Objective NB-3 is a member of the F3/contactin family of neural recognition molecules, which are crucial for cell morphogenesis and motility.NB-3 is expressed in neurons and plays an important role in axonal extension and neuronal survival.However, the role of NB-3 in cerebral ischemic injury remains unknown.Methods Adult male wild-type and NB-3 knockout (KO) mice were subjected to ischemic injury by unilateral middle cerebral carotid artery occlusion for 3 h, 6 h and 12 h.Ischemic infarction volumes were then determined by 2, 3, 5-triphenyltetrazolium chloride staining.Neurological dysfunction analysis was also performed.Primary culture of neuronal cells from wild-type and knockout animals were also used for analysis of neuronal survival and neurite outgrowth.Results NB-3 expression in the ischemic hemisphere was decreased after transient middle cerebral artery occlusion (MCAO).NB-3-KO mice developed a 2.6-fold larger infarct volume and exhibited increased neurological deficit scores after transient MCAO compared with control mice.Substrate with NB-3 promoted neuronal survival and neurite outgrowth in vitro, while neurite outgrowth and neuronal survival were significantly reduced in NB-3-deficient neurons.In addition, NB-3 deficiency renders neurons more susceptible to OGD-induced damage and NB-3 as substrate could partially via homophilic mechanisms.Conclusions These data demonstrate that NB-3 deficiency may aggravate brain damage after MCAO by impairing neuronal survival and neurite growth.