Background The transient global cerebral hypoperfusion/reperfusion achieved by induction of Bilateral Common Carotid Artery Occlusion followed by Reperfusion (BCCAO/R) may trigger a physiological response in an attempt to preserve tissue and function integrity. to evaluate whether the ECS the enzyme cyclooxygenase-2 (COX-2) and PPAR-alpha are involved during BCCAO/R in rat brain and to identify possible markers of the ongoing BCCAO/R-induced challenge in plasma. Methods Adult Wistar rats underwent BCCAO/R with 30?min hypoperfusion followed by 60?min reperfusion. The frontal and temporal-occipital cortices and plasma were analyzed by high performance liquid chromatography-mass spectrometry (HPLC-MS) to determine concentrations of endocannabinoids (eCBs) and related molecules behaving as ligands of PPAR-alpha and of oxidative-stress markers such as lipoperoxides while Western Blot Istradefylline and immunohistochemistry were used to study protein expression of cannabinoid receptors COX-2 and PPAR-alpha. Unpaired Student’s in (a) and (c … Fig. 5 Double immunofluorescence Istradefylline for cyclooxygenase-2 (COX-2) (a b e f) and either Iba1 (c e) or glial fibrillary acidic protein (GFAP) (d f) in the frontal cortex of BCCAO/R rats. Scale bars E?=?A C: 25?μm; F?=?B … Statistical analysis Data from the two experimental groups sham-operated animals and the BCCAO/R ones are depicted in the figures as mean ± standard deviation (S.D.) and statistical differences were determined by unpaired Student’s does not increase susceptibility to oxidative stress. Data in rodents and healthy humans support this inference as dietary supplementation with n-3 PUFAs does not affect lipid peroxidation [50 51 By contrast since DHA has been shown to have a role in neuroprotection after brain hypoxia and ischemia it is relevant that recent prophylactic and therapeutic approaches for cerebrovascular disease take into account the pathways of brain accretion and delivery of DHA [48 52 Istradefylline In our study eCB changes were further characterized by a marked increase of tissue and plasmatic concentrations of lipoperoxides that is one of the hallmarks of the reperfusion-induced oxidative stress [53 54 Lipoperoxides are quite unstable compounds that are capable of extending?the free radical oxidative damage and forming proinflammatory substances [53 54 The increase in lipoperoxide levels without any evident histological alteration of cerebral tissue is in line with previous experimental findings on a rat model of BCCAO/R similar to ours . Clinical implications Increase of levels of lipoperoxides and a concomitant increase of their catabolism in peroxisomes have been shown to be directly correlated with the hypoperfusion/reperfusion-induced oxidative challenge in patients undergoing carotid endoarterectomy . In particular it is interesting that peroxisomal beta-oxidation increased during the first 30?min of reperfusion only in patients having contralateral carotid stenosis higher Istradefylline than Istradefylline 50% to decrease thereafter within 2?h from reperfusion . Several physiopathological conditions not necessarily associated with early obvious neurological signs  share the occurrence of cerebral hypoperfusion episodes for which the detection of molecular indicators in the early hours may be useful in FCRL5 clinical settings to prevent irreversible cerebral damage. The question of whether plasmatic changes of AEA and lipoperoxides could represent additional specific markers in humans should be further investigated. Conclusions The present study showed that the 30/60?min BCCAO/R procedure activates the ECS in rat and induces parallel changes in the fatty acid tissue profile (namely decreased levels of DHA and increased the lipoperoxides) and COX-2 levels in the rat frontal cortex. In addition we found that BCCAO/R increased plasmatic levels of anandamide and lipoperoxides. The molecular changes induced by the BCCAO/R are evaluated on the basis of a single time point Istradefylline of reperfusion and so far this aspect represents an intrinsic limitation. Additional studies are warranted to evaluate both the time course of these changes during longer time points of reperfusion (e.g. at 6 12 and 24?h after BCCAO/R) and the possible effects of dietary compounds in preventing BCCAO/R-induced oxidative stress. In conclusion this study shows that BCCAO/R-induced positive modulation of the ECS. As far as we aware this is the first study that has investigated early changes that can be easily traced in brain tissue as well as in plasma and may be interpreted as indicative of the tissue physiological response to the oxidative stress induced by the BCCAO/R. The variations observed suggest that the activation of the ECS and.