Mesenchymal stem cells (MSCs) have been shown in various animal models to be capable of neurorepair and neuroprotection. To carry out a therapeutic function, MSCs must be delivered to the target organ. MSCs are administered to patients via systemic infusion, which has many drawbacks, including a low engraftment rate and the migration of MSCs to non-target organs. However, other approaches such as direct intracerebral injection of MSCs might cause cerebral bleeding. In this study, a traumatic brain injury (TBI) was induced over the right parietal cerebral cortex in Sprague Dawley rats, and green fluorescent protein (GFP)-expressing MSCs (GFP-MSCs), together with a thin layer of fibrin, were applied to the external surface of the contralateral side 2 days later. Within 5 days of topical application, the GFP-MSCs had migrated from the site of application on the cortical surface, through the white matter, and had emerged at the cortical surface of the TBI site on the contralateral cerebral hemisphere, apparently following axons along the corpus callosum. In sham-injured control animals, the topically applied GFP-MSCs proliferated superficially on the cortex at the site of application, and no GFP-MSCs were found at the contralateral cortical surface. In all instances, GFP-MSCs were not detected in other organs of either the test or the control animals. Our study demonstrated that MSCs topically applied to the brain surface can migrate to a TBI site.