The noncaloric sweeteners (NCSs) cyclamate (Cycl) and acesulfame K (AceK) are widely added to foods and beverages. Little is known about their impact on gastric acid secretion (GAS), which is stimulated by dietary protein and bitter-tasting compounds. Since Cycl and AceK have a bitter off taste in addition to their sweet taste, we hypothesized they modulate mechanisms of GAS in human gastric parietal cells (HGT-1). HGT-1 cells were exposed to sweet tastants (50 mM of glucose, d-threonine, Cycl, or AceK) and analyzed for their intracellular pH index (IPX), as an indicator of proton secretion by means of a pH-sensitive dye, and for mRNA levels of GAS-associated genes by RT-qPCR. Since the NCSs act via the sweet taste-sensing receptor T1R2/T1R3, mRNA expression of the corresponding genes was analyzed in addition to immunocytochemical localization of the T1R2 and T1R3 receptor proteins. Exposure of HGT-1 cells to AceK or d-threonine increased the IPX to 0.60 ± 0.05 and 0.80 ± 0.04 ( P ≤ 0.05), respectively, thereby indicating a reduced secretion of protons, whereas Cycl demonstrated the opposite effect with IPX values of -0.69 ± 0.08 ( P ≤ 0.05) compared to controls (IPX = 0). Cotreatment with the T1R3-inhibitor lactisole as well as a TAS1R3 siRNA knock-down approach reduced the impact of Cycl, AceK, and d-thr on proton release ( P ≤ 0.05), whereas cotreatment with 10 mM glucose enhanced the NCS-induced effect ( P ≤ 0.05). Overall, we demonstrated Cycl and AceK as modulators of proton secretion in HGT-1 cells and identified T1R3 as a key element in this response.

The careful evaluation of food is important for survival throughout the animal kingdom, and specialized chemoreceptors have evolved to recognize nutrients, minerals, acids, and many toxins. Vertebrate bitter taste, mediated by the taste receptor type 2 (T2R) family, warns against potentially toxic compounds. During evolution T2R receptors appear first in bony fish, but the functional properties of bony fish T2R receptors are mostly unknown. We performed a phylogenetic analysis showing the “living fossil” coelacanth (Latimeria chalumnae) and zebrafish (Danio rerio) to possess T2R repertoires typical for early-diverged species in the lobe-finned and the ray-finned clade, respectively. Receptors from these two species were selected for heterologous expression assays using a diverse panel of bitter substances. Remarkably, the ligand profile of the most basal coelacanth receptor, T2R01, is identical to that of its ortholog in zebrafish, consistent with functional conservation across >400 Myr of separate evolution. The second coelacanth receptor deorphaned, T2R02, is activated by steroid hormones and bile acids, evolutionary old molecules that are potentially endogenously synthesized agonists for extraoral T2Rs. For zebrafish, we report the presence of both specialized and promiscuous T2R receptors. Moreover, we identified an antagonist for one of the zebrafish receptors suggesting that bitter antagonism contributed to shape this receptor family throughout evolution.

The sense of taste plays an important role in the evaluation of the nutrient composition of consumed food. Bitter taste in particular is believed to serve a warning function against the ingestion of poisonous substances. In the past years enormous progress was made in the characterization of bitter taste receptors, including their gene expression patterns, pharmacological features and presumed physiological roles in gustatory as well as in non-gustatory tissues. However, due to a lack in TAS2R-specifc antibodies the localization of receptor proteins within gustatory tissues has never been analyzed. In the present study we have screened a panel of commercially available antisera raised against human bitter taste receptors by immunocytochemical experiments. One of these antisera was found to be highly specific for the human bitter taste receptor TAS2R38. We further demonstrate that this antibody is able to detect heterologously expressed TAS2R38 protein on Western blots. The antiserum is, however, not able to interfere significantly with TAS2R38 function in cell based calcium imaging analyses. Most importantly, we were able to demonstrate the presence of TAS2R38 protein in human gustatory papillae. Using double immunofluorescence we show that TAS2R38-positive cells form a subpopulation of PLCbeta2 expressing cells. On a subcellular level the localization of this bitter taste receptor is neither restricted to the cell surface nor particularly enriched at the level of the microvilli protruding into the pore region of the taste buds, but rather evenly distributed over the entire cell body.

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