The gut microbiota affects tissue physiology, metabolism, and function of both the immune and nervous systems. We found that intrinsic enteric-associated neurons (iEANs) in mice are functionally adapted to the intestinal segment they occupy; ileal and colonic neurons are more responsive to microbial colonization than duodenal neurons. Specifically, a microbially responsive subset of viscerofugal CART(+) neurons, enriched in the ileum and colon, modulated feeding and glucose metabolism. These CART(+) neurons send axons to the prevertebral ganglia and are polysynaptically connected to the liver and pancreas. Microbiota depletion led to NLRP6- and caspase 11-dependent loss of CART(+) neurons and impaired glucose regulation. Hence, iEAN subsets appear to be capable of regulating blood glucose levels independently from the central nervous system.