The present study was designed to explore the mechanisms involved in the anti-ischemic action of lumbrokinase (LK) in brain. The enzyme immunoassay, spectrofluorimeter and flow cytometry were used to detect the level of adenosine 3',5'-cyclic monophosphate (cAMP) and guanosine 3',5'-cyclic monophosphate (cGMP), the Ca(2+) mobilization, and human platelet surface antigen expression in order to elucidate the anti-platelet action involved in LK cerebroprotection. RT-PCR and western blot were used to identify the role of Intercellular adhesion molecule-1 (ICAM-1) and Janus Kinase1/Signal Transducers and Activators of Transcription1 (JAK1/STAT1) pathway in protecting brain against ischemic injury by anti-thrombosis and anti-apoptosis. Results showed that LK significantly potentiated the activity of adenylate cyclase (AC), increased the cAMP level in vivo, remarkably inhibited the rise of rat platelet intracellular Ca(2+) ([Ca(2+)](i)), and attenuated the expression of Glycoprotein IIB/IIIA (GPIIB/IIIA) and P-selectin in human platelet stimulated by thrombin in vitro. Furthermore, the expressions of ICAM-1 and JAK1/STAT1 were remarkably regulated by LK in Human Umbilical Vein Endothelial Cell (HUVEC) and ischemic cerebral tissues. These data indicated that the anti-ischemic activity of LK was due to its anti-platelet activity by elevating cAMP level and attenuating the calcium release from calcium stores, the anti-thrombosis action due to inhibiting of ICAM-1 expression, and the anti-apoptotic effect due to the activation of JAK1/STAT1 pathway.