PROJECT SUMMARY Ischemic stroke is a significant public health concern in the United States. Current therapeuticapproaches for stroke involve thrombolytic therapy with recombinant tissue plasminogen activator (r-tPA) orendovascular treatments; however many patients still experience disability. The goal of improving post-strokeoutcomes requires novel neuroprotective drugs for stroke treatment. While many such compounds have beenidentified in preclinical stroke studies none of these have been successfully translated to the clinic. In contrast3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitors (i.e. statins) are routinely given tostroke patients due to an inherent ability of these drugs to improve post-stroke functional outcomes. In this grant we will test the mechanistic hypothesis that neuroprotection from statins resultsfrom transport across the blood-brain barrier (BBB) that is mediated by the critical uptake transporterorganic anion transporting polypeptide 1a4 (Oatp1a4). This hypothesis will be tested by two aims. Aim 1: To investigate CNS delivery of statins mediated by Oatp1a4 in ischemic stroke. To evaluatethe role of Oatp1a4 in CNS drug delivery during stroke we will investigate Oatp1a4-mediated statin transport atthe BBB using the transient middle cerebral artery occlusion (tMCAO) model. In these studies age-matchedmale and female rats will be subjected to tMCAO for 90 min (Aim 1A). We will then demonstrate that Oatp1a4-mediated statin delivery improves both biomarkers of neuroprotection and BBB integrity (Aim 1B). We will alsoperform neurocognitive sensorimotor and motor performance studies (i.e. functional neurological tests) inanimals administered statins and subjected to tMCAO with reperfusion times of up to 21 days (Aim 1C). In allstudies statins will be administered intravenously either at the time of reperfusion or after 2 h of reperfusion toshow that early administration of neuroprotective drugs can improve post-stroke outcomes. Aim 2: Transforming Growth Factor-b (TGF-b) signaling can be targeted to control Oatp1a4-mediated CNS statin delivery in ischemic stroke. In these experiments we will perform dose-responsestudies and multiple-dosing experiments in age-matched male and female Sprague-Dawley rats using the TGF-b/ALK1 agonist bone morphogenetic protein (BMP)-9 and the TGF-b/ALK5 antagonist SB431542. We will studythe activation of specific Smad proteins that control TGF-b signaling in brain microvascular endothelial cells (Aim2A). We will also determine the time course of Oatp1a4 expression and activity changes following BMP-9 orSB431542 treatment and their effects on CNS delivery of statins (Aim 2B). Additionally we will measure indicesof neuroprotection markers of BBB protection and neurological outcomes in rats subjected to tMCAOadministered BMP-9 or SB431542 and injected intravenously with a statin (Aim 2C). Overall these studies are clinically relevant because they will demonstrate the effective BBBtransport mechanisms are required to confer effectiveness of neuroprotective drugs in stroke.