Integration of sensory and molecular inputs from the environment shapes animal behavior. A major site of exposure to environmental molecules is the gastrointestinal tract, where dietary components are chemically transformed by the microbiota 1 and gut-derived metabolites are disseminated to all organs, including the brain 2 . In mice, the gut microbiota impacts behavior 3 , modulates neurotransmitter production in the gut and brain 4,5 , and influences brain development and myelination patterns 6,7 . Mechanisms mediating gut-brain interactions remain poorly defined, though broadly involve humoral or neuronal connections. We previously reported that levels of the microbial metabolite 4-ethylphenyl sulfate (4EPS) were elevated in a mouse model of atypical neurodevelopment 8 . Herein, we identified biosynthetic genes from the gut microbiome that mediate conversion of dietary tyrosine to 4-ethylphenol (4EP), and bioengineered gut bacteria to selectively produce 4EPS in mice. 4EPS entered the brain and was associated with changes in region-specific activity and functional connectivity. Gene expression signatures revealed altered oligodendrocyte function in the brain, and 4EPS impaired oligodendrocyte maturation in mice as well as decreased oligodendrocyte-neuron interactions in ex vivo brain cultures. Mice colonized with 4EP-producing bacteria exhibited reduced myelination of neuronal axons. Altered myelination dynamics in the brain have been associated with behavioral outcomes 7,[9][10][11][12][13][14]13,14 . Accordingly, we observed that mice exposed to 4EPS displayed anxiety-like behaviors, and pharmacologic treatments that promote oligodendrocyte differentiation prevented the behavioral effects of 4EPS. These findings reveal that a gut-derived molecule influences complex behaviors in mice via effects on oligodendrocyte function and myelin patterning in the brain.

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Editorial summaryThe gut microbiome contributes to the development and function of the immune, metabolic, and nervous systems. Furthermore, commensal bacteria modulate symptoms and pathology in mouse models of neuropsychiatric and neurodevelopmental diseases. Uncovering mechanisms that are utilized by the microbiome to mediate gut–brain connections may provide novel opportunities to target therapies to the gut in order to treat neurologic disorders.

Significance Many human diseases are causally linked to the gut microbiota, yet the field still lacks mechanistic understanding of the underlying complex interactions, because existing tools cannot simultaneously quantify microbial communities and their native context. In this work, we provide an approach to tissue clearing and preservation that enables 3D visualization of the biogeography of the host–microbiota interface. We combine this tool with sequencing and multiplexed microbial labeling to provide the field with a platform on which to discover patterns in the spatial distribution of microbes. We validated this platform by quantifying bacterial distribution in cecal mucosa at different stages of antibiotic exposure. This approach may enable researchers to formulate and test new hypotheses about host–microbe and microbe–microbe interactions.

The enteric nervous system (ENS) integrates cues from the brain and from local signals in the gut to coordinate responses that shape the intestinal milieu. Tools to study, and knowledge of, the ENS lag behind brain research. Herein, we deploy novel recombinant adeno-associated viral (rAAV) vectors with enhanced tropism for the gut to map and activate enteric neurons with spatial and temporal resolution. rAAV-mediated fluorescent labelling coupled with whole-tissue clearing methods in the small intestine and colon of mice enables efficient and thorough characterization of the neuronal architecture of the ENS. We also delivered "designer receptors exclusively activated by designer drugs" (DREADDs) to specifically activate gut neurons that express choline acetyltransferase (ChAT+) and tyrosine hydroxylase (TH+). Targeted activation of ChAT+ or TH+ neuronal populations associated with the gastrointestinal (GI) tract altered the intestinal transcriptome of mice, host and microbial proteome, metagenome, and fecal metabolome. We reveal previously unknown and broad roles for neurons in modulating intestinal physiology, mucosal immunity, and gut microbiome structure, and propose novel interactions for the ENS such as regulating fungal colonization and shaping of bile acid profiles. Experimental tools and rich datasets with multi-parameter characterization of the gut ecosystem may enable further study of the ENS.