Exposure to the herbicide glyphosate leads to inappropriate threat responses and alters gut microbial composition

Mauricio Cáceres-Chacón¹Osmarie Martínez-Guzmán²Héctor A. Haddock-Martínez¹Alexdiel Figueroa-Pérez¹Sian Rodríguez-Rosado¹Jaleniz Suarez-Perez³Raúl Y. Ramos-Sánchez¹Filipa Godoy-Vitorino⁴Demetrio Sierra-Mercado^1*

• ¹Anatomy & Neurobiology, School of Medicine, University of Puerto Rico, San Juan, Puerto Rico
• ²Anatomy & Neurobiology ; Microbiology & Medical Zoology, School of Medicine, University of Puerto Rico, San Juan, Puerto Rico
• ³Microbiology & Medical Zoology, School of Medicine, University of Puerto Rico, San Juan, Puerto Rico
• ⁴School of Medicine, University of Puerto Rico, San Juan, Puerto Rico

Background: Exposure to the herbicide glyphosate may contribute to anxiety-related disorders. The mechanisms by which this occurs may involve effects on brain regions or alterations of gut microbiota implicated in emotions. Pre-clinical studies use unusually high doses in which humans may not normally be exposed. The effects of glyphosate on anxiety at doses considered safe are largely unexplored. Methods: Adult male rats were administered glyphosate at a dose considered safe for 16 weeks. After 4 and 10 weeks, anxiety-like behaviors were tested in the open field and elevated plus maze, respectively. After 14 weeks, rats interacted with a novel neutral object, followed by a naïve rat of the same age and sex as a conspecific. Lastly, after 16 weeks, rats underwent fear conditioning, and freezing was quantified. Upon completion of the experiments, cellular activity was assessed using immunohistochemistry in brain regions implicated in anxiety and fear. Fecal pellets were collected to extract DNA and perform 16S rRNA community analyses. Results: Glyphosate increases anxiety in the elevated plus maze after 10 weeks. Moreover, glyphosate decreases interaction to a novel object, but not to the conspecific, after 14 weeks. Furthermore, freezing increases to a novel neutral tone, but not a conditioned tone, after 16 weeks. Behavioral changes correspond to increases in cellular activity in the bed nucleus of the stria terminalis. Furthermore, we observed that glyphosate causes taxonomic changes in microbiota with a decrease of Lactobacillus genus. Conclusion: Glyphosate increases threat interpretation and alters cellular activity in brain regions implicated in promoting anxiety. Also, glyphosate induces gut dysbiosis and reduces the abundance of Lactobacillus, bacteria that play a role in the production of serotonin, which may further exacerbate the anxiogenic effect of glyphosate.