Glibenclamide ameliorates ischemia–reperfusion injury via modulating oxidative stress and inflammatory mediators in the rat hippocampus

BR A I N R ES E A RC H 1 3 8 5 ( 2 01 1 ) 2 5 7 –2 62

available at www.sciencedirect.com

www.elsevier.com/locate/brainres

Research Report

Glibenclamide ameliorates ischemia–reperfusion injury via modulating oxidative stress and inflammatory mediators in the rat hippocampus Dalaal M. Abdallah, Noha N. Nassar⁎, Rania M. Abd-El-Salam Department of Pharmacology and Toxicology, Faculty of Pharmacy, Cairo University, Cairo, Egypt

A R T I C LE I N FO

AB S T R A C T

Article history:

Stroke remains a debilitating disease with high incidence of morbidity and mortality, where

Accepted 3 February 2011

many reports provide promising venues for prevention/treatment of such ailment.

Available online 5 March 2011

Glibenclamide, a selective blocker of KATP channels, was reported to protect against ischemia and ischemia–reperfusion (IR) injury in several experimental models. Hence, the

Keywords:

present study aimed to investigate the possible involvement of free radicals as well as

Glibenclamide

inflammatory and anti-inflammatory mediators in the hippocampus of rats exposed to IR. To

Ischemia–reperfusion

this end, male Wistar rats were divided into 3 groups: group I served as sham operated controls;

Oxidative stress

group II was subjected to 15 min ischemia by occlusion of both common carotid arteries,

Nitric oxide

followed by 60 min reperfusion; group III was injected with glibenclamide (1 mg/kg, i.p.) 10 min

Cytokines

before ischemic–reperfusion injury. IR increased lipid peroxides, myeloperoxidase activity,

Prostaglandin E2

TNF-α and PGE2, while decreasing glutathione, total antioxidant capacity, nitric oxide and IL-10 levels in the hippocampus. Glibenclamide reversed all the former alterations, thus highlighting a potential therapeutic utility for this sulphonyl urea in IR brain injury via modulating oxidative stress and inflammatory mediators. © 2011 Elsevier B.V. All rights reserved.

1.

Introduction

Despite advances in cellular and molecular mechanisms, stroke remains the second leading debilitating disease worldwide (Bakhai, 2004). Recent understanding of the pathophysiology of this ailment implicates multiple cell types and pathways including excitotoxicity, apoptosis and inflammation (Bielewicz et al., 2010). In the brain, ischemia for more than

5 min leads to considerable neuronal death and infarction (Deb et al., 2010; Krnjevic, 2008). Early reperfusion injury results in generation of reactive oxygen species (ROS) via activation of macrophages (Tapuria et al., 2008). This event further jeopardizes endothelial injury and augments recruitment of neutrophils with subsequent release of proinflammatory cytokines, prime contributors in the late phase of ischemia– reperfusion (IR) injury (Lakhan et al., 2009; Tapuria et al., 2008).

⁎ Corresponding author at: Department of Pharmacology and Toxicology, Faculty of Pharmacy, Cairo University, Kasr El-Aini Street, Cairo 11562, Egypt. Fax: +20 22 484 1841. E-mail addresses: [email protected], [email protected] (N.N. Nassar). Abbreviations: ATP, adenosine triphosphate; GSH, glutathione; IL-10, interleukin10; IR, ischemia–reperfusion; MPO, myeloperoxidase; NC(Ca-ATP) channel, Ca2+-activated [ATP]i-sensitive nonspecific cation channel; PGE2, prostaglandin E2; ROS, reactive oxygen species; SUR-1, sulphonyl urea receptor-1; TAC, total antioxidant capacity; TBARS, thiobarbituric acid reactive substances; TNF-α, tumor necrosis factor-alpha 0006-8993/$ – see front matter © 2011 Elsevier B.V. All rights reserved. doi:10.1016/j.brainres.2011.02.007

258

BR A I N R ES E A RC H 1 3 8 5 ( 2 01 1 ) 2 5 7 –26 2

Among the various factors involved in molecular mechanisms of stroke, special interest is directed towards ATP depletion, a crucial player in subsequent cytotoxicity (Taoufik and Probert, 2008). Notably, this event results in the upregulation of the non-constitutive SUR1-regulated NC(Ca-ATP) during hypoxic insult in astrocytes, neurons and endothelial cells (Simard et al., 2006). Opening of these channels creates a strong inward current, cell depolarization and cytotoxic edema that eventually results in cell death (Krnjevic, 2008; Simard et al., 2006). Moreover, another contributor is the KATP channel, which is usually closed under normal conditions and rapidly activated in response to an increase of intracellular ADP and hypoxia conditions causing K+ efflux; hence, they directly couple the metabolic cellular state to its electrical activity (Sun and Hu, 2010). These channels are abundantly distributed throughout the brain, being found in neurons, glial cells and in the brain vasculature (Sun and Hu, 2010), which have been shown to elicit controversial effects in ischemic insult. Although a plethora of literature supports a beneficial effect for opening of KATP channels in IR injury (Sun and Hu, 2010; Zarch et al., 2009), others advocate a protective role for glibenclamide, either by blocking KATP channel (Nistico et al., 2007) and/or SUR-1 in multiple ischemic models (Simard et al., 2006 and 2009). Moreover, glibenclamide has been shown to ameliorate damage caused by renal and intestinal IR (Pompermayer et al., 2005 and 2007). Via its KATP channel blockade, glibenclamide suppresses neutrophil migration and chemotaxis in acute inflammatory conditions (Da SilvaSantos et al., 2002; Pompermayer et al., 2007). However, independent of its KATP blocking activity, this sulphonyl urea demonstrates direct antioxidant potential (Al-Azzam et al., 2010; Nazaroglu et al., 2009), which makes it a suitable candidate for stalling neuronal damage induced by IR insult. Accordingly, in the current study, we hypothesized that glibenclamide, the KATP channel blocker, by virtue of its antioxidant/anti-inflammatory properties may ameliorate damage induced by IR injury in the hippocampus.

Table 1 – Effect of IR alone or following glibenclamide pretreatment on basal blood glucose level Blood glucose (mmol/L) Groups

Baseline

10 min preischemia

15 min postischemia

60 min reperfusion

Control IR Gliben (1 mg/kg, i.p.)

5.2 ± 0.35 5.3 ± 0.56 4.8 ± 0.08

5.2 ± 0.42 4.8 ± 0.62 4.9 ± 0.24

5.4 ± 0.35 4.8 ± 0.61 4.6 ± 0.12

5.4 ± 0.31 3.8 ± 0.34 ⁎ 4.2 ± 0.19

Values are means ± S.E.M (n = 8, each group). ⁎ Significantly different from baseline p < 0.05 using two-way ANOVA followed by Bonferroni post hoc test.

(118%), TAC (182%) as well as NO production (150%) following IR in hippocampal homogenate.

2.3. Effect of glibenclamide on inflammatory mediators and cytokines induced by IR Following reperfusion, there was a marked activation of myeloperoxidase (MPO; approximately 2.5-fold above control value), indicative of increased neutrophil infiltration (Fig. 2a). This finding corroborated with the increased production of the proinflammatory cytokine tumor necrosis factor-α (TNF-α; 119%; Fig. 2b) that coincided with a prominent decline in the anti-inflammatory cytokine interleukin-10 (IL-10; 60%; Fig. 2c). Furthermore, the prostanoid prostaglandin E2 (PGE2; 151%; Fig.2d) was elevated by the insult. On the other hand, glibenclamide ameliorated neutrophil infiltration by 66% as well as both inflammatory markers, TNF-α (85%) and PGE2 (84%). Moreover, the sulphonyl urea boosted IL-10 level to 134% above that induced by IR.

3. 2.

Results

2.1.

Effect of IR and glibenclamide on blood glucose level

The baseline values for blood glucose were similar in all groups used. Blood glucose levels remained insignificant from baseline in all groups over the first 25 min (Table 1). However, following reperfusion, the IR group showed a significant hypoglycemic effect compared to baseline. Interestingly, the group receiving glibenclamide showed insignificant effect from baseline values and IR (Table 1).

2.2. by IR

Effect of glibenclamide on altered redox status induced

IR induced ROS as evidenced by increased thiobarbituric acid derivatives (Fig. 1a) by 168% from its respective vehicle control, which was substantiated by a decline in glutathione (GSH; 72%; Fig. 1b), total antioxidant capacity (TAC; 46%; Fig. 1c) as well as nitric oxide (NO) production (68%; Fig. 1d). Glibenclamide reduced lipid peroxide level (80%) and increased GSH

Discussion

Despite advances in discerning the etiology of brain ischemic stroke, therapeutic options remain constrained. Increasing evidence shows that ischemic injury due to KATP channel activation (Sun and Hu, 2010) and the consequent ensuing inflammation/oxidant status disturbance that irreversibly damages neurons (Deb et al., 2010; Krnjevic, 2008; Lakhan et al., 2009). Furthermore, reperfusion renders the brain more vulnerable to neuronal injury (Lakhan et al., 2009). In the current study, glibenclamide, the KATP channel blocker, conferred neuropotection against IR as evidenced by: (i) reducing neutrophil recruitment; (ii) restoring proxidant/antioxidant balance; (iii) ameliorating inflammatory mediators; (iv) boosting the anti-inflammatory cytokine IL-10 and (v) attenuating reperfusion induced hypoglycemia. Though several reports supported the view that glibenclamide antagonizes the preconditioning effects of various pharmacological agents (Ma et al., 2004; Rehni et al., 2007; Velly et al., 2009), the current study documents its neuroprotective efficacy in a global IR model. The present results are consistent with those of Smirad et al. (2006 and 2009) in different ischemic

BR A I N R ES E A RC H 1 3 8 5 ( 2 01 1 ) 2 5 7 –2 62

259

Fig. 1 – Effect of IR alone or with glibenclamide (Gliben; 1 mg/kg, i.p.) administered 10 min prior to ischemia on thiobarbituric reactive substances (TBARS; a), glutathione (GSH; b), total antioxidant capacity (TAC; c) and nitric oxide (NO; d). Data represent the mean of 8 experiments ± S.E.M.; *, #p < 0.05 compared to control and IR group, respectively using one-way ANOVA followed by Student–Newman–Keuls multiple comparisons test.

models in the brain as well as other studies in combating IR injury in kidney and intestinal tissue (Pompermayer et al., 2005 and 2007). Evidence exists that KATP channels are located on the inner mitochondrial membrane and that their activation in the early stage of ischemic insult is due to either hypoxia, glucose deprivation or both (Busija et al., 2008; Krnjevic, 2008).

Consequently, Ca2+ overload in the mitochondria via opening of KCa2+ channels is triggered by a variety of factors, including Ca2+ release from internal stores, acidosis and extrinsic agents, notably glutamate and adenosine (Krnjevic, 2008). This translocation of Ca2+ activates cellular proteases and lipases with the subsequent tissue breakdown (Krnjevic, 2008) that ultimately results in lipid peroxidation and cellular death. Another

Fig. 2 – Effect of IR alone or with glibenclamide (Gliben; 1 mg/kg, i.p.) administered 10 min prior to ischemia on myeloperoxidase (MPO; a) concentration, tumor necrosis factor-alpha (TNF-α; b), interleukin-10 (IL-10, c) and prostaglandin E2 (PG E2, d). Data represent the mean of 8 experiments ± S.E.M.; *, #p < 0.05 compared to control and IR group, respectively using one-way ANOVA followed by Student–Newman–Keuls multiple comparisons test.

260

BR A I N R ES E A RC H 1 3 8 5 ( 2 01 1 ) 2 5 7 –26 2

contributor of propagated destruction of cell membranes is the formation of ROS (Deb et al., 2010; Lakhan et al., 2009). The current investigation highlights elevation of neutrophil infiltration and lipid peroxides, consistent with other reports (Anaya-Prado et al., 2008; Elango et al., 2009), along with a decreased NO content. Both former effects were reflected on the antioxidant defense systems, where GSH and TAC concentrations were suppressed; these findings are in line with previous ones (Elango et al., 2009; Gaur and Kumar, 2010). By virtue of its KATP channel blocking potential (Nistico et al., 2007) and antioxidant activity (Al-Azzam et al., 2010; Nazaroglu et al., 2009), glibenclamide might just hamper those events, thus favoring neuronal salvaging. Similarly, the sulphonyl urea, via blockade of KATP channels, suppresses neutrophil migration and chemotaxis and indirectly reduces the proinflammatory cytokine TNF-α released by these cells (Da SilvaSantos et al., 2002; Pompermayer et al., 2005 and 2007). In fact, a lower level of TNF-α associated by increased production of the anti-inflammatory cytokine IL-10 was documented by the sulphonyl urea treatment in this study. Interestingly, in the current investigation we report a hypoglycemic effect in the IR group, which is in line with a recent clinical report (Naidech et al., 2010). A plausible explanation for this observation might be attributed to the increased level of TNFα, where a previous study indicates that the increase in the this cytokine might induce hypoglycemia (Battelino et al., 1996). Hence, glibenclamide by reducing TNF-α might hamper IR induced hypoglycemic effect. During ischemia, NO production via nNOS is upregulated along with ROS production (Leker and Shohami, 2002). Excess NO in the vicinity of the increased levels of superoxide anion produces peroxynitrite (Alonso et al., 2002), which further exacerbates cellular injury (Leker and Shohami, 2002) and depletes thiol containing proteins, such as GSH, the body's most powerful endogenous antioxidant (Radi et al., 1991). Moreover, a previous study suggests that peroxynitrite induces opening of brain mitoKATP channels (Lacza et al., 2003). In the present study, NO concentration was reduced following IR insult, an effect that could be attributed to its exhaustion in formation of peroxynitrite (Alonso et al., 2002). The former finding is further supported by the enhanced production of thiobarbituric acid reactive substances (TBARS) and reduced levels of antioxidants in this study. Conversely, glibenclamide by prohibiting KATP activation and by virtue of its antioxidant potential could ameliorate damage caused by IR. In the present investigation, we report increased concentration of PGE2 in the hippocampi of IR rats, which corroborates with enhanced ROS formation, consistent with a previous report (Candelario-Jalil et al., 2003). Cyclooxygenase-2 (COX-2) is a crucial enzyme in several interlocking metabolic pathways, which mediates the transformation of structural membrane lipids into a plethora of biologically active eicosanoids including the inflammatory mediator PGE2 (Strauss, 2008). Early on after moderate brain injury, neuronal COX-2 levels are upregulated, especially in the hippocampus (Nakayama et al., 1998). Meanwhile, elevated intracellular free Ca2+ activates phospholipases that release arachidonic acid from membranes (Strauss, 2008). In turn, arachidonic acid produces ROS during its conversion to eicosanoids, hence boosting free radical formation that triggers

a central inflammatory reaction (Adibhatla and Hatcher, 2003). COX-2 via its peroxidative power oxidizes GSH during PGE2 formation (Hamberg et al., 1974), an effect that might afford another explanation to its decreased levels in the current investigation. On the other hand, glibenclamide effectively normalized PGE2 level, beside hippocampal redox status. The former finding is in line with a previously reported study (Bouchard et al., 1994). The present investigation highlights a potential therapeutic utility for glibenclamide, the KATP channel blocker, in IR brain injury via modulating oxidative stress and inflammatory mediators.

4.

Experimental procedures

4.1.

Animals

Adult male albino Wistar rats (200–250 g) were used in the present experiment. Animals were kept under controlled environmental conditions at a constant temperature (23 ± 2 ºC), humidity (60 ± 10%) and a light/dark (12/12 h) cycle. Food and water were available ad libitum until the beginning of the experiment. Animal care and experimental protocol was approved by the Research Ethical Committee of Faculty of Pharmacy Cairo University (Cairo Egypt).

4.2.

Groups and treatments

Experimental procedures were subdivided into 3 subsets. In each subset, animals were randomly allocated into 3 groups (n = 8 per group). In all subsets, rats were subjected to 15 min ischemia followed by 60 min reperfusion to serve as IR group. Another group was pretreated with glibenclamide (SigmaAldrich, CA, USA; 1 mg/kg, i.p.) injection 10 min before IR performance (Ma et al., 2004). A last group was administered 1% Tween 80 i.p., the vehicle used to suspend the suphonylurea to serve as the sham operated control.

4.3.

Induction of global ischemia–reperfusion (IR) injury

Rats were anaesthetized with phenobarbital (50 mg/kg, i.p.) and midline ventral incision was made in neck to expose right and left common carotid arteries. After separating from surrounding tissues and vagus nerve, bilateral carotid artery occlusion was performed for 15 min to induce global ischemia, and then reperfusion was allowed for 60 min (Racay et al., 2009). Rectal temperature was maintained close to 37 °C by means of a warming plate and an overhead incandescent bulb. Sham operated control rats underwent the same procedure except of carotid occlusion.

4.4.

Blood and tissue collection

Blood droplets were collected from the tail vein and blood glucose concentration was determined with rapid blood glucose meter (Wang et al., 2010) at four different time points (baseline, which represented zero time; 10 min pre-ischemia; 15 min postischemia and finally 60 min following reperfusion). After IR/ sham operation, all animals were euthanized under deep ether

BR A I N R ES E A RC H 1 3 8 5 ( 2 01 1 ) 2 5 7 –2 62

anesthesia. Brains were removed immediately and both hippocampi were dissected on ice-cold plates and homogenized immediately in 0.1 M phosphate (pH 7.4) buffer, containing 1 mM EDTA and 0.1 μM indomethacin for PGE2 measurement, hexadecyltrimethylammonium bromide (1%) in potassium phosphate buffer (100 mM, pH 6) for MPO activity or ice-cold saline for other determinations.

4.5.

Lipid peroxides determination

The thiobarbituric acid reaction of Mihara and Uchiyama (1978) was adopted for estimation of lipid peroxide level. To hippocampal homogenates orthophosphoric acid (1%) and thiobarbituric acid (0.6%) were added, and mixtures were boiled for 45 min at 100 °C, then cooled. The colored product was extracted by n-butanol, vortexed and centrifuged at 3000g for 15 min. The absorbance of the organic layer was read at 535 and 520 nm and the difference in absorbance was calculated as lipid peroxide level expressed as TBARS.

4.6.

Glutathione (GSH) estimation

The method for the assessment of GSH in the hippocampi was based on that of Beutler et al. (1963). Homogenates were deproteinated with 5-sulfuosalicylic acid (10%) for 30 min at 4 °C and then centrifuged at 3000g for 15 min at 4 °C. An aliquot of the acid soluble supernatant was diluted with phosphate buffer (0.3 M, pH 7.7) and 5,5′-dithiobis-2-nitrobenzoic acid (1 mM) was added to the samples, where its optical density was determined at 412 nm.

4.7.

Total antioxidant capacity (TCA) estimation

The method for the assessment of TAC of hippocampi was based on that of Koracevic et al. (2001) using commercial kit supplied by Biodiagnostic Co. (Giza, Egypt). Antioxidants present in the sample eliminate hydrogen peroxide (H2O2) and its residual level is determined by an enzymatic reaction at 505 nm.

4.8.

Nitric oxide (NO) estimation

Nitric oxide was assayed according to the method of Miranda et al. (2001), where hippocampal homogenates were deproteinated with absolute ethanol for 48 h at 4 °C, and then centrifuged at 12,000g for 15 min at 4 °C. To an aliquot of the supernatant vanadium trichloride (0.8% in 1 M HCl) was added for the reduction of nitrate to nitrite, followed by the rapid addition Griess reagent consisting of N-(1-naphthyl)ethylenediamine dihydrochloride (0.1%) and sulfanilamide (2% in 5% HCl), incubated for 30 min at 37 °C, and cooled where the absorbance was measured at 540 nm.

4.9.

Myeloperoxidase (MPO) activity

A slight modification of the method described by Krawisz et al. (1984) was used for the estimation of MPO(EC 1.15.1.1) activity (U/g tissue). Homogenates were subjected to 3 freeze and thaw cycles and sonicated for 10 s followed by centrifugation at 10,000g for 15 min at 4 °C. To the supernatant, a reaction

261

mixture consisted of o-dianisidine hydrochloride (0.167%) and H2O2 (0.0005%) in potassium phosphate buffer (50 mM, pH 6) was added. The change in absorbance was monitored at 1 min intervals at 460 nm for 4 min.

4.10.

TNF-α, IL-10 and PGE2 estimations

Hippocampal TNF-α, IL-10, and PGE2 were measured by ELISA kits purchased from Invitrogen (California, USA), Bender MedSystems (Vienna, Austria) and Cayman Chemical (MI, USA), respectively. All the procedures of the used kits were performed following manufacturers' instruction.

4.11.

Statistical analysis

Data are expressed as mean of eight experiments ± S.E.M., and statistical comparisons were carried out using one- or twoway analysis of variance (ANOVA) followed by Student– Newman–Keuls multiple or Bonferroni post hoc test, respectively. All analyses utilized GraphPad Prism 5.0 statistical package for Windows (La Jolla, CA, USA). The minimal level of significance was identified at p < 0.05.

REFERENCES

Adibhatla, R.M., Hatcher, J.F., 2003. Citicoline decreases phospholipase A2 stimulation and hydroxyl radical generation in transient cerebral ischemia. J. Neurosci. Res. 73, 308–315. Al-Azzam, S.I., Abdul-Razzak, K.K., Jaradat, M.W., 2010. The nephroprotective effects of pioglitazone and glibenclamide against gentamicin-induced nephrotoxicity in rats: a comparative study. J. Chemother. 22, 88–91. Alonso, D., Serrano, J., Rodriguez, I., Ruiz-Cabello, J., Fernandez, A.P., Encinas, J.M., Castro-Blanco, S., Bentura, M.L., Santacana, M., Richart, A., Fernandez-Vizarra, P., Uttenthal, L.O., Rodrigo, J., 2002. Effects of oxygen and glucose deprivation on the expression and distribution of neuronal and inducible nitric oxide synthases and on protein nitration in rat cerebral cortex. J. Comp. Neurol. 443, 183–200. Anaya-Prado, R., Perez-Gomez, N., Toledo-Pereyra, L.H., Walsh, J., Jordan, J., Ward, P.A., 2008. Small molecule selectin inhibitor in global cerebral ischemia and controlled hemorrhagic shock. J. Trauma 65, 678–684. Bakhai, A., 2004. The burden of coronary, cerebrovascular and peripheral arterial disease. Pharmacoeconomics 22 (Suppl 4), 11–18. Battelino, T., Goto, M., Zeller, W.P., 1996. Dexamethasone attenuates hypoglycemia in ten day old rats treated with TNF alpha. Res. Commun. Mol. Pathol. Pharmacol. 92, 149–154. Beutler, E., Duron, O., Kelly, B.M., 1963. Improved method for the determination of blood glutathione. J. Lab. Clin. Med. 61, 882–888. Bielewicz, J., Kurzepa, J., Lagowska-Lenard, M., Bartosik-Psujek, H., 2010. The novel views on the patomechanism of ischemic stroke. Wiad. Lek. 63, 213–220. Bouchard, J.F., Dumont, E., Lamontagne, D., 1994. Evidence that prostaglandins I2, E2, and D2 may activate ATP sensitive potassium channels in the isolated rat heart. Cardiovasc. Res. 28, 901–905. Busija, D.W., Gaspar, T., Domoki, F., Katakam, P.V., Bari, F., 2008. Mitochondrial-mediated suppression of ROS production upon exposure of neurons to lethal stress: mitochondrial targeted preconditioning. Adv. Drug Deliv. Rev. 60, 1471–1477.

262

BR A I N R ES E A RC H 1 3 8 5 ( 2 01 1 ) 2 5 7 –26 2

Candelario-Jalil, E., Gonzalez-Falcon, A., Garcia-Cabrera, M., Alvarez, D., Al-Dalain, S., Martinez, G., Leon, O.S., Springer, J.E., 2003. Assessment of the relative contribution of COX-1 and COX-2 isoforms to ischemia-induced oxidative damage and neurodegeneration following transient global cerebral ischemia. J. Neurochem. 86, 545–555. Da Silva-Santos, J.E., Santos-Silva, M.C., Cunha Fde, Q., Assreuy, J., 2002. The role of ATP-sensitive potassium channels in neutrophil migration and plasma exudation. J. Pharmacol. Exp. Ther. 300, 946–951. Deb, P., Sharma, S., Hassan, K.M., 2010. Pathophysiologic mechanisms of acute ischemic stroke: an overview with emphasis on therapeutic significance beyond thrombolysis. Pathophysiology. 17, 197–218. Elango, C., Jayachandaran, K.S., Niranjali Devaraj, S., 2009. Hawthorn extract reduces infarct volume and improves neurological score by reducing oxidative stress in rat brain following middle cerebral artery occlusion. Int. J. Dev. Neurosci. 27, 799–803. Gaur, V., Kumar, A., 2010. Behavioral, biochemical and cellular correlates in the protective effect of sertraline against transient global ischemia induced behavioral despair: possible involvement of nitric oxide-cyclic guanosine monophosphate study pathway. Brain Res. Bull. 82, 57–64. Hamberg, M., Svensson, J., Samuelsson, B., 1974. Prostaglandin endoperoxides. A new concept concerning the mode of action and release of prostaglandins. Proc. Natl. Acad. Sci. USA. 71, 3824–3828. Koracevic, D., Koracevic, G., Djordjevic, V., Andrejevic, S., Cosic, V., 2001. Method for the measurement of antioxidant activity in human fluids. J. Clin. Pathol. 54, 356–361. Krawisz, J.E., Sharon, P., Stenson, W.F., 1984. Quantitative assay for acute intestinal inflammation based on myeloperoxidase activity. Assessment of inflammation in rat and hamster models. Gastroenterology 87, 1344–1350. Krnjevic, K., 2008. Electrophysiology of cerebral ischemia Neuropharmacology 55, 319–333. Lacza, Z., Snipes, J.A., Kis, B., Szabo, C., Grover, G., Busija, D.W., 2003. Investigation of the subunit composition and the pharmacology of the mitochondrial ATP-dependent K+ channel in the brain. Brain Res. 994, 27–36. Lakhan, S.E., Kirchgessner, A., Hofer, M., 2009. Inflammatory mechanisms in ischemic stroke: therapeutic approaches. J. Transl. Med. 7, 97. Leker, R.R., Shohami, E., 2002. Cerebral ischemia and trauma-different etiologies yet similar mechanisms: neuroprotective opportunities. Brain Res. Brain Res. Rev. 39, 55–73. Ma, S.G., Fu, R.F., Feng, G.Q., Wang, Z.J., Ma, X.Q., Weng, S.A., 2004. Effect of G(alphaq/11) protein and ATP-sensitive potassium channels on prostaglandin E(1) preconditioning in rat hearts. Acta Pharmacol. Sin. 25, 587–592. Mihara, M., Uchiyama, M., 1978. Determination of malonaldehyde precursor in tissues by thiobarbituric acid test. Anal. Biochem. 86, 271–278. Miranda, K.M., Espey, M.G., Wink, D.A., 2001. A rapid, simple spectrophotometric method for simultaneous detection of nitrate and nitrite. Nitric Oxide 5, 62–71. Naidech, A.M., Levasseur, K., Liebling, S., Garg, R.K., Shapiro, M., Ault, M.L., Afifi, S., Batjer, H.H., 2010. Moderate hypoglycemia is associated with vasospasm, cerebral infarction, and 3-month disability after subarachnoid hemorrhage. Neurocrit. Care 12, 181–187. Nakayama, M., Uchimura, K., Zhu, R.L., Nagayama, T., Rose, M.E., Stetler, R.A., Isakson, P.C., Chen, J., Graham, S.H., 1998. Cyclooxygenase-2 inhibition prevents delayed death of CA1

hippocampal neurons following global ischemia. Proc. Natl Acad. Sci. USA 95, 10954–10959. Nazaroglu, N.K., Sepici-Dincel, A., Altan, N., 2009. The effects of sulfonylurea glyburide on superoxide dismutase, catalase, and glutathione peroxidase activities in the brain tissue of streptozotocin-induced diabetic rat. J. Diab. Complications 23, 209–213. Nistico, R., Piccirilli, S., Sebastianelli, L., Nistico, G., Bernardi, G., Mercuri, N.B., 2007. The blockade of K(+)-ATP channels has neuroprotective effects in an in vitro model of brain ischemia. Int. Rev. Neurobiol. 82, 383–395. Pompermayer, K., Souza, D.G., Lara, G.G., Silveira, K.D., Cassali, G.D., Andrade, A.A., Bonjardim, C.A., Passaglio, K.T., Assreuy, J., Cunha, F.Q., Vieira, M.A., Teixeira, M.M., 2005. The ATP-sensitive potassium channel blocker glibenclamide prevents renal ischemia/reperfusion injury in rats. Kidney Int. 67, 1785–1796. Pompermayer, K., Amaral, F.A., Fagundes, C.T., Vieira, A.T., Cunha, F.Q., Teixeira, M.M., Souza, D.G., 2007. Effects of the treatment with glibenclamide, an ATP-sensitive potassium channel blocker, on intestinal ischemia and reperfusion injury. Eur. J. Pharmacol. 556, 215–222. Racay, P., Chomova, M., Tatarkova, Z., Kaplan, P., Hatok, J., Dobrota, D., 2009. Ischemia-induced mitochondrial apoptosis is significantly attenuated by ischemic preconditioning. Cell. Mol. Neurobiol. 29, 901–908. Radi, R., Beckman, J.S., Bush, K.M., Freeman, B.A., 1991. Peroxynitrite oxidation of sulfhydryls. The cytotoxic potential of superoxide and nitric oxide. J. Biol. Chem. 266, 4244–4250. Rehni, A.K., Shri, R., Singh, M., 2007. Remote ischaemic preconditioning and prevention of cerebral injury. Indian J. Exp. Biol. 45, 247–252. Simard, J.M., Chen, M., Tarasov, K.V., Bhatta, S., Ivanova, S., Melnitchenko, L., Tsymbalyuk, N., West, G.A., Gerzanich, V., 2006. Newly expressed SUR1-regulated NC(Ca-ATP) channel mediates cerebral edema after ischemic stroke. Nat. Med. 12, 433–440. Simard, J.M., Yurovsky, V., Tsymbalyuk, N., Melnichenko, L., Ivanova, S., Gerzanich, V., 2009. Protective effect of delayed treatment with low-dose glibenclamide in three models of ischemic stroke. Stroke 40, 604–609. Strauss, K.I., 2008. Antiinflammatory and neuroprotective actions of COX2 inhibitors in the injured brain. Brain Behav. Immun. 22, 285–298. Sun, X.L., Hu, G., 2010. ATP-sensitive potassium channels: a promising target for protecting neurovascular unit function in stroke. Clin. Exp. Pharmacol. Physiol. 37, 243–252. Taoufik, E., Probert, L., 2008. Ischemic neuronal damage. Curr. Pharm. Des. 14, 3565–3573. Tapuria, N., Kumar, Y., Habib, M.M., Abu Amara, M., Seifalian, A.M., Davidson, B.R., 2008. Remote ischemic preconditioning: a novel protective method from ischemia reperfusion injury—a review. J. Surg. Res. 150, 304–330. Velly, L.J., Canas, P.T., Guillet, B.A., Labrande, C.N., Masmejean, F.M., Nieoullon, A.L., Gouin, F.M., Bruder, N.J., Pisano, P.S., 2009. Early anesthetic preconditioning in mixed cortical neuronal-glial cell cultures subjected to oxygen-glucose deprivation: the role of adenosine triphosphate dependent potassium channels and reactive oxygen species in sevoflurane-induced neuroprotection. Anesth. Analg. 108, 955–963. Wang, C.F., Li, D.Q., Xue, H.Y., Hu, B., 2010. Oral supplementation of catalpol ameliorates diabetic encephalopathy in rats. Brain Res. 1307, 158–165. Zarch, A.V., Toroudi, H.P., Soleimani, M., Bakhtiarian, A., Katebi, M., Djahanguiri, B., 2009. Neuroprotective effects of diazoxide and its antagonism by glibenclamide in pyramidal neurons of rat hippocampus subjected to ischemia–reperfusion-induced injury. Int. J. Neurosci. 119, 1346–1361.