Acute stress exposures are the building blocks of chronic stress. Unravelling the mechanisms underpinning the whole-body response to acute stress is critical to understand this neglected aspect of health and disease. A single acute stress exposure rapidly modulates gut-brain axis signalling including intestinal permeability and hippocampal plasticity. The gut microbiota is increasingly recognized as an important modulator of mood and cognition, and microbially-produced metabolites such as butyrate can be a key regulator of brain and gut function via the gut-brain axis. The aim of this work is to determine if acute stress regulates the microbial production of butyrate in the gut to modulate stress-induced alterations in barrier function and hippocampal plasticity. Mice underwent acute restraint stress, and butyrate production was analysed in caecal content at contemporaneous timepoints to long-term potentiation (LTP) measured by electrophysiology. Colonic T84 epithelial and mouse brain endothelial (bEnd3) cells were used to evaluate barrier function. Butyrate, which was reduced in stressed mice, prevented Lipopolysaccharides (LPS)-induced disruption of gut and brain barrier function in a dose-dependent manner. We also expanded our evaluation of key electrophysiological indices of hippocampal plasticity, showing that LTP was modified following acute stress. In conclusion, the production of butyrate was reduced by acute restraint stress and has protective effects on gut and brain barrier disruption. Future work will evaluate the role of butyrate in acute stress-induced alterations in hippocampal plasticity.