Inhibition of occludin S490 phosphorylation prevents MCAO-induced permeability. (A) PDGFiCre⁺ mice and PDGFiCre⁺; S490AOCC^+/+ mice were subjected to MCAO, and BBB permeability was determined 24 hours later by quantifying a 70-kDa fluorescent dextran leak in cross-sections from both the ipsilateral (Ipsi) and the contralateral (Contra) region. (B) WT mice were given vehicle (Veh) or the PKCβ inhibitor (PKCβi; 10 mg/kg) once a day for 3 days and then subjected to MCAO followed by one additional dose of the inhibitor 1 hour later. pS490 occludin (red) and total occludin (green) were detected by immunostaining in brain sections 24 hours after MCAO. Scale bar, 25 µm. Quantification of pS490 at the junction (C) and total occludin from confocal images (D) 24 hours after MCAO. (E) Image of BBB permeability to 70-kDa dextran in entire coronal sections in both contralateral and ipsilateral hemispheres 24 hours after MCAO. Scale bar, 600 µm. (F) Quantification of dextran leak in cross-sections 24 hours after MCAO. (G) BBB permeability to 70-kDa dextran was also assessed by quantifying the amount of extravasated dye in brain homogenates 24 hours after MCAO. One-way analysis of variance followed by a Holm-Šídák post hoc test. *P < .05, ****P < .0001. ns, nonsignificant.