Flow-induced cerebral vasodilatation involves activation of PI3-kinase and production of reactive oxygen species

Reactive oxygen species (ROS) such as superoxide and hydrogen peroxide are known to be cerebral vasodilators. A major source of ROS in the vasculature is the flavin-containing enzyme NADPH-oxidase. We have previously demonstrated that activation of NADPH-oxidase leads to dilatation of the basilar artery in vivo, via a mechanism involving production of hydrogen peroxide from superoxide dismutase. The endogenous stimuli for this unique vasodilator mechanism are yet to be characterised, however shear stress is known to activate phosphatidylinositol 3-kinase (PI3-K) and NADPH-oxidase in cultured cells. This study investigated whether increased intraluminal blood flow could induce cerebral vasodilatation via the activation of PI3-K and/or NADPH-oxidase. A cranial window preparation in anaesthetised rats was used to examine vasomotor responses of the basilar artery in the absence and presence of inhibitors of ROS production and breakdown. Bilateral occlusion of the common carotid arteries to increase basilar artery flow resulted in reproducible dilatations of this vessel (77±2% of maximum, n=39) that were rapidly reversed when normal flow was restored. Flow-dependent dilatation was profoundly inhibited following treatment with the PI3-K inhibitor wortmannin (1 μM). Treatment of the basilar artery with the NADPH-oxidase inhibitor diphenyleneiodonium at 0.5 and 5 μM also caused a significant reduction in flow-dependent dilatation (54±9% and 37±7 % of maximum) without affecting nitric oxide-mediated dilatations to acetylcholine. Treatment with the hydrogen peroxide scavenger catalase also reduced flow-dependent dilatation from 79±7% to 55±7%, indicating a role for NADPH-oxidase-derived hydrogen peroxide in this response. The nitric oxide synthase (NOS) inhibitor L-NAME caused a small reduction (from 83±8% to 57±6%) in flow-dependent dilatation. Furthermore, combined treatment with a ROS inhibitor (diphenyleneiodonium or catalase) and L-NAME caused a greater reduction in flow-dependent dilatation than seen with inhibition of either pathway alone. Thus, flow-dependent cerebral vasodilatation in vivo involves production of both ROS and nitric oxide, and is dependent on PI3-K activation.