Intestinal enteric neurons are required for luminal nutrient sensing and vagal afferent neurotransmission and are dysregulated in diet-induced obesity

Introduction: Intestinal nutrient sensing and vagal afferent neurotransmission are mediated by neuroendocrine mechanisms. Although the endocrine players have been described, the role of enteric neurons remains unknown. Hypotheses: (1) Enteric neurons of the small intestine (SI) are activated by glucose and oleic acid (OA); Diet-induced obesity alters (2) activation profiles of small intestinal enteric neurons, and (3) activation of afferent vagal cell bodies in the nodose ganglia. Methods: Designer Receptor Activated by Designer Drugs (DREADD) strategy was used to selectively inactivate enteric neurons in Baf53b-Cre mice that targets enteric neurons. Mice were fed low fat (10% calories) or high-fat diet (45% calories) for 12 weeks, transduced with Cre-dependent AAV-hSYN-hM4D-mCherry and inactivated with Clozapine N-oxide (1mg/kg) or saline (control). Blood was drawn at baseline and 15-90 mins after intestinal infusions of glucose (2mg/kg) or oleic acid (16µL/g) for gut hormone measurements, and nodose ganglia were harvested for vagal activation measurements. In separate cohorts, primary neuronal cultures were generated from small intestinal submucosal plexus (SP) and longitudinal muscle myenteric plexus (LMMP) of lean and diet-induced obese (DIO) wildtype (WT) mice. Real-time activity of enteric neurons was measured using live imaging of intracellular calcium transients (Cai2+), surrogate for neuronal activation. Results: Glucose (25mM) and OA (50μM) induced a 2-4-fold increase in Cai2+ fluorescence intensity (ΔF/F0= 2.2-4.3; p =0.003, 0.02, respectively) in neurons isolated from SPs of lean WT mice. No response was observed in DIO mice at the above concentrations, suggesting a decline in nutrient sensitivity. At higher concentrations of glucose (50mM) and OA (100μM), DIO mice showed a marked decrease in ΔF/F0 (1.3-2.7; p =0.03, 0.005) along with significant delay in response (glucose: 24.4s, p =0.004; OA: 18.8s, p =0.006). LMMP neurons were less responsive to nutrient stimulation (ΔF/F0=1.6-2.3; p =0.06). Chemogenetic inactivation of enteric neurons dramatically blunted glucose and OA-induced release of acyl ghrelin (-67.3%, p =0.025; -73%, p =0.001), cholecystokinin (-58.5%, p =0.006; -82.6%, p =0.005), GLP-1 (57.3%, p =0.035; -43%, p =0.023), and peptide YY (-41.8%, -56.3%, p =0.03) in lean Baf53b-Cre mice, indicating that enteric neurons are required for nutrient-driven gut hormone release. Concomitantly, inactivation of enteric neurons led to a marked reduction in c-Fos-positive cell bodies in the nodose ganglia (53%, p =0.004; -43.7%, p =0.05). Conclusion: This is the FIRST report that provides evidence of direct activation of enteric neurons by nutrients and demonstrates that DIO blunts intestinal neuronal activation and afferent vagal activation, laying the foundation for therapeutic strategies targeting dysregulated intestinal sensing for DIO amelioration. CCOM Michael Walczak Research Award for the study of Diabetes, Heart attack, and Stroke Midwestern Start-up Funds This abstract was presented at the American Physiology Summit 2025 and is only available in HTML format. There is no downloadable file or PDF version. The Physiology editorial board was not involved in the peer review process.