Microplastics (MPs) raise environmental concerns. However, their effects on the ruminal–gastro-intestinal system have not yet been studied. This study aims to investigate the effects of polyethylene terephthalate (PET) MPs on the ability of the ruminal–gastro-intestinal system to degrade and digest mixed hay. Using a three-step in vitro ruminal–gastro-intestinal incubation system, PET MPs were introduced at concentrations of 0, 5, 10, and 15 g/L in ruminal and gastro-intestinal solutions. Ruminal fluid was collected from three 16-month-old Piedmontese bulls. The experiment was conducted on three mixed hays and was repeated three times, with triplicate incubations in each run. The results reveal that PET MPs reduced the degradability and digestibility of crude protein. Specifically, crude protein degradation was reduced by 9% at medium and 16% at high PET MP concentrations in the ruminal phase, while the crude protein digestibility of undegraded crude protein was reduced by 8% at the lowest PET MPs concentration in the gastro-intestinal tract. Additionally, PET MPs reduced the degradation of neutral detergent fiber at medium and high PET MP concentrations in the ruminal phase by 9% and 13%, respectively. These results highlight the risks of PET MPs contamination on ruminal–gastro-intestinal functions and underscore the urgent need to mitigate MPs contamination in the livestock sector.