Gut microbes promote colonic serotonin production through an effect of short-chain fatty acids on enterochromaffin cells

Abstract

Gut microbiota alterations have been described in several diseases with altered gastrointestinal (GI) motility, and awareness is increasing regarding the role of the gut microbiome in modulating GI function. Serotonin [5-hydroxytryptamine (5-HT)] is a key regulator of GI motility and secretion. To determine the relationship among gut microbes, colonic contractility, and host serotonergic gene expression, we evaluated mice that were germ-free (GF) or humanized (HM; ex-GF colonized with human gut microbiota). 5-HT reduced contractile duration in both GF and HM colons. Microbiota from HM and conventionally raised (CR) mice significantly increased colonic mRNAs Tph1 [(tryptophan hydroxylase) 1, rate limiting for mucosal 5-HT synthesis; P < 0.01] and chromogranin A (neuroendocrine secretion; P < 0.01), with no effect on monoamine oxidase A (serotonin catabolism), serotonin receptor 5-HT4, or mouse serotonin transporter. HM and CR mice also had increased colonic Tph1 protein (P < 0.05) and 5-HT concentrations (GF, 17 ± 3 ng/mg; HM, 25 ± 2 ng/mg; and CR, 35 ± 3 ng/mg; P < 0.05). Enterochromaffin (EC) cell numbers (cells producing 5-HT) were unchanged. Short-chain fatty acids (SCFAs) promoted TPH1 transcription in BON cells (human EC cell model). Thus, gut microbiota acting through SCFAs are important determinants of enteric 5-HT production and homeostasis.

Keywords: 5-hydroxytryptamine; gnotobiotic; gut motility; host-microbe interaction; microbiome.

Figure 1.

GF and HM colonic segments show similar contractile responses to exogenous, luminal 5-HT ex vivo. A) Intracolonic pressure was measured in midcolonic segments ex vivo (arrowhead indicates migrating contraction). B) Representative recordings from GF and HM segments, with and without addition of luminal 5-HT (10 μM). C) Basal contractile frequency of HM and GF colonic segments (4 mice per group). Student’s t test: P > 0.05. D) Mean contractile duration (seconds) measured at half-maximal amplitude over 10 minutes (4 mice per group). Two-way ANOVA: *P < 0.05.

Figure 2.

Tph1 mRNA and protein expression are increased in the proximal colon of HM and CR mice compared with GF mice, without alteration of 5-HT catabolic (Maoa) and transporter (Slc6a4) mRNAs. A) Relative expression of Tph1 mRNA was determined in the proximal colon by qRT-PCR (14–16 mice per group). One-way ANOVA: **P < 0.01; ****P < 0.0001. B) Western blot detection of Tph1 and Gapdh from proximal colonic lysates. C) Quantification of relative Tph1 protein in the proximal colon of GF, HM, and CR mice (9 mice per group). One-way ANOVA: *P < 0.05.

Figure 3.

Presence of human- or mouse-derived microbiota increases tissue concentration of 5-HT and Chga mRNA in the mouse proximal colon. A) 5-HT concentration was assessed in tissue lysates from the proximal colon by ELISA [8 mice per group (4 males)]. One-way ANOVA: *P < 0.05; ***P < 0.001. B) Relative expression of Chga mRNA (Chga, a neuroendocrine secretory marker) was determined in the proximal colon by qRT-PCR (14–16 mice per group). One-way ANOVA: *P < 0.05; ***P < 0.001.

Figure 4.

SCFAs, but not LPS, modulate TPH1 expression in a human model of EC cells. Relative expression is shown of TPH1 mRNA in BON cells treated with different concentrations of SCFAs (A) acetate and (B) butyrate or LPSs from (C) commensal K12 E. coli and (D) pathogenic O111:B4 E. coli (3 replicate treatments per group). One-way ANOVA, Dunnett multiple comparison test: **P < 0.01; ***P < 0.001.