The oral lipid sensor GPR120 is not indispensable for the orosensory detection of dietary lipids in mice

Abstract

Implication of the long-chain fatty acid (LCFA) receptor GPR120, also termed free fatty acid receptor 4, in the taste-guided preference for lipids is a matter of debate. To further unravel the role of GPR120 in the "taste of fat", the present study was conducted on GPR120-null mice and their wild-type littermates. Using a combination of morphological [i.e., immunohistochemical staining of circumvallate papillae (CVP)], behavioral (i.e., two-bottle preference tests, licking tests and conditioned taste aversion) and functional studies [i.e., calcium imaging in freshly isolated taste bud cells (TBCs)], we show that absence of GPR120 in the oral cavity was not associated with changes in i) gross anatomy of CVP, ii) LCFA-mediated increases in intracellular calcium levels ([Ca(2+)]i), iii) preference for oily and LCFA solutions and iv) conditioned avoidance of LCFA solutions. In contrast, the rise in [Ca(2+)]i triggered by grifolic acid, a specific GPR120 agonist, was dramatically curtailed when the GPR120 gene was lacking. Taken together, these data demonstrate that activation of lingual GPR120 and preference for fat are not connected, suggesting that GPR120 expressed in TBCs is not absolutely required for oral fat detection in mice.

Keywords: G-protein; diet and dietary lipids; fat taste; feeding behavior; lipids; mouse; nutrition; receptors.

Fig. 1.

Impacts of GPR120 gene invalidation on body weight, blood glucose, plasma insulin levels and circumvallate papilla morphology. A: Genotyping of studied mice: agarose gel electrophoresis of PCR products. B: RT-quantitative PCR of CVP (n = 5, ud, undetectable). C: Typical GPR120^−/− phenotypes compared with littermates GPR120^+/+ mice: evolution of body weight in mice fed a standard chow (n = 7), fasted blood glucose and plasma insulin levels (n = 8–12). Means ± SEM. *, P < 0.05; **, P < 0.01. D: Morphology of CVP from wild-type and GPR120^−/− mice. Gross anatomy: hematoxylin-eosin-stained CVP. Black arrows show apical taste pore areas of taste buds. Identification of taste buds cells by α-gustducin staining and visualization of taste bud cells proliferation by Ki67 staining.

Fig. 2.

Effect of GPR120 gene invalidation on the preference for lipids. Two bottles (control and experimental solutions) were simultaneously offered to wild-type and GPR120^−/− mice for 12 h. Experimental solutions contained: A: 0.02%, 0.2% or 2.0% of rapeseed oil (w/v), n = 10-12. B:2.5%, 0.25% of Intralipid (v/v), n = 10. C: 0.5% or 1% LA, n = 7. D: 2% sucrose (w/v) or 0.3 mM quinine (bitter taste), n = 10–12. Means ± SEM. Dotted line represents the absence of preference.

Fig. 3.

Effect of GPR120 gene invalidation on the conditioned avoidance of LA. Conditioned taste aversion (CTA) performed by pairing a conditioned stimulus, i.e., LA with a digestive pain obtained by intraperitoneal (i.p.) injection of LiCl (150 mM, 16µl/g of body weight), was assessed by using 1 h two-bottle preference tests. Unconditioned controls (GPR120^+/+ and GPR120^−/− mice) received an i.p. NaCl injection. A: LA-mediated CTA. Data shown are the average of two successive tests (48 h interval). There was no inter­action between the tests and no effect of the time. Means ± SEM (n = 7–9). ***, P ≤ 0.001. B: To demonstrate the specificity of LA-taste aversion, two-bottle preference test was performed with sucrose (n = 7–9). Dotted line represents the absence of preference.

Fig. 4.

Effect of GPR120 gene invalidation on the attraction for LCFAs. Short-term (1 min) licking tests in GPR120^−/− and their littermate controls (GPR120^+/+). Animals were subjected successively in a randomized manner to a control solution (mineral oil) and an experimental solution of 0.5% OLA or LA in mineral oil. Means ± SEM (n = 12). *, P < 0.05; **, P < 0.01; ***, P < 0.001.

Fig. 5.

Effects of LA or GA on Ca²⁺ signaling in mouse circumvallate TBCs. Ca²⁺ imaging studies were performed in calcium-containing (100% Ca²⁺) media. The changes in intracellular Ca²⁺ (F340/F380) were monitored under a Nikon microscope (TiU) by using S-fluor 40× oil immersion objectives, as described in the Materials and Methods section. Colored time-lapse changes show the kinetics of the rise in [Ca²⁺]_i levels in a CD36-positive TBC freshly isolated from CVP from GPR120^+/+ and GPR120^−/− mice following addition of 20 µM LA or GA in the medium. The arrows indicate when LA or GA were added into the cuvette without interruptions in the recording. A: changes (F₃₄₀/F₃₈₀) in intracellular Ca²⁺ evoked by LA, 20 µM and (B) by GA, 20 µM. C: Means ± SEM (n = 7). ***, P < 0.001. ns, nonsignificant.

Fig. 6.

Differential intracellular Ca²⁺ signaling responses induced by LA and GA in TBCs. Ca²⁺ imaging studies were performed in calcium-containing (100% Ca²⁺) media. The changes in intracellular Ca²⁺ (F340/F380) were monitored under the Nikon microscope (TiU) by using S-fluor 40× oil immersion objectives, as described in the Materials and Methods section. Colored time-lapse changes show the kinetics of the rise in [Ca²⁺]_i levels in a CD36-positive taste bud cell freshly isolated from CVP from GPR120^+/+ and GPR120^−/− mice following addition of 20 µM LA and GA one after another in the medium. The arrows indicate when LA or GA were added into the cuvette without interruptions in the recording. A: in GPR120^+/+ mice and (B) in GPR120^−/− mice. C: Means ± SEM (n = 7). *, P < 0.05; **, P < 0.01.