Glyphosate Alters the Neurochemical Phenotype of Enteric Neurons and Modulates Cytokine Levels in the Porcine Ileum.

Glyphosate is one of the most widely used herbicides worldwide, yet its effects on the enteric nervous system (ENS) remain insufficiently understood. This study aimed to evaluate the impact of glyphosate exposure on the neurochemical phenotype of enteric neurons and levels of selected pro-inflammatory cytokines in the porcine ileum. Fifteen immature gilts were randomly assigned to three groups: control (C), low-dose glyphosate (G1; corresponding to the theoretical maximum daily intake, 0.05mg/kg body weight/day), and higher-dose glyphosate (G2; corresponding to the acceptable daily intake, 0.5mg/kg body weight/day) for 28 days. Double immunofluorescence staining was used to assess neurons immunoreactive to CART, nNOS, PACAP, CGRP, and VAChT in the intramural plexuses. Cytokine levels (IL-1β, IL-6, and TNF-α) were determined in ileal tissue homogenates using ELISA. Glyphosate exposure induced dose-dependent alterations in neurochemical coding of enteric neurons. The number of CART-, nNOS-, PACAP- and CGRP-immunoreactive neurons increased, whereas VAChT-positive neurons decreased, particularly in the high-dose group. These changes were observed across all plexuses, although their extent varied depending on neuronal subtype and localization. Additionally, IL-1β and IL-6 levels were significantly elevated in the high-dose group, while TNF-α levels remained unchanged. In conclusion, glyphosate exposure at doses corresponding to current safety thresholds is associated with alterations in the neurochemical phenotype of enteric neurons and changes in the local cytokine milieu in the ileum. These findings suggest glyphosate may contribute to disturbances in gut homeostasis through concurrent modulation of enteric neuronal phenotype and local inflammatory signalling.