Effect of salivary gland removal on taste preference in mice.

To evaluate the effect of decreased salivary secretion on taste preference, we investigated taste preference for five basic tastes by a 48 h two-bottle preference test using a mouse model (desalivated mice) that underwent surgical removal of three major salivary glands: the parotid, submandibular, and sublingual glands. In the desalivated mice, the avoidance behaviors for bitter and salty tastes and the attractive behaviors for sweet and umami tastes were significantly decreased. We confirmed that saliva is necessary to maintain normal taste preference. To estimate the cause of the preference changes, we investigated the effects of salivary gland removal on the expression of taste-related molecules in the taste buds. No apparent changes were observed in the expression levels or patterns of taste-related molecules after salivary gland removal. When the protein concentration and composition in the saliva were compared between the control and desalivated mice, the protein concentration decreased and its composition changed after major salivary gland removal. These results suggest that changes in protein concentration and composition in the saliva may be one of the factors responsible for the changes in taste preferences observed in the desalivated mice.

Allosteric modulation of the fish taste receptor type 1 (T1R) family by the extracellular chloride ion.

Many G protein-coupled receptors (GPCRs) are allosterically modulated by inorganic ions. Although the intraoral ionic composition of the oral cavity varies depending on the living environment and feeding behavior, little is known about whether and how it affects the function of taste receptor type 1 (T1R), a member of the class C GPCR family. Here, we report that chloride ions allosterically modulate the functions of specific fish T1Rs, namely, mfT1R2a/mfT1R3 and zfT1R2a/zfT1R3. Site-directed mutagenesis revealed mfT1R2a K265, which lies in the extracellular domain of mfT1R2a, to be as a critical residue for the modulation of mfT1R2a/mfT1R3 by Cl-. However, this residue is not conserved in zfT1R2a, and the introduction of the key residue at the corresponding site of another T1R, mfT1R2b, did not confer Cl- susceptibility. These results indicate the variability of the determinants of Cl- susceptibility.

Modeling the structure of the transmembrane domain of T1R3, a subunit of the sweet taste receptor, with neohesperidin dihydrochalcone using molecular dynamics simulation.

Neohesperidin dihydrochalcone (NHDC) is a sweetener, which interacts with the transmembrane domain (TMD) of the T1R3 subunit of the human sweet taste receptor. Although NHDC and a sweet taste inhibitor lactisole share similar structural motifs, they have opposite effects on the receptor. This study involved the creation of an NHDC-docked model of T1R3 TMD through mutational analyses followed by in silico simulations. When certain NHDC derivatives were docked to the model, His7345.44 was demonstrated to play a crucial role in activating T1R3 TMD. The NHDC-docked model was then compared with a lactisole-docked inactive form, several residues were characterized as important for the recognition of NHDC; however, most of them were distinct from those of lactisole. Residues such as His6413.33 and Gln7947.38 were found to be oriented differently. This study provides useful information that will facilitate the design of sweeteners and inhibitors that interact with T1R3 TMD.

Zebrafish and medaka T1R (taste receptor type 1) proteins mediate highly sensitive recognition of l-proline.

In vertebrates, nutritional tastants, such as amino acids and sugars, are recognized by G-protein-coupled receptors of the taste receptor type 1 (T1R) family. Previous studies have shown that fish T1Rs are functionally distinct from mammalian T1Rs in certain regards. Here, we report the existence of oral receptors with high sensitivity to amino acids in zebrafish and medaka fish. We describe the construction of multiple cell lines stably expressing functional T1Rs (from medaka fish or zebrafish) with a chimeric G-protein (G16gust44) using the Flp-In system. Through functional assays with these cell lines, medaka fish and zebrafish were confirmed to possess particular T1Rs highly sensitive to l-proline, possibly reflecting the physiological importance of l-proline in teleosts, in line with previous studies.

Identification of mouse bitter taste receptors that respond to resveratrol: a bitter-tasting polyphenolic compound.

The mouse bitter taste receptors (Tas2rs) that respond to resveratrol, a bitter-tasting polyphenolic compound, were identified. Among 35 members of the Tas2r family, Tas2r108, 109, 131, and 137 responded to resveratrol treatment. mRNA expression levels of Tas2r108 and Tas2r137 were higher than those of Tas2r109 and Tas2r131 in mouse circumvallate papillae, indicating that Tas2r108 and Tas2r137 may play important roles in detecting the bitterness of resveratrol in the oral cavity. The mRNA expression levels of Tas2r137 and Tas2r108 were also observed in several tissues, suggesting that Tas2r108 and Tas2r137 may also be involved in the physiological action of resveratrol.

Expression of Olfactory-Related Genes in the Olfactory Epithelium of an Alzheimer's Disease Mouse Model.

Using an amyloid precursor protein (App) gene knock-in (KI) mouse of Alzheimer's disease (AD), we investigated the expression of olfactory-related genes in olfactory impairment caused by AD. We observed the change in olfactory behavior in the App-KI mice. There was no significant difference, however, in the mRNA expression levels of olfactory-related genes between the olfactory epithelia of wild-type (WT) and App-KI mice. Amyloid-ß deposition was confirmed throughout the olfactory pathway in App-KI mice, but not in WT mice. These show that the change in olfactory behavior in the App-KI mice might cause by the impairment of the olfactory pathway.

Transmembrane channel-like 4 is involved in pH and temperature-dependent modulation of salty taste.

Human susceptibility to NaCl varies depending on temperature and pH, the molecular mechanisms of which remain unclear. The voltage-dependent chloride channel, transmembrane channel-like 4 (TMC4), is activated at approximately 40 °C and is suppressed at pH 5.5. As these are similar in character to human sensory evaluations, human TMC4 may be involved in human salt taste reception.

TMC4 is a novel chloride channel involved in high-concentration salt taste sensation.

"Salty taste" sensation is evoked when sodium and chloride ions are present together in the oral cavity. The presence of an epithelial cation channel that receives Na+ has previously been reported. However, no molecular entity involving Cl- receptors has been elucidated. We report the strong expression of transmembrane channel-like 4 (TMC4) in the circumvallate and foliate papillae projected to the glossopharyngeal nerve, mediating a high-concentration of NaCl. Electrophysiological analysis using HEK293T cells revealed that TMC4 was a voltage-dependent Cl- channel and the consequent currents were completely inhibited by NPPB, an anion channel blocker. TMC4 allowed permeation of organic anions including gluconate, but their current amplitudes at positive potentials were less than that of Cl-. Tmc4-deficient mice showed significantly weaker glossopharyngeal nerve response to high-concentration of NaCl than the wild-type littermates. These results indicated that TMC4 is a novel chloride channel that responds to high-concentration of NaCl.

Evolution of the primate glutamate taste sensor from a nucleotide sensor.

Taste perception plays an essential role in food selection. Umami (savory) tastes are sensed by a taste receptor complex, T1R1/T1R3, that detects proteinogenic amino acids.1 High sensitivity to l-glutamate (l-Glu) is a characteristic of human T1R1/T1R3, but the T1R1/T1R3 of other vertebrates does not consistently show this l-Glu response.1,2 Here, we demonstrate that the l-Glu sensitivity of T1R1/T1R3 is a derived state that has evolved repeatedly in large primates that rely on leaves as protein sources, after their divergence from insectivorous ancestors. Receptor expression experiments show that common amino acid substitutions at ligand binding sites that render T1R1/T1R3 sensitive to l-Glu occur independently at least three times in primate evolution. Meanwhile T1R1/T1R3 senses 5'-ribonucleotides as opposed to l-Glu in several mammalian species, including insectivorous primates. Our chemical analysis reveal that l-Glu is one of the major free amino acids in primate diets and that insects, but not leaves, contain large amounts of free 5'-ribonucleotides. Altering the ligand-binding preference of T1R1/T1R3 from 5'-ribonucleotides to l-Glu might promote leaf consumption, overcoming bitter and aversive tastes. Altogether, our results provide insight into the foraging ecology of a diverse mammalian radiation and help reveal how evolution of sensory genes facilitates invasion of new ecological niches.

Modulatory Effect of Theaflavins on Apical Sodium-Dependent Bile Acid Transporter (ASBT) Activity.

Inhibiting apical sodium-dependent bile acid transporter (ASBT) has been identified as a potential strategy to reduce plasma cholesterol levels. Thus, in this study, we aimed to identify polyphenols that inhibited ASBT activity and to elucidate their mechanism. ASBT is responsible for most of the taurocholic acid (TC) uptake in Caco-2 cells. Of the 39 polyphenols examined, theaflavin (TF)-3-gallate (TF2A) and theaflavin-3'-gallate (TF2B) have been found to significantly reduce TC uptake in Caco-2 cells to 37.4 ± 2.8 and 33.8 ± 4.0%, respectively, of that in the untreated cells. The results from the TC uptake assay using N-acetylcysteine suggested that the inhibitory effect of TF2A and TF2B was attributed to the oxidization of their benzotropolone rings and their covalent bonding with ASBT's cysteine. TC uptake was reduced in the COS-7 cells expressing recombinant ASBT whose cysteine residues were mutated to alanine. Finally, the substrate concentration-dependent TC uptake assay showed that TFs competitively inhibited TC uptake.

Ibuprofen inhibits oral NaCl response through transmembrane channel-like 4.

Nonsteroidal anti-inflammatory drugs, such as ibuprofen, are known to modify salty taste perception in humans. However, the underlying molecular mechanisms remain unknown. We investigated the inhibitory effect of ibuprofen on the NaCl stimulation of epithelium sodium channel (ENaC) and transmembrane channel-like 4 (TMC4), which are involved in salty taste detection. Although ibuprofen only minimally inhibited the response of the ENaC to NaCl, it significantly inhibited the TMC4 response to NaCl with an IC50 at 1.45 mM. These results suggest that ibuprofen interferes with detection of salty taste via inhibition of TMC4.

Change in Taste Preference to Capsaicin and Catechin Due to Aging in Mice.

Taste is a chemical sensation that primarily detects nutrients present in food, and maintenance of taste sensations is important for ensuring that older people have a balanced nutritional diet. While several reports have suggested that taste sensitivity changes with age, the molecular mechanisms underlying this phenomenon are still unclear. Previous studies on the matter have focused mainly on the relationship between aging and taste detection of specific basic taste-inducing substances, and other than for these basic substances, understanding of how aging affects the detection of taste is limited. Therefore, to understand the effect that aging has on the taste detection of some familiar substances found in our daily meals, namely capsaicin and catechin, we investigated age-related changes in taste preferences to capsaicin and catechin in young and old C57BL/6J mice using a 48-h two-bottle preference test. For the capsaicin stimuli, the mice showed avoidance behavior in a concentration-dependent manner. However, we observed that there was no significant difference in the preference ratio for capsaicin between young and old mice. For the catechin stimuli, although both age groups showed avoidance behavior in a concentration-dependent manner, the preference ratio in old mice showed significantly higher values than those in young mice. This suggests that catechin sensitivity is declined due to aging. Thus, we observed that catechin sensitivity decreases with age, but capsaicin sensitivity does not.

Early origin of sweet perception in the songbird radiation.

Early events in the evolutionary history of a clade can shape the sensory systems of descendant lineages. Although the avian ancestor may not have had a sweet receptor, the widespread incidence of nectar-feeding birds suggests multiple acquisitions of sugar detection. In this study, we identify a single early sensory shift of the umami receptor (the T1R1-T1R3 heterodimer) that conferred sweet-sensing abilities in songbirds, a large evolutionary radiation containing nearly half of all living birds. We demonstrate sugar responses across species with diverse diets, uncover critical sites underlying carbohydrate detection, and identify the molecular basis of sensory convergence between songbirds and nectar-specialist hummingbirds. This early shift shaped the sensory biology of an entire radiation, emphasizing the role of contingency and providing an example of the genetic basis of convergence in avian evolution.

De novo transcriptome analysis and comparative expression profiling of genes associated with the taste-modifying protein neoculin in Curculigo latifolia and Curculigo capitulata fruits.

BACKGROUND: Curculigo latifolia is a perennial plant endogenous to Southeast Asia whose fruits contain the taste-modifying protein neoculin, which binds to sweet receptors and makes sour fruits taste sweet. Although similar to snowdrop (Galanthus nivalis) agglutinin (GNA), which contains mannose-binding sites in its sequence and 3D structure, neoculin lacks such sites and has no lectin activity. Whether the fruits of C. latifolia and other Curculigo plants contain neoculin and/or GNA family members was unclear. RESULTS: Through de novo RNA-seq assembly of the fruits of C. latifolia and the related C. capitulata and detailed analysis of the expression patterns of neoculin and neoculin-like genes in both species, we assembled 85,697 transcripts from C. latifolia and 76,775 from C. capitulata using Trinity and annotated them using public databases. We identified 70,371 unigenes in C. latifolia and 63,704 in C. capitulata. In total, 38.6% of unigenes from C. latifolia and 42.6% from C. capitulata shared high similarity between the two species. We identified ten neoculin-related transcripts in C. latifolia and 15 in C. capitulata, encoding both the basic and acidic subunits of neoculin in both plants. We aligned these 25 transcripts and generated a phylogenetic tree. Many orthologs in the two species shared high similarity, despite the low number of common genes, suggesting that these genes likely existed before the two species diverged. The relative expression levels of these genes differed considerably between the two species: the transcripts per million (TPM) values of neoculin genes were 60 times higher in C. latifolia than in C. capitulata, whereas those of GNA family members were 15,000 times lower in C. latifolia than in C. capitulata. CONCLUSIONS: The genetic diversity of neoculin-related genes strongly suggests that neoculin genes underwent duplication during evolution. The marked differences in their expression profiles between C. latifolia and C. capitulata may be due to mutations in regions involved in transcriptional regulation. Comprehensive analysis of the genes expressed in the fruits of these two Curculigo species helped elucidate the origin of neoculin at the molecular level.

Comparison between the timing of the occurrence of taste sensitivity changes and short-term memory decline due to aging in SAMP1 mice.

Several studies have suggested that cognitive impairment affects taste sensitivity. However, the mechanism behind this is still unclear. In this study, we focused on short-term memory. Using senescence-accelerated mouse prone 1 (SAMP1) mice, we compared whether the effects of aging are observed earlier in taste sensitivity or short-term memory. We used 8-week-old mice as the young group, and 70- and 80-week-old mice as aged groups. Taste sensitivity was evaluated using a 48-hour two-bottle preference test, and short-term memory was evaluated using the Y-maze test. SAMP1 mice showed apparently changes in taste sensitivity at 70-weeks-old. However, the influence of aging on spontaneous alternation behavior, which is indicative of short-term memory alterations, was not observed in 70-week-old mice. At 80-weeks-old, the influence of aging was observed, and spontaneous alternation behavior was significantly decreased. This suggests that age-dependent changes in taste sensitivity occur prior to short-term memory function decline. In addition, there was no significant influence of aging on the mRNA expression of long-term potentiation-related genes in the hippocampus of 80-week-old mice. Therefore, the age-related decline of short-term memory may not affect taste sensitivity.

Characterization of the Human Bitter Taste Receptor Response to Sesquiterpene Lactones from Edible Asteraceae Species and Suppression of Bitterness through pH Control.

Vegetables are important sources of nutrients and bioactive compounds; however, their consumption is often insufficient, partly because of unpleasant taste characteristics. This study aimed to investigate the mechanisms underlying bitter taste reception and to develop methods to suppress bitterness. We focused on sesquiterpene lactones found in edible Asteraceae species. HEK293T cells that heterologously expressed human bitter taste receptors (including TAS2R46) together with a chimeric G protein were analyzed using calcium imaging, and cellular responses to four sesquiterpene lactones contained in lettuce were examined. We found that TAS2R46-expressing cells responded most strongly to bitter compounds. The EC50 value of 11ß,13-dihydrolactucopicrin was 2.0 ± 0.6 µM, in agreement with the previously reported bitterness threshold of the compound. Adjustment of pH from neutral to weak acidic conditions reduced the response of TAS2R46-expressing cells to sesquiterpene lactones. We demonstrate the possibility of regulating the bitterness of Asteraceae species by controlling the pH.

Recent progress in the use of diaziridine-based sweetener derivatives to elucidate the chemoreception mechanism of the sweet taste receptor.

All sweeteners are recognized by the sweet taste receptor (T1R2-T1R3). The elucidation of the chemoreception mechanism of receptor-ligand interactions is an attractive topic for researchers. Molecular biology and computational biology techniques can reveal the proposed mechanisms for this topic. Other approaches, including chemical biology (bioorganic chemistry), have helped to identify mechanisms on the basis of molecular structure. In this mini-review, we have summarized the recent progress in the synthesis of sweetener derivatives, which includes the use of photoaffinity labeling of diazirine-based derivatives to elucidate the chemoreception of sweeteners.

Bitter taste receptor activation by hop-derived bitter components induces gastrointestinal hormone production in enteroendocrine cells.

Matured hop bitter acids (MHBA) are bitter acid oxides derived from hops, widely consumed as food ingredients to add bitterness and flavor in beers. Previous studies have suggested a potential gut-brain mechanism in which MHBA simulates enteroendocrine cells to produce cholecystokinin (CCK), a gastrointestinal hormone which activates autonomic nerves, resulting in body fat reduction and cognitive improvement; however, the MHBA recognition site on enteroendocrine cells has not been fully elucidated. In this study, we report that MHBA is recognized by specific human and mouse bitter taste receptors (human TAS2R1, 8, 10 and mouse Tas2r119, 130, 105) using a heterologous receptor expression system in human embryonic kidney 293T cells. In addition, knockdown of each of these receptors using siRNA transfection partially but significantly suppressed an MHBA-induced calcium response and CCK production in enteroendocrine cells. Furthermore, blocking one of the essential taste signaling components, transient receptor potential cation channel subfamily M member 5, remarkably inhibited the MHBA-induced calcium response and CCK production in enteroendocrine cells. Our results demonstrate that specific bitter taste receptor activation by MHBA drives downstream calcium response and CCK production in enteroendocrine cells. These findings reveal a mechanism by which food ingredients derived from hops in beer activate the gut-brain axis for the first time.