Short-chain fatty acids suppress food intake by activating vagal afferent neurons

Abstract

Fermentable carbohydrates including dietary fibers and resistant starch produce short-chain fatty acids (SCFAs), including acetate, propionate and butyrate, through microbial fermentation in the intestine of rodents and humans. Consumption of fermentable carbohydrate and SCFAs suppress food intake, an effect involving the brain. However, their signaling pathway to the brain remains unclear. Vagal afferents serve to link intestinal information to the brain. In the present study, we explored possible role of vagal afferents in the anorexigenic effect of SCFAs. Intraperitoneal (ip) injection of three SCFA molecules (6 mmol/kg) suppressed food intake in fasted mice with the rank order of butyrate > propionate > acetate. The suppressions of feeding by butyrate, propionate and acetate were attenuated by vagotomy of hepatic branch and blunted by systemic treatment with capsaicin that denervates capsaicin-sensitive sensory nerves including vagal afferents. Ip injection of butyrate induced significant phosphorylation of extracellular-signal-regulated kinase 1/2, cellular activation markers, in nodose ganglia and their projection site, medial nucleus tractus solitaries. Moreover, butyrate directly interacted with single neurons isolated from nodose ganglia and induced intracellular Ca²⁺ signaling. The present results identify the vagal afferent as the novel pathway through which exogenous SCFAs execute the remote control of feeding behavior and possibly other brain functions. Vagal afferents might participate in suppression of feeding by intestine-born SCFAs.

Introduction

Several intestinal and pancreatic hormones are released in response to meal and induce satiety, which include cholecystokinin (CCK), glucagon-like peptide-1 (GLP-1), peptide YY_3–36 (PYY_3–36), insulin and pancreatic polypeptide (PP) [1], [2]. These peripheral hormones send their information to the brain regions regulating feeding via two distinct routes: (a) penetrating the blood–brain barrier (BBB) to act directly on the brain and (b) interacting with the vagal afferent nerves that transmit peripheral signals to the nucleus tractus solitaries (NTS) of the medulla [1], [2]. In general, the entry of hormones into the brain is strictly limited by BBB [3]. The vagal afferents terminate in the lamina propria of intestinal mucosa, portal area and pancreas [4], [5], [6], and can be easily accessed by intestinal and pancreatic hormones in active forms immediately after they are released. In fact, it has been shown that vagal afferent nerves are directly activated by the postprandial hormones GLP-1, CCK, PYY_3–36, nesfatin-1, insulin, PP and glucagon [2], [7]. Some of these interactions are linked to suppression of food intake [2], [7].

Accumulating evidences indicate that supplementing fermentable carbohydrate, including dietary fibers and resistant starch, in diet reduces food intake and body weight gain and improves glucose metabolism in rodents and humans [8], [9]. These effects are suggested to be mediated partly by short-chain fatty acids (SCFAs). SCFAs are produced by microbiota in the colon and the distal small intestine from indigestible carbohydrate (resistant starch, dietary fiber and other low-digestible polysaccharides) in a fermentation process [10], [11], [12]. Acetate, propionate and butyrate are the predominant SCFAs in the intestinal lumen in humans and rodents [13]. These SCFAs in intestinal lumen act on the enteroendocrine cells and promote the release of the anorectic gut hormones PYY and GLP-1 [14], [15]. The SCFAs also enter the circulation and can influence various organs and central and peripheral nerves systems [10], [11]. Frost et al. have reported that intraperitoneal (ip) injection of acetate reduces food intake via entering the brain over the BBB and directly activating hypothalamic neurons [10]. However, the anorexigenic effect of acetate was weaker after administration into the third ventricle than ip injection, suggesting that circulating SCFAs might also act on the sites other than the brain [10]. The possible interaction of SCFAs with vagal afferent nerves may play a key role.

The present study first examined the efficacy and temporal profile of the anorexigenic effects of three SCFAs (acetate, propionate and butyrate) ip injected in mice. We also explored whether the anorexigenic effects of three SCFAs, especially butyrate, are mediated by vagal afferents and furthermore through its direct interaction with nodose ganglion neurons (NGNs) of vagal afferents. Involvement of vagal afferents was examined by chemical denervation with capsaicin treatment and surgical hepatic vagotomy. In vivo activation of NGNs and medial NTS, to which vagal afferents project, was assessed by phosphorylation of extracellular-signal-regulated kinase 1/2 (ERK1/2), cellular activity markers [16], [17]. The direct effect of butyrate on single NGNs was monitored by imaging of cytosolic Ca²⁺ concentration ([Ca²⁺]_i) with fura-2. We found that ip butyrate had highest efficacy among the three SCFAs in inhibiting food intake. Systemic capsaicin treatment completely and hepatic vagotomy markedly counteracted the anorexigenic effects of acetate, propionate or butyrate. Butyrate ip injection increased ERK1/2 phosphorylation in NGN and NTS in vivo, and butyrate increased [Ca²⁺]_i in isolated NGNs ex vivo.