During stroke, blood-brain barrier (BBB) permeability increases and cannabidiol (CBD) has shown protection in animal stroke models. The aim of this study was to establish if CBD modulates BBB permeability following oxygen-glucose deprivation (OGD). To model the BBB co-cultures of human brain microvascular endothelial cells (HBMECs) and human astrocytes were grown on inserts. BBB permeability was measured by transepithelial electrical resistance (TEER) using STX2 electrodes and an EVOM2 meter. Inserts were subjected to 4h OGD by incubation in GasPak pouches with glucose-free RPMI medium. Reperfusion was established by returning cells to normoxia and their normal medium; TEER was measured at various time-points over 28h. CBD, receptor antagonists, or vehicle were added pre or post-OGD. The effect of giving 10µM CBD 2 or 4h into reperfusion was also investigated. Lactate dehydrogenase (LDH) was measured using a kit and VCAM-1 was measured by ELISA. Levels of cytokines/chemokine’s known to be altered in stroke were measured by Multiplex. HBMEC-monocultures were also grown on 6 well-plates, and IL-6 and VCAM-1 were measured by ELISA, both in the presence and absence of OGD. Statistics: Student’s t-test or one-way ANOVA with Dunnett’s. Exposing the BBB to 4h OGD increased its permeability (P<0.001). Pre-treatment with 10μM CBD attenuated this increase (P<0.05), and 100nM, 1 or 10µM CBD decreased barrier permeability during reperfusion (P<0.001-0.05, n=8-11). CBD given post-OGD decreased barrier permeability during reperfusion (P<0.01-0.05, n=8-16). Receptor antagonists for CB1, CB2, PPARα, adenosine-A2A, 5-HT1A and TRPV1 did not affect CBD responses (n=6-10). However, the effect of CBD was inhibited by PPARγ antagonism (P<0.001-0.05, n=6). Administration of the PPARγ agonist Pioglitazone post-OGD also decreased barrier resistance during reperfusion (P<0.01-0.05, n=6). CBD given 2h into reperfusion decreased permeability (P<0.05, n=8-11), but CBD given 4h into reperfusion did not (n=6-9). LDH was raised following 4h OGD and 28h reperfusion (P<0.05), this was inhibited by CBD (P<0.05) in a non-PPARγ manner (n=6-9). Detectable levels of IL-10, IL-6, MIP-1α, MIP-1β and VEGF did not differ with CBD treatment. However, VCAM-1 at the end of reperfusion was attenuated by CBD (P<0.05, n=6), which was insensitive to PPARγ-antagonism. VCAM-1 in medium from HBMEC-monoculture was also attenuated by CBD both in the presence (P<0.05, n=3) or absence (P<0.01, n=3) of OGD. IL-6 from HBMEC-monoculture medium was increased by OGD exposure (P<0.001, n=6), and this increase was even greater in CBD-treated cells (P<0.001, n=6). Furthermore, CBD increased IL-6 in a concentration-dependent manner in HBMEC-monocultures that had not been exposed to OGD (P<0.05, n=3). In conclusion, CBD given pre- or post-OGD decreases barrier permeability.