evidence suggests that females' preferences for sweet foods are affected by hormonal fluctuations across the reproductive cycle. In rats, the preference for sweet foods may involve estrogen-mediated changes in response to the taste of sweets. Our recent work showed that ovariectomized female rats lick less to dilute sucrose solutions when given estrogen than when given the oil vehicle. These findings suggest that estrogen decreases the preference for less concentrated sucrose solutions; however, an alternative explanation is that estrogen interferes with the ability to detect dilute sucrose solutions. To distinguish between these possibilities, we conditioned a taste aversion to 0.2 M sucrose in ovariectomized rats by pairing it with injection of LiCl and then examined the generalization of that taste aversion to 0.075 and 0.025 M sucrose solutions during estrogen or oil treatment. Oil-treated rats generalized the LiCl-induced aversion conditioned to 0.2 M sucrose to both 0.075 and 0.025 M sucrose. Estrogen-treated rats generalized the LiCl-induced taste aversion to 0.075 M sucrose but not to 0.025 M sucrose. Moreover, two weeks later, when estrogen had cleared the system, both groups generalized the aversion to both 0.075 and 0.025 M sucrose. These results show that estrogen affects the ability to discriminate dilute sucrose from water and suggest that estrogen may have short-term effects on the detection threshold for sucrose taste in rats.     

Pharyngeal arch deficiencies affect taste bud development in the circumvallate papilla with aberrant glossopharyngeal nerve formation

Background: The pharyngeal arches (PAs) generate cranial organs including the tongue. The taste placodes, formed in particular locations on the embryonic tongue surface, differentiate into taste buds harbored in distinct gustatory papillae. The developing tongue also has a complex supply of cranial nerves through each PA. However, the relationship between the PAs and taste bud development is not fully understood. Results: Ripply3 homozygous mutant mice, which have impaired third/fourth PAs, display a hypoplastic circumvallate papilla and lack taste buds, although the taste placode is normally formed. Formation of the glossopharyngeal ganglia is defective and innervation toward the posterior tongue is completely missing in Ripply3 mutant embryos at E12.5. Moreover, the distribution of neuroblasts derived from the epibranchial placode is severely, but not completely, atenuated, and the neural crest cells are diminished in the third PA region of Ripply3 mutant embryos at E9.5–E10.5. In Tbx1 homozygous mutant embryos, which exhibit another type of deficiency in PA development, the hypoplastic circumvallate papilla is observed along with abnormal formation of the glossopharyngeal ganglia and severely impaired innervation. Conclusions: PA deficiencies affect multiple aspects of taste bud development, including formation of the cranial ganglia and innervation to the posterior tongue. Developmental Dynamics 244:874–887, 2015. © 2015 Wiley Periodicals, Inc.     

Sweet taste of dilute NaCl: Psychophysical evidence for a sweet stimulus

Dilute NaCl tastes sweet. This sweetness could result from coding confusions in the nervous system such that weak NaCl produces neural signals resembling those for sweeteners like sucrose. On the other hand, an analysis of the structural chemistry of water-salt interactions suggests that water shells organized around cations may actually provide a sweet stimulus indistinguishable (to the receptor molecules) from more conventional sweet stimuli. In the present experiment, the sweetness of weak NaCl was abolished in two ways: by adaptation to sucrose and by topical application of Gymnema sylvestre. Since these two operations also abolish the sweet taste of a variety of conventional sweetners, the sweet taste of NaCl appears to result from the presence of a sweet stimulus.     

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.     

Expression of alpha-gustducin in the circumvallate papillae of taste buds of diabetic rats

Taste impairment is a complication of Diabetes mellitus and some studies have shown this taste disorder in diabetes. Diabetes can decrease the ability of individuals to detect and recognize sweet, salty and bitter tastes. alpha-Gustducin is a transducin-like G-protein selectively expressed in 20 - 30% of taste receptor cells, which has been shown to be involved in bitter, sweet and umami taste responses. The present study was performed to explore the protein and mRNA expression of alpha-gustducin in the taste buds of diabetic and control rat circumvallate papillae. Our results showed that the positive expression of alpha-gustducin in diabetic rat taste bud cells is higher than that in normal controls as shown by both immunohistochemistry and RT-PCR. There may be some variant of bitter, sweet or umami taste transduction during diabetes and that taste transduction variant may be one cause of diabetic taste impairment.     

Isolation and characterization of porcine circumvallate papillae cells

Animal food intake is primarily controlled by appetite, which is affected by food quality, environment, and the management and status of animal health. Sensing of taste is mediated by taste receptor cells and is central to appetite. Taste receptor cells possess distinctive physiological characteristics that permit the recognition of various stimuli in foods. Thus, cultures of porcine circumvallate papillae cells provide a model for identification of the molecular and functional characteristics of taste receptor cells. In this study, we described the isolation and culture of porcine circumvallate papillae, using tissue explants and enzymatic digestion, and showed continuous viability and expression of pivotal taste marker proteins for more than 9 passages. In addition, cultured cells showed dramatic rises in intracellular calcium upon stimulation with several taste stimuli (sweet, umami, bitter, and fat). These cultures of porcine taste receptor cells provide a useful model for assessing taste preferences of pigs and may elucidate interactions between various taste stimuli.     

Apical CD36 immunolocalization in human and porcine taste buds from circumvallate and foliate papillae

CD36 is the receptor for long chain fatty acids (LCFA), and is expressed in lingual taste cells from rodents. In these animals, CD36 has been proposed to play an important role in oral detection of LCFA, and subsequently, determines their dietary fat preference. Humans also seem to detect LCFA in the oral cavity, however, information on the molecular mechanism of this human orosensory LCFA recognition is currently lacking. The aim of our study was to investigate whether CD36 is also expressed in lingual human and porcine taste buds cells. Using fluorescence immunohistochemistry, apical CD36 expression was revealed in human and porcine taste bud cells from circumvallate and foliate papillae. These data suggest CD36 as the putative orosensory receptor for dietary LCFA in human, and, therefore, may be involved in our preference for fatty foods.     

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

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

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 (IP₃R₃)_, α-gustducin, protein-specific gene product 9.5 (PGP9.5)] and a subpopulation of type III (IP₃R₃, PGP9.5) taste cells. On average, 72%, 50%, and 32% of the espin-positive taste cells were labeled with antibodies to IP₃R₃, α-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 ∼3 μm-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 μm 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.     

Taste discriminations in rats lacking anterior insular gustatory neocortex

Previous neurobehavioral investigations have demonstrated that the anterior insular gustatory neocortex (AIGN) mediates taste-illness learning. The present experiment evaluated taste discriminations in rats lacking AIGN. Two groups of rats received distinct surgical treatments. One-half of the animals received bilateral electrolytic lesion placements in the AIGN: Remaining animals received anesthesia and scalp incisions only. Following postoperative recovery animals received standard two-bottle preference tests with various concentrations of sucrose to evaluate gustatory reactivity. Animals thereafter received two-bottle discrimination tests with selected sucrose concentrations. At the conclusion of preference tests and discrimination tests with sucrose, preference tests and discrimination tests were conducted with sodium chloride. Following those tests animals received taste aversion conditioning to determine whether or not AIGN lesions impaired taste-illness learning. Results of two-bottle taste tests indicated that AIGN lesions do not obviously alter taste reactivity nor taste discriminations to preferred concentrations of sucrose and NaCl. Anterior insular lesions did, however, impair normal taste aversion learning. These results, in combination with those of previous investigators, provide further evidence that the AIGN preferentially contributes to taste learning functions.     

Discrimination of taste qualities among mouse fungiform taste bud cells

Multiple lines of evidence from molecular studies indicate that individual taste qualities are encoded by distinct taste receptor cells. In contrast, many physiological studies have found that a significant proportion of taste cells respond to multiple taste qualities. To reconcile this apparent discrepancy and to identify taste cells that underlie each taste quality, we investigated taste responses of individual mouse fungiform taste cells that express gustducin or GAD67, markers for specific types of taste cells. Type II taste cells respond to sweet, bitter or umami tastants, express taste receptors, gustducin and other transduction components. Type III cells possess putative sour taste receptors, and have well elaborated conventional synapses. Consistent with these findings we found that gustducin-expressing Type II taste cells responded best to sweet (25/49), bitter (20/49) or umami (4/49) stimuli, while all GAD67 (Type III) taste cells examined (44/44) responded to sour stimuli and a portion of them showed multiple taste sensitivities, suggesting discrimination of each taste quality among taste bud cells. These results were largely consistent with those previously reported with circumvallate papillae taste cells. Bitter-best taste cells responded to multiple bitter compounds such as quinine, denatonium and cyclohexamide. Three sour compounds, HCl, acetic acid and citric acid, elicited responses in sour-best taste cells. These results suggest that taste cells may be capable of recognizing multiple taste compounds that elicit similar taste sensation. We did not find any NaCl-best cells among the gustducin and GAD67 taste cells, raising the possibility that salt sensitive taste cells comprise a different population.