Glycogen synthase kinase-3β (GSK-3β),a serine/threonine protein kinase,is widely distributed in mammalian brains.Since GSK-3β plays a vital role in the development of neurodegenerative disorders,the present study was designed to investigate the role of GSK-3β in the blood-brain barrier (BBB) permeability in aged mice.Morris water maze test was used to examine mouse cognitive function.BBB permeability was examined by the leakage of fluorescence signals of low-molecular weight dextran.GSK-3β inhibitor,4-benzyl-2-methyl-1,2,4-thiadiazolidine-3,5-dione (TDZD-8),was administrated in aged mice and in cultured mouse brain microvascular endothelial cells (bEnd.3).Compared with young mice,aged mice had increased leftover signals of dextran in the hippocampus and a lower score in the maze test,suggesting that aged mice have abnormal leakage of BBB and cognitive dysfunction.The protein expression of Toll-like receptor 4 (TLR4) was increased,whereas the protein expressions of junction proteins (claudin1 and claudin5) were reduced in endothelial cells of BBB in aged mice.Phosphorylated level of serine 9,an inhibitory residue in GSK-3β protein,was decreased.TDZD-8 treatment downregulated TLR4 protein expression,upregulated claudin1 and claudin5 protein expressions,and significantly improved cognitive function in aged mice.In bEnd.3 cells,TDZD-8 treatment reduced TLR4 expression and increased claudin5 expression in cells stimulated with lipopolysaccharides.In conclusion,the inhibition of GSK-3β activity downregulates aging-induced TLR4 expression and restores the BBB integrity,resulting in the improvement of cognitive function in aged mice.