Recent research has shown that transplanted bone marrow stromal cells (MSCs) migrate to the injured regions and exert their therapeutic effects in cases of intracranial trauma, stroke, inflammation and degenerative disease. The specific mechanisms involved in their migration to lesions are still to be fully elucidated. In the present study, a rat model of transient middle cerebral artery occlusion (MCAO) was established. At 24 h after reperfusion, human bone marrow stromal cells (hMSCs) were transplanted by intravenous injection to explore the effects of fractalkine/CX3CR1 on the migration of transplanted MSCs to lesions. In vitro study using real-time PCR and western blot revealed that CX3CR1, the only known receptor of fractalkine, was expressed in cultured hMSCs. The expression of fractalkine in the ischemic brain was significantly increased. The directional migration of transplanted hMSCs to the damaged region was observed through detection of green fluorescence protein (GFP). The results indicated the cells were mainly distributed in the ischemic boundary zone with high fractalkine expression. In a further study, lentivirus-mediated RNA interference of CX3CR1 expression was employed. The results of these experiments indicated that CX3CR1 knock-down dramatically decreased the migration of hMSCs to the ischemic brain. The present study suggests that fractalkine and its specific receptor CX3CR1 are involved in the directional migration of transplanted MSCs to the ischemic damaged brain region.