Searching for the bitter truth: Insights into receptors for taste

Abstract

Mueller KL, Hoon MA, Erlenbach I, Chandrashekar J, Zuker CS, Ryba NJ (University of California at San Diego, San Diego, California; National Institute of Dental and Craniofacial Research, Bethesda, Maryland) The receptors and coding logic for bitter taste. Nature 2005;434:225–229. The ability to detect bitter taste is proposed to protect organisms from ingesting dangerous substances. Receptors on specialized cells on the tongue have been characterized which distinguish the different tastes. T2Rs are G protein–coupled receptors that participate in bitter taste perception. In this study, Mueller and colleagues demonstrate the sufficiency and necessity of T2R receptors for bitter taste sensation, that interspecies differences in T2Rs produce differential selectivity of bitter taste responsiveness, and that the cell type in which the T2R is expressed determines the behavioral response to a given stimulus. The investigators first assessed if T2Rs are sufficient for bitter taste perception. Transgenic mice were generated that express human receptors under the control of a mouse T2R promoter for β-glucopyranosides (hT2R16) and phenylthiocarbamide (PTC)(hT2R38), substances which are not perceived as bitter in mice. T2R-hT2R16 transgenic mice were able to detect phenyl-β-D-glucopyranoside at concentrations similar to those detected by humans in two-bottle intake preference assays and licking responses on a gustometer. Similarly, T2R-hT2R38 transgenic animals perceived PTC in comparable assays. The investigators then used homologous recombination to generate mice lacking mT2R5, the Cyx receptor responsive to the bitter compound cycloheximide, to test if T2Rs are necessary for bitter taste detection. T2R5−/− mice exhibited reduced responses to cycloheximide and were no longer behaviorally averse to the substance. Conversely, T2R5 knockout mice exhibited normal responses to sweet, sour, umami, salty, and non-cycloheximide bitter substances. Next, these investigators determined if taste receptor cells are responsive to single or multiple bitter taste modalities. PLCβ2 is an effector molecule involved in signaling after activation of a broad range of taste receptors. PLCβ2 deletion abolishes taste of sweet, umami, and bitter substances. PLCβ2 was reintroduced in T2R cells in PLCβ2−/− knockout mice under the control of one of the T2R-promoters on 3 separate chromosomal locations (T2R5, T2R19, T2R32) and the engineered animals were exposed to a number of bitter substances. PLCβ2 rescued mice with only one of the T2R-promoters responded normally to all of the bitter compounds, indicating that individual T2R cells perform as broadly tuned bitter receptors. Finally, mice were generated that expressed an inducible RASSL receptor on bitter taste cells. RASSL is a κ-opioid receptor responsive to the tasteless opioid agonist spiradoline. In prior studies, sweet cells expressing RASSL responded to spiradoline, showing that activating the cells rather than the sweet receptor itself produced the sensation of sweetness. In the present study, mice expressing RASSL in bitter taste cells showed aversion to spiradoline. Furthermore, mice expressing the bitter receptor for β-glucopyranosides on sweet cells exhibited attraction rather than aversion to the bitter compound, indicating that taste sensation is a property of the cells activated rather than the substance being tasted. The authors concluded that bitter taste coding is determined by dedicated cell types that act as sensors to a broad range of bitter tastants and elicit behavioral aversion. The gastrointestinal tract begins in the mouth, where contact of the consumed bolus is made with taste buds on the tongue, yet most gastroenterologists have a limited understanding of the process of taste. Perceptual discrimination of different tastes plays a critical role in deciding which substances are ingested for subsequent digestion and absorption and which are rejected. Pleasing compounds such as sweets are avidly consumed, whereas unpleasant bitter substances elicit aversion and are not ingested. The fungiform papillae on the anterior tongue and the circumvallate and foliate papillae on central and lateral regions contain the taste buds (Int J Dev Biol 2004;48:157–161). Taste molecules bind to microvillus tips and to small, membrane-bounded cytoplasmic blebs which are shed, providing a possible means of elimination of tastant-receptor complexes (Scan Micros 1987;1:351–357). The keratinized epithelium overlying the taste buds is permeable, facilitating the process of taste (Micros Res Tech 1993;26:94–105). In mice, 38% of taste cells respond to multiple taste qualities, whereas other cells activate in response to tastants of only a single class (J Physiol 2002;544:501–509). Taste cells synapse with afferent neurons that project to the central nervous system, and receive efferent projections (J Comp Neurol 2000;417:315–324). The sense of taste is mediated by cranial nerves VII (facial), IX (glossopharyngeal), and X (vagus), which project to the oral somatosensory and gustatory cortex (Brain Behav Evol 2004;64:198–206, J Neurosci 2001;21:4478–4489). Cortical regions respond to an array of stimulus properties including taste, temperature, and texture of the ingested substance (Physiol Behav 2005;85:45–56). Roughly 40% of gustatory cortex neurons respond to specific tastes, although many neurons respond to several different stimulants (J Neurosci 2001;21:4478–4489). Mammals sense tastes in 4 broad categories: bitter, sweet, salty and sour, and umami (taste of glutamate) (Nature 2000;404:601–604). Sweet substances activate chorda tympani nerves to greater degrees than glossopharyngeal nerves, whereas some bitter compounds preferentially stimulate the glossopharyngeals (BMC Neurosci 2003;4:5). Thirty-eight to 50% of S fibers in both nerves respond to sweets, whereas 40%–46% of Q fibers predominantly respond to bitter substances (J Neurophysiol 2002;88:579–594). Furthermore, first order neurons in the glossopharyngeal and chorda tympani nerves can discriminate the different bitter compounds quinine, caffeine, nicotine, and phenylthiocarbamide (PTC) (Brain Res 1997;756:22–34). Consistent with this is the observation that taste cells respond to bitter substances through multiple transduction pathways (J Neurophysiol 1997;78:734–745). There is significant interindividual variability in the sense of taste. Taster and nontaster phenotypes for the ability to detect PTC are described which are present in characteristic proportions in different geographic regions (Am J Human Gen 2004;74:637–646). Specific receptors that are members of the G protein–coupled receptor superfamily with 7 transmembrane domains have been characterized for bitter, sweet, and umami tastes by genetic linkage and positional cloning methods (J Dent Res 2004;83:448–453, Cell 1999;96:541–551, Nature 2000;404:601–604). A family of 40–80 human and rodent T2Rs expressed on selected taste receptor cells has been identified which detects chemically dissimilar bitter substances (Cell 2000;100:693–702, Nutr Rev 2001;59:163–169). Individual taste receptor cells may express a number of T2Rs, but many bitter-sensing cells are activated by a small number of bitter compounds, indicating that different populations of taste cells respond to distinct tastants (Science 2001;291:1557–1560). Several T2Rs have been described, including mouse receptors for cycloheximide (mT2R5) and 6-n-propyl-2-thiouracil (mT2R8) and human receptors for denatonium (hT2R4, hT2R44), β-glucopyranosides (hT2R16), 6-nitrosaccharin (hT2R61, hT2R44), and PTC (hT2R38) (Cell 2000;100:703–711, Nat Gen 2002;32:397–401, Chem Sense 2004;29:583–593, Curr Biol 2005;15:322–327). Amino acid substitutions in mT2R5 render mice less responsive to cycloheximide, whereas expression of the same receptor in insects elicits responsiveness of signaling systems for bitter taste sensation. In humans, 5 haplotypes of the T2R gene that detects PTC account for the variable sensitivity to that substance in the population (Science 2003;299:1221–1225, Clin Genet 2005;67:275–280). Bitter taste receptors for the sweet substances saccharin and acesulfame K are believed to participate in the bitter aftertaste sensed after ingesting these compounds (J Neurosci 2004;24:10260–10265). Our understanding of detection of nonbitter tastants also has improved. In mice, tasters and nontasters of selected sweet substances have different amino acid sequences in the sweet receptor T1R3 (Nat Neurosci 2001;4:492–498). Two receptors, T1R1 and T1R3, combine as a broadly tuned sensor that activates in response to nearly 20 amino acids (Nature 2002;416:199–202). In humans, one heterodimer (T1R2/T1R3) responds to sweets while another (T1R1/T1R3) detects umami tastes (Proc Natl Acad Sci U S A 2002;99:4692–4696). Elimination of one T1R subunit abolishes detection of one of the taste modalities (Cell 2003;115:255–266). Lingual exposure to acid produces a sour taste. Proposed mechanisms of acid sensation include altered function of potassium, calcium, sodium, and chloride channels in taste cells (Resp Physiol 2001;129:231–245, Brain Res 2001;923:58–70, J Physiol 2003;547:475–483). Upon screening of a rat papilla cDNA library, two amiloride-sensitive, proton-activated cation channels were found in taste cells, including acid sensing ion channel 2a (ASIC2a) and ASIC2b (Anat Sci Int 2003;78:205–210). These channels exhibit greater activation on exposure to acetic acid than hydrochloric acid, corresponding to the observation that acetic acid is perceived as the more sour substance. Signaling in response to T2R and T1R activation is mediated by the G-protein gustducin (Cell 2000;100:703–711). Mice with knockouts to the alpha subunit of gustducin show reduced neural and behavioral responses to many bitter and sweet substances (Chem Sense 2002;27:719–727, Chem Sense 2005;30:299–316). Gustducin is not expressed in cells responsive to acid stimulation or sour tastants (Nature 2001;413:631–615). Furthermore, knockouts of TRPM5, an ion channel involved in taste, and PLCβ2, a phospholipase that is responsible for signaling after activation of an extended array of taste molecules, abolish responses to bitter, sweet, and umami tastants (Cell 2003;112:293–301). The present study provides important insight into the link between T2R activation after bitter tastant exposure and the physiologic response of the whole organism. Using mice engineered to express human T2Rs and mice with knockouts to specific T2Rs, the investigators showed that T2Rs are both sufficient and required for detection of and behavioral responses to bitter substances. Next, reintroduction of the signaling apparatus under the control of single T2R promoter to individual taste cells in mice with selective knockouts of a specific effector molecule restored reactivity to several bitter tastants, indicating that individual T2R cells are broadly responsive to bitter stimuli. Finally, expression of receptors for nonbitter compounds on bitter taste cells led to aversion to the nonbitter substances, whereas expression of bitter receptors on sweet taste cells produced attraction, indicating that the behavioral response to a specific tastant results from activation of a given taste cell type rather than the receptor itself. The findings of this investigation advance our understanding of the receptor-mediated processes of taste. A sizeable research commitment has been made in academic centers and the pharmaceutical industry to characterize selected receptor-effector pathways in an effort to develop novel receptor agonists and antagonists for use in the clinical arena. Future study into the mechanisms underlying the sense of taste will provide insight into how individuals choose what to ingest and what to reject. Better understanding of these pathways may allow for engineering of ingested nutritive supplements or pharmaceuticals which are designed to activate only selected taste receptor pathways to optimize their tolerability.