<p>Multiple sclerosis constitutes a chronic, inflammatory, and demyelinating disorder affecting the central nervous system, with an estimated global prevalence of 2.5 million individuals. Emerging research underscores the significant influence of the gut microbiota on the immune system, suggesting a potential role in the initiation and progression of inflammatory diseases. This study investigated the potential therapeutic effects of <i>Lactobacillus</i> and <i>Bifidobacterium</i> species isolated from traditional dairy products on cuprizone-induced demyelination in a rat model. 48 adults male Wistar rats were randomly assigned to six groups. Demyelination was induced by daily oral administration of 0.6% (w/w) cuprizone mixed with food for 30&#xa0;days. Subsequently, treated groups received oral administration of mixed of <i>Lactiplantibacillus plantarum</i>, <i>Lactobacillus acidophilus</i>, and <i>Lactobacillus reuteri</i>: and mixed of <i>Bifidobacterium bifidum</i> and <i>Bifidobacterium animalis</i>. A control group received no bacteria intervention. Behavioral deficits were assessed using grip-traction, beam-walking, and grid-walking tests. Oxidative stress biomarkers were quantified using colorimetric assays. The extent of demyelination was evaluated by hematoxylin and eosin staining of the corpus callosum. Serum levels of vitamin D₃ and B₁₂ were measured by ELISA. The results demonstrated that lactic acid bacteria supplementation significantly improved behavioral deficits and reduced demyelination in the corpus callosum. Furthermore, these bacteria administration was associated with reduced oxidative stress and increased serum levels of vitamin D₃ and B₁₂. These findings suggest that <i>Lactobacillus</i> and <i>Bifidobacterium</i> species may offer a supplementary therapeutic strategy for demyelinating disorders such as multiple sclerosis, potentially by mitigating oxidative stress, promoting remyelination, and enhancing vitamin D₃ and B₁₂ levels.</p>