Amino Acid Taste Receptor Research Articles

Impact of feeding diets with enhanced vegetable protein content and presence of umami taste-stimulating additive on gastrointestinal amino acid sensing and feed intake regulation in rainbow trout

The regulation of feed intake in fish is dependent upon different neuroendocrine and metabolic mechanisms including amino acid sensing in the gastrointestinal tract (GIT). However, there is little information regarding the impact of diets on such mechanisms. Therefore, in this study, we fed rainbow trout (Oncorhynchus mykiss) with 3 diets: a diet with high content of fishmeal and low content of soybean protein concentrate (SPC) (HF), a diet with a reduced content of fishmeal and high content of SPC (LF), and the LF diet supplemented with an umami taste-stimulating additive (LFU). Fish were fed ad libitum once a day for 4 weeks, with no significant differences being registered in feed intake among groups. At the end of the feeding trial, we collected samples of different areas of the GIT (stomach, proximal and distal intestine) and hypothalamus at different times: after 48 h of fasting (time 0), and 1 h, 4 h, and 24 h after feeding. We evaluated the activity of pepsin in the stomach and trypsin and chymotrypsin in the proximal intestine, as well as mRNA abundance of transcripts encoding amino acid transporters and taste receptors, intracellular signalling molecules, and hormones. Moreover, we assessed the hypothalamic mRNA abundance of neuropeptides involved in feed intake regulation. Feeding rainbow trout with LF did not result in marked alterations in parameters related to digestive function and amino acid sensing in the rainbow trout GIT, nor in the expression of gastrointestinal hormones (except cck) and hypothalamic neuropeptides. In contrast, supplementation of the LF diet with an umami taste-stimulating additive resulted in a general improvement of digestive or absorptive function (increased protein, dry matter and energy digestibility, and earlier peak in plasma amino acid levels) and activation of gut-brain axis mechanisms involved in feed intake regulation through the transcriptional activation of amino acid transporters, taste receptors, signalling molecules, and hormones. These results demonstrate that the dietary inclusion of umami receptor stimulants has the potential to improve fish physiological responses to the rise in levels of vegetable protein in the diet.

The avian taste system.

Taste or gustation is the sense evolving from the chemo-sensory system present in the oral cavity of avian species, which evolved to evaluate the nutritional value of foods by detecting relevant compounds including amino acids and peptides, carbohydrates, lipids, calcium, salts, and toxic or anti-nutritional compounds. In birds compared to mammals, due to the relatively low retention time of food in the oral cavity, the lack of taste papillae in the tongue, and an extremely limited secretion of saliva, the relevance of the avian taste system has been historically undermined. However, in recent years, novel data has emerged, facilitated partially by the advent of the genomic era, evidencing that the taste system is as crucial to avian species as is to mammals. Despite many similarities, there are also fundamental differences between avian and mammalian taste systems in terms of anatomy, distribution of taste buds, and the nature and molecular structure of taste receptors. Generally, birds have smaller oral cavities and a lower number of taste buds compared to mammals, and their distribution in the oral cavity appears to follow the swallowing pattern of foods. In addition, differences between bird species in the size, structure and distribution of taste buds seem to be associated with diet type and other ecological adaptations. Birds also seem to have a smaller repertoire of bitter taste receptors (T2Rs) and lack some taste receptors such as the T1R2 involved in sweet taste perception. This has opened new areas of research focusing on taste perception mechanisms independent of GPCR taste receptors and the discovery of evolutionary shifts in the molecular function of taste receptors adapting to ecological niches in birds. For example, recent discoveries have shown that the amino acid taste receptor dimer T1R1-T1R3 have mutated to sense simple sugars in almost half of the living bird species, or SGLT1 has been proposed as a part of a T1R2-independent sweet taste sensing in chicken. The aim of this review is to present the scientific data known to date related to the avian taste system across species and its impact on dietary choices including domestic and wild species.

Conditioned Taste Aversion to L-Amino Acid Taste Stimuli and Oral Transcriptional Changes to Type 1 Taste Receptors T1R1 and T1R3 on Chronic Exposure to L-Alanine Solution in Chickens.

Elucidating taste sensing systems in chickens is an important step toward understanding poultry nutrition. Amino acid taste receptors, type 1 taste receptors 1 and 3 (T1R1 and T1R3, respectively), are expressed in chicken taste cells, and chicken T1R1/T1R3 is activated by L-alanine (L-Ala) and L-serine (L-Ser), but not by L-proline (L-Pro). However, it is not clear whether chickens have a gustatory perception of L-amino acids. Here, we found that chickens conditioned to avoid either L-Ala, L-Ser, or L-Pro solutions could successfully learn to avoid the corresponding L-amino acid solution in the conditioned taste aversion (CTA) test. Because CTA is a well-established learning paradigm generated specifically by pairing gustatory perception and gastrointestinal malaise, the present study suggests that chickens can sense L-amino acids by gustatory perception. In addition, we found that the expression of the T1R1 and T1R3 genes was significantly downregulated in response to chronic exposure to L-Ala solution, but not to acute oral stimulation. Taken together, the present study suggests that chickens have a gustatory perception of L-amino acids, and the expression of T1R1/T1R3 mRNAs in the oral cavity can be regulated by L-amino acid intake. Since chickens can detect L-Pro solutions, additional amino acid receptors, other than T1R1/T1R3, may be involved in L-amino acid taste detection in chickens.

The umami receptor T1R1\u2013T1R3 heterodimer is rarely formed in chickens

The characterization of molecular mechanisms underlying the taste-sensing system of chickens will add to our understanding of their feeding behaviors in poultry farming. In the mammalian taste system, the heterodimer of taste receptor type 1 members 1/3 (T1R1/T1R3) functions as an umami (amino acid) taste receptor. Here, we analyzed the expression patterns of T1R1 and T1R3 in the taste cells of chickens, labeled by the molecular markers for chicken taste buds (vimentin and α-gustducin). We observed that α-gustducin was expressed in some of the chicken T1R3-positive taste bud cells but rarely expressed in the T1R1-positive and T2R7-positive taste bud cells. These results raise the possibility that there is another second messenger signaling system in chicken taste sensory cells. We also observed that T1R3 and α-gustducin were expressed mostly in the vimentin-positive taste bud cells, whereas T1R1 and bitter taste receptor (i.e., taste receptor type 2 member 7, T2R7) were expressed largely in the vimentin-negative taste bud cells in chickens. In addition, we observed that T1R1 and T1R3 were co-expressed in about 5% of chickens' taste bud cells, which express T1R1 or T1R3. These results suggest that the heterodimer of T1R1 and T1R3 is rarely formed in chickens’ taste bud cells, and they provide comparative insights into the expressional regulation of taste receptors in the taste bud cells of vertebrates.

Methionine and valine activate the mammalian target of rapamycin complex 1 pathway through heterodimeric amino acid taste receptor (TAS1R1/TAS1R3) and intracellular Ca2+ in bovine mammary epithelial cells

Amino acids play a key role in regulating milk protein synthesis partly through activation of the mammalian target of rapamycin (mTOR) signaling pathway. However, the involvement of extracellular AA sensing receptors in this process is not well understood. In nonruminants, it is well established that the AA taste 1 receptor member 1/3 (TAS1R1/TAS1R3) heterodimer contributes to the sensing of most l-AA. Whether this receptor is functional in bovine mammary cells is unknown. The objective of this study was to determine essential AA signaling through TAS1R1/TAS1R3 and their roles in regulating mTOR signaling pathway and casein mRNA abundance in primary bovine mammary epithelial cells and the Mac-T cell line. The bovine mammary epithelial cells were stimulated with complete Dulbecco's modified Eagle's medium (+EAA), medium without EAA (-EAA), or medium supplemented with only 1 of the 10 essential AA, respectively. The nonessential AA levels were the same across all treatments. Small interference RNA targeting TAS1R1 were designed and transfected into bovine primary mammary epithelial cells (bPMEC). Supplementation of a complete mixture of essential AA or Arg, Val, Leu, His, Phe, Met, and Ile individually led to greater mTOR phosphorylation. Phosphorylation of ribosomal protein S6 kinase β-1 was greater in the presence of Val, Leu, Trp, Met, and Ile. Valine, Leu, Met, and Ile led to greater eIF4E-binding protein 1 phosphorylation. Although +EAA and a few individual AA tested induced increases in intracellular calcium, Met and Val were the most potent. Knockdown of TAS1R1 decreased intracellular calcium in bPMEC cultured with both Val and Met. Phosphorylation of mTOR, ribosomal protein S6 kinase β-1, and eIF4E-binding protein 1 was lower when TAS1R1 was knocked-down in bPMEC supplemented with Val and Met. In addition, small interference RNA silencing of TAS1R1 resulted in lower β-casein (CSN2) abundance. The TAS1R1/TAS1R3 receptor may sense extracellular AA and activate mTOR signaling in bovine mammary cells, likely by elevating intracellular calcium concentration. This mechanism appears to have a role in Met- and Val-induced changes in CSN2 mRNA abundance. Further in vivo studies will have to be performed to assess the relevance of this mechanism in the mammary gland.

Obesity Impairs Oligopeptide/Amino Acid-Induced Ghrelin Release and Smooth Muscle Contractions in the Human Proximal Stomach.

The satiation properties of proteins involve effects on gut peptide release and gastrointestinal motility which may be altered during obesity. This study compares the in vitro response and role of amino acid (AA) taste receptors (TASR) in the effect of AAs and a casein hydrolysate on ghrelin release and smooth muscle (SM) contractions in the proximal gut of lean and obese patients. Basal ghrelin release, measured from mucosal segments, is maximal in the fundus and decreased distally. Obesity selectively impaires the stimulatory effect of a casein hydrolyaste on ghrelin release in the fundus but does not affect its inhibitory effect in the small intestine (SI). The SM contractions induced by a casein hydrolysate and AAs are stronger in strips from the SI than from the fundus but are reduced in the stomach of obese patients. The region-dependent expression of AA-TASRs in the mucosa and SM layer is affected by obesity. Most of the AA-induced responses are reduced by the umami antagonist, lactisole. l-Met-induced responses involve bitter taste receptors. Region-specific targeting of AA taste receptors on both enteroendocrine and SM cells with specific AA-enriched diets might be a useful strategy to combat obesity as well as hypomotility disorders.

Chemosensory signalling pathways involved in sensing of amino acids by the ghrelin cell.

Taste receptors on enteroendocrine cells sense nutrients and transmit signals that control gut hormone release. This study aimed to investigate the amino acid (AA) sensing mechanisms of the ghrelin cell in a gastric ghrelinoma cell line, tissue segments and mice. Peptone and specific classes of amino acids stimulate ghrelin secretion in the ghrelinoma cell line. Sensing of L-Phe occurs via the CaSR, monosodium glutamate via the TAS1R1-TAS1R3 while L-Ala and peptone act via 2 different amino acid taste receptors: CaSR & TAS1R1-TAS1R3 and CaSR & GPRC6A, respectively. The stimulatory effect of peptone on ghrelin release was mimicked ex vivo in gastric but not in jejunal tissue segments, where peptone inhibited ghrelin release. The latter effect could not be blocked by receptor antagonists for CCK, GLP-1 or somatostatin. In vivo, plasma ghrelin levels were reduced both upon intragastric (peptone or L-Phe) or intravenous (L-Phe) administration, indicating that AA- sensing is not polarized and is due to inhibition of ghrelin release from the stomach or duodenum respectively. In conclusion, functional AA taste receptors regulate AA-induced ghrelin release in vitro. The effects differ between stomach and jejunum but these local nutrient sensing mechanisms are overruled in vivo by indirect mechanisms inhibiting ghrelin release.

No Taste for Health: How Tastes are Being Manipulated to Favour Foods that are not Conducive to Health and Wellbeing

No Taste for Health: How Tastes are Being Manipulated to Favour Foods that are not Conducive to Health and Wellbeing

Genetic loss or pharmacological blockade of testes-expressed taste genes causes male sterility

TAS1R taste receptors and their associated heterotrimeric G protein gustducin are involved in sugar and amino acid sensing in taste cells and in the gastrointestinal tract. They are also strongly expressed in testis and sperm, but their functions in these tissues were previously unknown. Using mouse models, we show that the genetic absence of both TAS1R3, a component of sweet and amino acid taste receptors, and the gustducin α-subunit GNAT3 leads to male-specific sterility. To gain further insight into this effect, we generated a mouse model that expressed a humanized form of TAS1R3 susceptible to inhibition by the antilipid medication clofibrate. Sperm formation in animals without functional TAS1R3 and GNAT3 is compromised, with malformed and immotile sperm. Furthermore, clofibrate inhibition of humanized TAS1R3 in the genetic background of Tas1r3(-/-), Gnat3(-/-) doubly null mice led to inducible male sterility. These results indicate a crucial role for these extraoral "taste" molecules in sperm development and maturation. We previously reported that blocking of human TAS1R3, but not mouse TAS1R3, can be achieved by common medications or chemicals in the environment. We hypothesize that even low levels of these compounds can lower sperm count and negatively affect human male fertility, which common mouse toxicology assays would not reveal. Conversely, we speculate that TAS1R3 and GNAT3 activators may help infertile men, particularly those that are affected by some of the mentioned inhibitors and/or are diagnosed with idiopathic infertility involving signaling pathway of these receptors.

Sensing of amino acids by the gut-expressed taste receptor T1R1-T1R3 stimulates CCK secretion

CCK is secreted by endocrine cells of the proximal intestine in response to dietary components, including amino acids. CCK plays a variety of roles in digestive processes, including inhibition of food intake, consistent with a role in satiety. In the lingual epithelium, the sensing of a broad spectrum of L-amino acids is accomplished by the heteromeric amino acid (umami) taste receptor (T1R1-T1R3). T1R1 and T1R3 subunits are also expressed in the intestine. A defining characteristic of umami sensing by T1R1-T1R3 is its potentiation by IMP or GMP. Furthermore, T1R1-T1R3 is not activated by Trp. We show here that, in response to L-amino acids (Phe, Leu, Glu, and Trp), but not D-amino acids, STC-1 enteroendocrine cells and mouse proximal small intestinal tissue explants secrete CCK and that IMP enhances Phe-, Leu-, and Glu-induced, but not Trp-induced, CCK secretion. Furthermore, small interfering RNA inhibition of T1R1 expression in STC-1 cells results in significant diminution of Phe-, Leu-, and Glu-stimulated, but not Trp-stimulated, CCK release. In STC-1 cells and mouse intestine, gurmarin inhibits Phe-, Leu-, and Glu-induced, but not Trp-stimulated, CCK secretion. In contrast, the Ca(2+)-sensing receptor antagonist NPS2143 inhibits Phe-stimulated CCK release partially and Trp-induced CCK secretion totally in mouse intestine. However, NPS2143 has no effect on Leu- or Glu-induced CCK secretion. Collectively, our data demonstrate that functional characteristics and cellular location of the gut-expressed T1R1-T1R3 support its role as a luminal sensor for Phe-, Leu-, and Glu-induced CCK secretion.

Amino acid taste receptor regulates insulin secretion in pancreatic β-cell line MIN6 cells

Amino acids such as L-glutamate and L-arginine are potent stimuli for insulin secretion from pancreatic β-cells. However, the precise molecular mechanisms of amino acid-induced insulin secretion have only partly understood. In this study, we show here that mouse pancreatic β-cell line MIN6 cells expressed amino acid taste receptor (heterodimer of type 1 taste G protein-coupled receptor member 1 and member 3; Tas1R1 and Tas1R3, respectively). We found that administration of L-glutamate or L-arginine to MIN6 cells caused the increase in free intracellular concentrations of both inositol-1,4,5-triphosphate (IP(3)) and Ca(2+) , and umami substance inocinate enhanced the effects of l-glutamate. Effects of amino acids on intracellular IP(3) and Ca(2+) concentration were diminished by application of lactisole, a Tas1R3 receptor antagonist. Furthermore, we investigated the effect of amino acids on the insulin release from MIN6 cells by both ELISA and total internal reflection fluorescence microscopy. Application of L-glutamate or L-arginine significantly increased the release of insulin, whereas inhibited by the application of lactisole. Based on these findings, we propose that heterodimer of Tas1R1 and Tas1R3 is the fundamental receptor for the sensing amino acids and regulates the amino acid-induced insulin secretion in pancreatic β-cells.

Errata

The paper by Mace et al. contains two errors on page 389. On line 43 ‘sucralose > acesulfame potassium ∼ saccharin’ was written incorrectly and should read ‘sucralose ∼ acesulfame potassium > saccharin’. On line 28 ‘cholecystokinin’ was incorrectly written as ‘cholecystokinic’. The sentence beginning on line 30 of the Discussion page 389 should read: ‘By contrast, the classic amino acid taste receptor components, T1R1 and transducin, and also PLC β2, behave reciprocally.’ The publishers wish to apologize for these errors.

The G‐protein coupling properties of the human sweet and amino acid taste receptors

The human T1R taste receptors are family C G-protein-coupled receptors (GPCRs) that act as heterodimers to mediate sweet (hT1R2 + hT1R3) and umami (hT1R1 + hT1R3) taste modalities. Each T1R has a large extracellular ligand-binding domain linked to a seven transmembrane-spanning core domain (7TMD). We demonstrate that the 7TMDs of hT1R1 and hT1R2 display robust ligand-independent constitutive activity, efficiently catalyzing the exchange of GDP for GTP on Galpha subunits. In contrast, relative to the 7TMDs of hT1R1 and hT1R2, the 7TMD of hT1R3 couples poorly to G-proteins, suggesting that in vivo signaling may proceed primarily through hT1R1 and hT1R2. In addition, we provide direct evidence that the hT1Rs selectively signal through Galpha(i/o) pathways, coupling to multiple Galpha(i/o) subunits as well as the taste cell specific Gbeta(1)gamma(13) dimer.

In Brief

A recent article in Nature describes the identification of a mammalian amino acid taste receptor, or rather combinations of two different G-protein-coupled receptors that function as taste receptors. The combination of T1R2 plus T1R3 receptors was activated by several d-amino acids that taste sweet to humans. By contrast, the T1R1–T1R3 combination only responded to l-amino acids. Interestingly, a single amino acid mutation in the sequence of T1R3 only affected its signalling when it was combined with T1R2, but not with T1R1. Finally, the authors showed that the T1R1 plus T1R3 combination might be involved in the unique taste perception induced by sodium monoglutamate. [Nelson, G. et al. (2002) Nature 416, 199–201] AB

A Savory Story

Specific G protein-coupled receptors (GPCRs) are activated during the sensation of light, smell, and taste. Nelson et al. used a transfected cell assay to show that the heterodimeric GPCR, comprised of TL1R1 and TL1R3, is a taste receptor activated by L-amino acids, but not sweet D-amino acids. Furthermore, this TL1R1-TL1R3 dimer also exhibited three properties consistent with its being involved in the detection of the taste of umami. Namely, monosodium glutamate (MSG) and the glutamate receptor agonist L-AP4 activate TL1R1-TL1R3, and its responsiveness is potentiated by purine nucleotides. Umami is the taste associated with foods containing MSG, such as Chinese foods. The TL1R3 GPCR is also part of a heterodimeric GPCR that responds to sweet substances. Thus, changing one subunit of the GPCR dimer confers completely different ligand specificity. In mice, the Sac gene encodes the TL1R3 receptor and "nontaster" Sac mice have a mutation in TL1R3 and do not respond to sweet taste, however, they respond normally to amino acids. One proposal was that the mutation in nontaster TL1R3 prevented the assembly of the sweet receptor dimer without affecting the amino acid receptor dimer. However, Nelson et al. showed that nontaster TL1R3 coprecipitated with TL1R1 and TL1R2, yet sweet substances did not activate the nontaster-containing receptor dimer. Mice and humans respond differently to various taste substances, and these differences can be explained by differences in the primary sequences of the TL1 receptors, because substituting the TL1 receptors from mouse or human in the transfected cell assay produced altered sensitivities to different taste stimuli. G. Nelson, J. Chandrashekar, M. A. Hoon, L. Feng, G. Zhao, N. J. P. Ryba, C. S. Zuker, An amino-acid taste receptor. Nature 416 , 199-202 (2002). [Online Journal]

An amino-acid taste receptor.

The sense of taste provides animals with valuable information about the nature and quality of food. Mammals can recognize and respond to a diverse repertoire of chemical entities, including sugars, salts, acids and a wide range of toxic substances. Several amino acids taste sweet or delicious (umami) to humans, and are attractive to rodents and other animals. This is noteworthy because L-amino acids function as the building blocks of proteins, as biosynthetic precursors of many biologically relevant small molecules, and as metabolic fuel. Thus, having a taste pathway dedicated to their detection probably had significant evolutionary implications. Here we identify and characterize a mammalian amino-acid taste receptor. This receptor, T1R1+3, is a heteromer of the taste-specific T1R1 and T1R3 G-protein-coupled receptors. We demonstrate that T1R1 and T1R3 combine to function as a broadly tuned L-amino-acid sensor responding to most of the 20 standard amino acids, but not to their D-enantiomers or other compounds. We also show that sequence differences in T1R receptors within and between species (human and mouse) can significantly influence the selectivity and specificity of taste responses.

Selective interactions of lectins with amino acid taste receptor sites in the channel catfish

Selective interactions of lectins with amino acid taste receptor sites in the channel catfish

Structure/activity relationships in the L-alanine taste receptor system of the channel catfish, Ictalurus punctatus

In order to understand the molecular determinants of amino acid taste receptor binding and activation, structure/activity studies were performed using analogs of L-alanine in a competitive ligand binding assay and a taste neurophysiological preparation. The presence of both the amine and carboxylic acid in a charged and unhindered form is a primary requisite for a ligand to both bind and activate L-alanine receptors. Although a number of carboxylic acid derivatives are moderately good stimuli, their neural activity derives from action at receptors different from L-alanine receptors. Of the molecular parameters examined, chirality and molecular volume of the side chain are the most important factors in determining the binding and stimulatory efficacy of L-amino acid taste stimuli. Electronegativity of the side chain did not correlate with receptor site binding. Heterologous ligand-induced enhancement of the binding of L-[ 3H]alanine by a purified taste membrane preparation is described. Neurophysiological experiments support the hypothesis that this phenomenon may be a basis of peripheral sensory interactions.

A stimulus-activated conductance in isolated taste epithelial membranes

Membrane vesicles isolated from the cutaneous taste epithelium of the catfish were incorporated into phospholipid bilayers on the tips of patch pipettes. Voltage-dependent conductances were observed in approximately 50% of the bilayers and single-channel currents having conductances from 8 to greater than 250 pS were recorded. In 40% of the bilayers displaying no voltage-dependent conductances, micromolar concentrations of L-arginine, a potent stimulus for one class of catfish amino acid taste receptors, activated a nonselective cation conductance. The L-arginine-gated conductance was concentration-dependent, showing half-maximal activation in response to approximately 15 microM L-arginine. L-Arginine-activated channels had unitary conductances of 40-50 pS and reversed between -6 and +18 mV with pseudointracellular solution in the pipette and Ringer in the bath. L-Alanine, a potent stimulus for the other major class of catfish amino acid taste receptors, did not alter bilayer conductance. D-Arginine, which is a relatively ineffective taste stimulus for catfish but a good cross-adapter of the L-arginine-induced neural response, had no effect on bilayer conductance at concentrations below 200 microM. However, increasing concentrations of D-arginine from 1 to 100 microM progressively suppressed the L-arginine-activated conductance, suggesting that D-arginine competed for the L-arginine receptor, but did not activate the associated cation channel. This interpretation is consonant with recent biochemical binding studies in this system. These results suggest that L-arginine taste receptor proteins in the catfish are part of or closely coupled to cation-selective channels which are opened by L-arginine binding.

Expression of catfish amino acid taste receptors inXenopus oocytes

We demonstrate that poly (A+)RNA isolated from catfish barbels directs the expression of functional amino acid taste receptors in the Xenopus oocyte. The activity of these receptors is monitored in ovo by the two electrode voltage clamp technique. Specific conductance changes recorded in response to amino acid stimulation are analogous to those recorded electrophysiologically from intact catfish barbels. These responses exhibit specificity, reproducibility, rapid onset and termination, and desensitization to repetitive stimulation. A functional assay system that encompasses the full complement of transduction events from the ligand-receptor interaction to subsequent conductance changes is necessary to identify molecular components responsible for these events. Our results demonstrate that the Xenopus oocyte can be used to characterize and identify clones coding for amino acid taste receptors analogous to its use in studying receptor molecules for other neuroactive compounds.