A gut-brain neural circuit for nutrient sensory transduction

INTRODUCTION In 1853, Sydney Whiting wrote in his classic Memoirs of a Stomach , “…and between myself and that individual Mr. Brain, there was established a double set of electrical wires, by which means I could, with the greatest ease and rapidity, tell him all the occurrences of the day as they arrived, and he also could impart to me his own feelings and impressions.” Historically, it is known that the gut must communicate with the brain, but the underlying neural circuits and transmitters mediating gut-brain sensory transduction still remain unknown. In the gut, there is a single layer of epithelial cells separating the lumen from the underlying tissue. Dispersed within this layer reside electrically excitable cells termed enteroendocrine cells, which sense ingested nutrients and microbial metabolites . Like taste or olfactory receptor cells, enteroendocrine cells fire action potentials in the presence of stimuli. However, unlike other sensory epithelial cells, no synaptic link between enteroendocrine cells and a cranial nerve has been described. The cells are thought to act on nerves only indirectly through the slow endocrine action of hormones, like cholecystokinin. Despite its role in satiety, circulating concentrations of cholecystokinin peak only several minutes after food is ingested and often after the meal has ended. Such a discrepancy suggests that the brain perceives gut sensory cues through faster neuronal signaling. Using a mouse model, we sought to identify the underpinnings of this neural circuit that transduces a sense from gut to brain. RATIONALE Our understanding of brain neural circuits is being propelled forward by the emergence of molecular tools that have high topographical and temporal precision. We adapted them for use in the gut. Single-cell quantitative real-time polymerase chain reaction and single-cell Western blot enabled the assessment of synaptic proteins. A monosynaptic rabies virus revealed the neural circuit’s synapse. The neural circuit was recapitulated in vitro by using nodose neurons cocultured with either minigut organoids or purified enteroendocrine cells. This system, coupled to optogenetics and whole-cell patch-clamp recording, served to determine the speed of transduction. Whole-nerve electrophysiology, along with optical excitation and silencing, helped to uncover the neurotransmission properties of the circuit in vivo. The underlying neurotransmitter was revealed by using receptor pharmacology and a fluorescent reporter called iGluSnFR. RESULTS Single-cell analyses showed that a subset of enteroendocrine cells contains presynaptic adhesion proteins, including some necessary for synaptic adhesion. Monosynaptic rabies tracing revealed that enteroendocrine cells synapse with vagal nodose neurons. This neuroepithelial circuit connects the intestinal lumen with the brainstem in one synapse. In coculture, this connection was sufficient to transduce a sugar stimulus from enteroendocrine cells to vagal neurons. Optogenetic activation of enteroendocrine cells elicited excitatory postsynaptic potentials in connected nodose neurons within milliseconds. In vivo recordings showed that enteroendocrine cells are indeed necessary and sufficient to transduce a sugar stimulus to the vagus. By using iGluSnFR, we found that enteroendocrine cells synthesize the neurotransmitter glutamate, and pharmacological inactivation of cholecystokinin and glutamate receptors revealed that these cells use glutamate as a neurotransmitter to transduce fast, sensory signals to vagal neurons. CONCLUSION We identified a type of gut sensory epithelial cell that synapses with vagal neurons. This cell has been referred to as the gut endocrine cell, but its ability to form a neuroepithelial circuit calls for a new name. We term this gut epithelial cell that forms synapses the neuropod cell. By synapsing with the vagus nerve, neuropod cells connect the gut lumen to the brainstem. Neuropod cells transduce sensory stimuli from sugars in milliseconds by using glutamate as a neurotransmitter. The neural circuit they form gives the gut the rapidity to tell the brain of all the occurrences of the day, so that he, too, can make sense of what we eat.