Activation of the sympathetic nervous system mediates hypophagic and anxiety-like effects of CB₁ receptor blockade

Abstract

Complex interactions between periphery and the brain regulate food intake in mammals. Cannabinoid type-1 (CB1) receptor antagonists are potent hypophagic agents, but the sites where this acute action is exerted and the underlying mechanisms are not fully elucidated. To dissect the mechanisms underlying the hypophagic effect of CB1 receptor blockade, we combined the acute injection of the CB1 receptor antagonist rimonabant with the use of conditional CB1-knockout mice, as well as with pharmacological modulation of different central and peripheral circuits. Fasting/refeeding experiments revealed that CB1 receptor signaling in many specific brain neurons is dispensable for the acute hypophagic effects of rimonabant. CB1 receptor antagonist-induced hypophagia was fully abolished by peripheral blockade of β-adrenergic transmission, suggesting that this effect is mediated by increased activity of the sympathetic nervous system. Consistently, we found that rimonabant increases gastrointestinal metabolism via increased peripheral β-adrenergic receptor signaling in peripheral organs, including the gastrointestinal tract. Blockade of both visceral afferents and glutamatergic transmission in the nucleus tractus solitarii abolished rimonabant-induced hypophagia. Importantly, these mechanisms were specifically triggered by lipid-deprivation, revealing a nutrient-specific component acutely regulated by CB1 receptor blockade. Finally, peripheral blockade of sympathetic neurotransmission also blunted central effects of CB1 receptor blockade, such as fear responses and anxiety-like behaviors. These data demonstrate that, independently of their site of origin, important effects of CB1 receptor blockade are expressed via activation of peripheral sympathetic activity. Thus, CB1 receptors modulate bidirectional circuits between the periphery and the brain to regulate feeding and other behaviors.

Fig. 1.

The hypophagic effect of rimonabant does not depend on CB₁ expression in brain neurons. Effect of rimonabant (3 mg/kg, i.p., black bars) or vehicle (white bars) in (A) mutant Glu-CB₁-KO mice; (B) GABA-CB₁-KO mice; (C) TPH2-CB₁-KO mice; (D) Sim1-CB₁-KO mice; (E) SF1-CB₁-KO mice, and respective WT littermates. (C, Right) PCR on genomic DNA extracted from dorsal raphe of WT and TPH2-CB₁-KO mice (KO). CB₁-floxed allele (Lower); deleted CB₁ allele (Upper). Data are means ± SEM. n = 6–9 mice per group. Statistics by two-way ANOVA followed by Bonferroni’s posttest. **P < 0.01, ***P < 0.001.

Fig. 2.

Blockade of peripheral β-adrenergic receptors prevents rimonabant-induced hypophagia. Effect of rimonabant (black bars, 3 mg/kg, i.p.; gray bars, 10 mg/kg, i.p.) or its vehicle (white bars) in fasted mice pretreated with (A) the β-blocker propranolol (Prop, 10 mg/kg, i.p.) or with the peripherally restricted β-blocker sotalol (Sot, 3 mg/kg, i.p.); (B) the β-blocker sotalol administered centrally (2 μg intracerebroventricularly); (C) the selective β3-blocker SR59320A (1 mg/kg, i.p.); (D) the ganglionic blocker hexamethonium (Hex, 20 mg/kg, i.p.). (E) Effect of β-blocker sotalol (3 mg/kg, i.p.) and rimonabant (Rim, 3 mg/kg, i.p.) in conditions of glucoprivation induced by 2-DG injection (250 mg/kg, i.p.). (F) Effect of β-blocker sotalol (3 mg/kg, i.p.) and rimonabant (3 mg/kg, i.p.) in conditions of lipoprivation induced by MA injection (68 mg/kg, i.p.). Data are means ± SEM. n = 5–8 mice per group. Statistics by two-way ANOVA followed by Bonferroni’s posttest. *P < 0.05, **P < 0.01, ***P < 0.001.

Fig. 3.

CB₁ receptor blockade enhances gastrointestinal activity and increases glutamatergic transmission in the caudal brainstem trough vagal innervation to reduce food intake. (A) Three-dimensional whole-body small animal PET images showing ¹⁸F-FDG uptake in the intestine of mice treated with vehicle, rimonabant (10 mg/kg, i.p.), or rimonabant + sotalol (3 mg/kg, i.p.). Gradation bar, signal intensity expressed as radioactive counts. (B) Quantification [standard uptake value (SUV) max of ¹⁸F-FDG uptake] of data in A. Black bars, rimonabant; white bars, vehicle. (C) Effect of capsaicin-induced deafferentiation on rimonabant- (3 mg/kg, i.p., black bars) or vehicle- (white bars) induced changes in food intake. (D) Effect of the combination of fourth-ventricle injection of MK801 (MK, 2 µg in 2 µL) and rimonabant (Rim, 3 mg/kg, i.p., black bars) or their respective vehicles (white bars) on food intake. The effects of a direct injection of vehicle (V, white bar) or rimonabant (Rim, 10 µg, gray bar) in the fourth ventricle are presented in the last two bars of the graph. (E) Effect of the combination of NTS bilateral injection of MK801 (0.2 µg in 0.2 µL) and rimonabant (3 mg/kg, i.p., black bars) or their respective vehicles (white bars) on food intake. The effects of a direct bilateral injection of vehicle (V, white bar) or rimonabant (Rim 1 µg per site, gray bar) in the NTS are presented in the last two bars of the graph. Data are means ± SEM. n = 4–12 mice per group. Statistics by two-way ANOVA followed by Bonferroni’s posttest. *P < 0.05, **P < 0.01.

Fig. 4.

Peripheral β-adrenergic signaling mediates the effects of rimonabant on fear-induced freezing and anxiety. (A) Effects of sotalol (3 mg/kg) and rimonabant (10 mg/kg) on fear-induced freezing. (Left) Time-course (1-min bins) over the 8-min tone presentation; (Right) total freezing. (B) Effects of sotalol (Sot, 3 mg/kg) and systemic rimonabant (Rim, 10 mg/kg) on the elevated-plus-maze test. (C) Effects of sotalol (3 mg/kg) and central rimonabant (10 µg intracerebroventricularly) on the elevated-plus-maze test. Data are means ± SEM. n = 8–10 mice per group. Statistics by using one- or two-way ANOVA followed by Bonferroni’s posttest. *P < 0.05; **P < 0.01.