2009 Abstracts - Association for Chemoreception Sciences

#P183 Poster session IV: Chemosensory transductionand perireceptor eventsGurmarin inhibits the Sweet Receptor by Binding to theVenus Fly Trap Module of T1R3Emeline L. Maillet, Laura Pelletier, Timothy J. Cardozo,Jeniffer Quijada, Prisca Silie, Baohua Zhao, Yuzo Ninomiya,Marianna Max, Robert F. MargolskeeMount Sinai School of Medicine, Department of Neuroscience.Box1065 New York, NY, USAGurmarin is a polypeptide of 35 amino-acids that suppressesbehavioral and gustatory neural responses of rodents to sweetcompounds without affecting responses to salty, sour, or bittersubstances. Gurmarin has no detectable effect in humanpsychophysical studies. Here, we show that gurmarin acts onmouse T1R3 to antagonize the heterologously expressed mousesweet receptor’s response to a panel of sweeteners. Co-expressingthe non-taster allele of mT1R3 with human T1R2 yields afunctional sweet receptor that is sensitive to gurmarin, indicatingthat these allelic variations in mT1R3 does not affect gurmarinbinding. Studies with human-mouse chimeras of T1R3 indicatedthat the first 150 amino acids (aa) of T1R3 must be from mouse tomaintain sensitivity to gurmarin. Additional chimeras narrowedthe region of importance to aa 40-80 of mT1R3. Based on thecrystal structure of mGluR1, we created a homology model of theVenus Fly Trap Module of mT1R3. According to this model a41-66 aa loop (loop1) is present within the upper lobe1 of thereceptor’s cleft. In our model, gurmarin docks to the receptorwithin the open cleft of the VFTM, directly in contact with theupper lobe. The difference in the 3D shape of the correspondingloop1 of human T1R3 suggests that steric hindrance preventsgurmarin from binding to human T1R3’s VFTM cleft. Wehypothesize that gurmarin may inhibit activation of the sweetreceptor by preventing proper adoption of T1R3 VFTM closestate. In addition, analysis of the interactions in the docked modelbetween gurmarin and the receptor accord with previous workidentifying key aromatic residues necessary for gurmarinfunction. Finally, point mutants altered at residues of mouseT1R3 predicted to interact with gurmarin displayed reducedsensitivity to gurmarin in in-vitro assays.#P184 Poster session IV: Chemosensory transductionand perireceptor eventsEffect of inosine monophosphate (IMP) on taste perceptionof methionine and valine by miceYuko Murata 1 , Alexander A. Bachmanov 2 , Gary K. Beauchamp 21National Research Institute of Fisheries Science Yokohama, Japan,2Monell Chemical Senses Center Philadelphia, PA, USAIn vitro heterologous expression studies showed that most L-amino acids, including L-methionine (Met) and L-Valine (Val),activate the mouse T1R1+T1R3 receptor when they are mixedwith IMP. However, Met and Val differ in their ability to activatethe mouse T1R1+T1R3 receptor without IMP. While Metstrongly activates this receptor, Val evokes only negligibleactivation (Nelson et. al., 2002). The goal of our study was toexamine whether addition of IMP changes taste quality perceptionof Met and Val. We have addressed this question using aconditioned taste aversion (CTA) technique. Separate groups ofC57BL/6J mice were exposed to one of four conditioned stimuli(50 mM Met, 50 mM Met + 2.5 mM IMP, 50 mM Val or 50 mMVal + 2.5 mM IMP) or to water (control) and injected with LiClto form CTA. Conditioned mice were presented with five basictaste solutions, Met and Val, and their lick responses wererecorded. An aversion to Met generalized to quinine, while anaversion to Met+IMP generalized to Met, 150 mM sucrose, amixture of 50 mM MSG and 30 M amiloride (Ami; added toblock sodium taste) with or without 2.5 mM IMP (i.e.,MSG+IMP+Ami and MSG+Ami), but not quinine. An aversionto Val generalized to quinine, while an aversion to Val+IMPgeneralized to MSG+IMP+Ami and MSG+Ami, but not quinine.This suggests that addition of IMP changes the taste quality ofMet and Val in vivo, which is consistent with results of in vitroexperiments. Supported by Fisheries Research Agency(Yokohama, Japan) research grant (YM) and NIH grant DC00882 (GKB and AAB).#P185 Poster session IV: Chemosensory transductionand perireceptor eventsMitigation of irradiation effects on taste epithelium in theProtein Kinase C delta null mouseH.M. Nguyen 1 , M.E. Reyland 2 , L.A. Barlow 11Dept. of Cell & Developmental Biology, and The RockyMountain Taste and Smell Center, School of Medicine, Universityof Colorado Denver Aurora, CO, USA, 2 Dept. of CraniofacialBiology, School of Dental Medicine, University of ColoradoDenver Aurora, CO, USARadiotherapy for head and neck cancer can result in taste loss;yet how radiation influences taste is unknown. One idea is thatdisruption of taste cell renewal may be causal. Taste cell turnoverspans 10-14 days (Beidler and Smallman, 1965) driven byproduction of transit amplifying (TA) cells in or near taste budsfrom as-of-yet unidentified stem cells. We propose that taste lossafter irradiation is due to apoptosis of replenishing TA cells, andconsequent attrition of mature taste cells. In support of this, wefind that proliferating TA cells are absent at 3 days postirradiation (dpi) including cells in S (BrdU-IR) and M (phosphohistone3(pH3)-IR) phases, in fungiform (ffp) and circumvallate(cvp) papillae. Thus, one strategy for averting taste loss may be toreduce epithelial cell death, with the prediction that cell divisionwould not be interrupted. Protein kinase C delta (PKCd) is amultifunctional kinase, which positively regulates apoptosis.To test if radiation effects on taste epithelium are mitigated inPKCd-/- mice, the heads of wild type (WT) and PKCd-/- adultswere irradiated with a single 8Gy dose and lingual epitheliaexamined for BrdU- and pH3-IR at progressive dpi. As in ourinitial results, in ffp and cvp, BrdU-IR cells are absent at 3 dpi,increase at 5 dpi, then decrease after 9 dpi. Cells in M phase(pH3-IR) are missing at 3 and 5 dpi, reappear by 7 dpi, thendecrease after 9 dpi. By contrast, the proliferative profile ofirradiated PKCd-/- mice does not drop; BrdU- and pH3-IR cellsare present at 3 and 5 dpi and dividing cell number is dramaticallyhigher from 7-11 dpi compared with WT mice. Our data suggestthat PKCd is required for apoptotic cell death and/or repressesproliferation in irradiated taste epithelium.84 | AChemS Abstracts 2009