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. 2010 Feb 19;107(10):4746–4751. doi: 10.1073/pnas.0911670107

Fig. 1.

Fig. 1.

Identification of a PAM for the human sweet receptor. (A) Chemical structure of SE-1. (B) SE-1 enhances sucralose-evoked increases in [Ca2+]int. Application of 75 μM SE-1 or D-PBS (A) did not evoke an increase in fluorescence. Subsequent application of 20 μM sucralose (final concentration) (B) produced a much greater increase in [Ca2+]int and fluorescence, on cells preexposed to SE-1 than on cells preexposed to D-PBS. (C) Dose–response effect of SE-1 on its own (•) and on the sucralose-evoked increase in [Ca2+]int (○). Cells were stimulated with SE-1 at concentrations up to 75 μM and the change in [Ca2+]int was monitored over the first 90 sec postaddition. At 220 sec, a suboptimal concentration of sucralose (10 μM final concentration) was then added onto the cells and the subsequent change in [Ca2+]int was monitored. SE-1 dose-dependently enhanced the sucralose effect with an EC50 of 3.0 ± 1.1 μM (average ± SD; n = 6). (D) SE-1 increases the sucralose potency in the assay. Sucralose dose–response effects were monitored after the cells had been preexposed to various concentrations of SE-1 of 1.5, 4.5, 15, and 45 μM, producing the depicted final enhancer concentrations after sweetener addition. In this representative experiment, the sucralose EC50 value was decreased from 60 μM to 39, 24, 9.4, and 3.3 μM at 1, 3, 10, and 30 μM of SE-1, respectively. (E) SE-1 is highly selective for sucralose. Cells were preexposed to either D-PBS or 75 μM SE-1 and 220 sec later, suboptimal concentrations of depicted sweeteners were added into the wells (bringing SE-1 concentration down to 50 μM final) and cell responses were monitored on the FLIPR system. Final concentrations of sweeteners used were as follows: 15 μM sucralose, 150 nM neotame, 25 mM sucrose, 5 μM alitame, 100 μM aspartame, 30 μM saccharin, 10 μM dulcin, 15 μM stevioside, 300 μM cyclamate, 10 μM mogroside, 1 μM perillartine, 20 μM glyccyrhizic acid, 13 μM NHDC, and 4 μM thaumatin. SE-1 significantly enhanced the effect of sucralose (***, P < 0.0001, Student's t test), neotame (**, P < 0.001), and sucrose (*, P < 0.05). Data points in C–E correspond to an average ± SD of a triplicate determination.