Prostaglandin FP receptor inhibitor reduces ischemic brain damage and neurotoxicity
Highlights
► Infarct volumes and behavioral deficits were reduced in FP antagonist AL8810 mice. ► Similar results in FP knockout mice after permanent ischemic stroke. ► FP receptor blockade protects against oxygen glucose deprivation cell death in organotypic slices. ► Neuronal death is less in cells treated with AL8810 not in cultured astrocytes. ► FP receptor agonist increases intracellular calcium levels of cultured neurons.
Introduction
We and others have described that the bioactive lipids, such as the prostaglandins, might affect neuronal outcomes after a variety of brain insults, including hypoxia, inflammation, and excitotoxicity (Dore et al., 2003). Therefore, we have focused our research on the unique properties of the prostaglandins and their receptors (Doré, 2006). Cyclooxgenases are the rate limiting enzymes for the conversion of arachidonic acid to prostaglandin PGH2, which is metabolized by specific prostaglandin synthases to at least five structurally related bioactive lipid molecules, including PGE2, PGD2, PGF2α, PGI2, and thromboxane A2 (Doré, 2006). PGF2α, which is synthesized from PGH2 via PGF synthase, plays a major role in initiation of parturition, renal function, control of cerebral blood flow autoregulation in newborn piglets (Chemtob et al., 1990), contraction of arteries, and myocardial dysfunction (Takayama et al., 2005). Recently, a report suggested that FP signaling can facilitate idiopathic pulmonary fibrosis (Oga et al., 2009). Although the critical biological functions of PGF2α were reported to be mediated through activation of the FP receptor, the effect and mechanism of FP activation are not well understood in ischemic stroke, especially in regard to stroke-related Ca2+ signaling. Based on reports of the Prostaglandin F (FP) receptor's presence in the central nervous system and its physiologic properties in regard to stroke and Ca2+ signaling (Abramovitz et al., 1994, Kitanaka et al., 1994, Saleem et al., 2009a), we hypothesize that activation of the FP receptor subsequent to injury contributes to excitotoxic and ischemic damage. Therefore, our goal is to investigate the role of the FP receptor in the brain and in neuronal cells and to further address relevant signaling pathways.
We have established the importance of the PGE2 EP1 receptor in ischemic models (Ahmad et al., 2008). Based on its similarity to the EP1 receptor in terms of amino acid sequence and cell signaling, we hypothesize that the FP receptor might contribute to excitotoxic and ischemic brain damage similar to that caused by the EP1 receptor. We speculate that the mechanism of injury might stem from dysregulation of Ca2+ homeostasis. Therefore, we studied the role of the FP receptor in a mouse model of permanent middle cerebral artery occlusion (pMCAO) and in ischemia-related Ca2+ signaling in cultured neurons.
Section snippets
Animals
All animal protocols were approved by the Institutional Animal Care and Use Committee. All mice were maintained and housed in the vivarium under controlled conditions (23 ± 2°C; 12 h light/dark periods) with access to food and water ad libitum. Adult male WT mice and FP−/− C57BL/6 mice were used at 8–10 weeks of age (20–25 g) in this study.
pMCAO, neurologic function, and infarct size determination
The pMCAO procedure was carried out as previously described (Saleem et al., 2009c). Mice were anesthetized with 250 mg/kg Avertin (Sigma), administered
Blockade of FP receptor is protective in a stroke mouse model
To determine the effect of FP receptor inhibition in stroke, we injected mice i.v. with 1 or 10 mg/kg of the FP antagonist AL-8810 (Cayman Chemical) immediately after pMCAO. The cortical infarct volume was significantly less in the AL-8810-treated mice than in the vehicle-treated group 48 h after pMCAO (Figs. 1A, B; P < 0.05 [1 mg/kg], P < 0.001 [10 mg/kg]). Mice treated with AL-8810 also had significantly reduced tape-removal times compared with mice in the vehicle-treated group at 48 h (Fig. 1C).
Discussion
In our previous study, the FP receptor significantly reduced infarct volume in a transient MCAO mouse model (Saleem et al., 2009a). In this study, we showed that the inhibitor of the FP receptor significantly improved the outcome in mice after ischemia in terms of neurobehavioral function and infarct volume. In a previous study, OGD resulted in expression of apoptotic and necrotic cell death phenotypes, especially in neurons (Newcomb-Fernandez et al., 2001). We also demonstrated that FP
Conclusion
In this study, we used AL-8810 to evaluate the role of FP receptor in ischemic damage. AL-8810 reduced OGD-induced neuronal cell death in slice cultures from WT mice but had no effect on cell death or ROS production in slices from FP−/− mice, indicating that the drug is selective for FP. However, potent and selective FP antagonists should be developed for more pharmacologic studies and therapeutic applications. Our findings provide novel targets for intervention and treatment of cerebral
Disclosures
None.
Acknowledgments
The authors thank all Doré lab members for their insightful contributions for this manuscript. This research was supported by grants from the National Institutes of Health NS046400 and AG022971 (SD), a fellowship from the Korea Research Foundation, KRF-2007-357-E00016, and by a postdoctoral grant from the American Heart Association, 09POST2060557 (YTK).
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These authors contributed equally to the work.