Activation of Ventral Tegmental Area 5-HT2C Receptors Reduces Incentive Motivation

Abstract

Obesity is primarily due to food intake in excess of the body's energetic requirements, intake that is not only associated with hunger but also the incentive value of food. The 5-hydroxytryptamine 2C receptor (5-HT_2CR) is a target for the treatment of human obesity. Mechanistically, 5-HT_2CRs are positioned to influence both homeostatic feeding circuits within the hypothalamus and reward circuits within the ventral tegmental area (VTA). Here we investigated the role of 5-HT_2CRs in incentive motivation using a mathematical model of progressive ratio (PR) responding in mice. We found that the 5-HT_2CR agonist lorcaserin significantly reduced both ad libitum chow intake and PR responding for chocolate pellets and increased c-fos expression in VTA 5-HT_2CR expressing γ-aminobutyric acid (GABA) neurons, but not 5-HT_2CR expressing dopamine (DA) neurons. We next adopted a chemogenetic approach using a 5-HT_2CR^CRE line to clarify the function of subset of 5-HT_2C receptor expressing VTA neurons in the modulation of appetite and food-motivated behavior. Activation of VTA 5-HT_2C receptor expressing neurons significantly reduced ad libitum chow intake, operant responding for chocolate pellets, and the incentive value of food. In contrast, chemogenetic inhibition of VTA 5-HT_2C receptor expressing neurons had no effect on the feeding behavior. These results indicate that activation of the subpopulation of 5-HT_2CR neurons within the VTA is sufficient to significantly reduce homeostatic feeding and effort-based intake of palatable food, and that this subset has an inhibitory role in motivational processes. These findings are relevant to the treatment of obesity.

Figure 1

Effect of lorcaserin on food intake and operant responding. (a) Lorcaserin 10 mg/kg reduced 1 h chow intake (n=14). (b) Lorcaserin 3 and 10 mg/kg reduced the breakpoint when mice were in ad libitum condition (n=9). (c) Lorcaserin 10 mg/kg reduced the breakpoint when mice were in food-restricted condition (n=9). (d–f) Effect of lorcaserin on performance on the progressive-ratio schedule. Ordinate, response rate; abscissa, response/reinforcer ratio, N. Points are group mean data: unfilled symbols indicate running response rate, filled symbols indicate overall response rate. The curves are best-fit functions defined by equations (2) and (3). (g) Schematic of the location of the VTA defined by the Mouse Brain Atlas (Paxinos and Franklin, 2001). (h) Lorcaserin 7 and 12 mg/kg increased FOS-IF (blue) within the VTA (n=6–7 per group). (i–k) FOS-IF (blue) within the VTA following administration of vehicle, lorcaserin 7 or 12 mg/kg (scale bar: 100 μm). All the data are presented as mean±SEM. ^∗p<0.05; ^∗∗p<0.01; ^∗∗∗p<0.001.

Figure 2

5-HT_2CR^CRE:YFP VTA neurons co-express GABA and dopamine. (a) A 5-HT_2CR^CRE mouse line in which Cre recombinase is driven by a 5-HT_2CR promoter was intercrossed with a ROSA26-stop-EYFP reporter mouse to generate a 5-HT_2CR^CRE:YFP line to facilitate the visualization of VTA 5-HT_2CR expressing cells. (b) VTA 5-HT_2CR^CRE:YFP containing neurons (GFP-IF) have a greater co-expression with GAD67-IF compared with TH-IF (n=4 per group, p<0.001). (c) VTA 5-HT_2CR^CRE:YFP (GFP-IF; green) are co-localized (overlay, yellow) with endogenous 5-HT_2CR-IF (red). (d) VTA 5-HT_2CR^CRE:YFP (GFP-IF; green) containing neurons and GAD67 (red) are co-localized (yellow). (e) VTA 5-HT_2CR^CRE:YFP (GFP-IF; green) containing neurons and TH (red) are co-localized (yellow). All the data are presented as mean±SEM. ^∗∗∗p<0.001. Scale bar: 100 μm.

Figure 3

Lorcaserin increases c-Fos in VTA 5-HT_2CR GABAergic, but not dopamine neurons. (a-c) c-Fos expression (FOS-IR, gray) in VTA 5-HT_2CR containing neurons (GFP-IF, green) co-expressing GAD67 (GAD67-IF, red) in 5-HT_2CR^CRE:YFP mice following (a) vehicle, (b) lorcaserin 7 mg/kg or (c) 12 mg/kg administration. (d–f) c-Fos expression (FOS-IR, gray) in VTA 5-HT_2CR containing neurons (GFP-IF, green) co-expressing TH (TH-IF, red) in 5-HT_2CR^CRE:YFP mice following (d) vehicle, (e) lorcaserin 7 mg/kg or (f) 12 mg/kg administration. (g) Lorcaserin dose-dependently increases FOS-IR in GAD67-positive 5-HT_2CR expressing neurons (n=4 per group, p<0.001), but not in (h) TH-positive 5-HT_2CR expressing neurons (n=4 per group, NS). All the data are presented as mean±SEM. ^∗p<0.05; ^∗∗p<0.01. Scale bar: 50 μm.

Figure 4

Chemogenetic activation of VTA 5-HT_2CR neurons reduces food intake and operant responding for food reward. (a) Representative image of Cre-dependent expression of hM3Dq-mCherry specifically within the VTA of a 5-HT_2CR^CRE:YFP mouse. (b) Firing rate of 5-HT_2CR^CRE:YPF::hM3D_q-mCherry^VTA neurons increased upon 10 μM CNO application. (c–e) CNO 2 mg/kg increased FOS-IR within 5-HT_2CR^CRE:YPF::hM3D_q-mCherry^VTA compared with vehicle (n=3 per group). (f) CNO 2 mg/kg reduced 1 h chow intake (n=14, p<0.05). (g) CNO 2 mg/kg reduced the breakpoint in the experimental group [AAV-hM3D (G_q)] compared with control [AAV-mCherry] in ad libitum condition (n=15) and (h) food-restricted condition (n=15). (i and j) Effect of chemogenetic activation of VTA 5-HT_2CR neurons on performance on a PR schedule. Ordinate, response rate; abscissa, response/reinforcer ratio, N. Points are group mean data: unfilled symbols indicate running response rate, filled symbols indicate overall response rate. The curves are best-fit functions defined by equations (2) and (3). All the data are presented as mean±SEM. ^∗p<0.05; ^∗∗p<0.01; ^∗∗∗p<0.001. Scale bar: 100 μm.

Figure 5

Chemogenetic inhibition of VTA 5-HT_2CR neurons does not alter food intake or operant responding for food reward. (a) Representative image of Cre-dependent expression of hM4D(Gi)-mCherry specifically within the VTA of a 5-HT_2CR^CRE:YFP mouse. (b) Firing rate of 5-HT_2CR^CRE:YPF::hM4D_i-mCherry^VTA neurons decreased upon 10 μM CNO application. (c) CNO 2 mg/kg had no effect on 1 h chow intake (n=14). (d) CNO 2 mg/kg had no effect on the breakpoint in the experimental group [AAV-hM4D (G_i)] compared with the control [AAV-mCherry] in ad libitum condition (n=29) or (e) food-restricted condition (n=30).