Postischemic Administration Research Articles

L‐theanine Administration Results in Neuroprotection and Prevents Glutamate Receptor Agonist‐Mediated Injury in the Rat Model of Cerebral Ischemia‐Reperfusion

While the neuroprotective effect of green tea (Camellia sinensis) might be explained by the presence of amino acid L-theanine in the tea leaves, it is not known whether postischemic administration of L-theanine could also provide neuroprotection. In the present study, we investigated the neuroprotective effect of L-theanine (1 and 4 mg/kg) administered at 3, 12, and 24 h after reperfusion in the rat model of stroke. We also studied the effect of L-theanine on brain injury caused by exogenous administration of N-methyl-D-aspartate and α-amino-3-hydroxy-5-methyl-isoxazole-4-propionate/kainate receptor agonists during reperfusion. Rats were subjected to 30-min middle cerebral artery occlusion followed by 48-h reperfusion. Neurological deficit and infarct size were determined at the end of reperfusion. At 3 and 12 h, but not at 24 h of reperfusion, L-theanine substantially reduced the size of brain infarct. Neurological status was improved when L-theanine was administered 3, 12, and 24 h after reperfusion. Repeated intrastriatal injections of L-theanine at a total dose of 800 µg/kg during reperfusion prevented brain injury caused by glutamate receptor agonists. In conclusion, L-theanine at reperfusion exerts neuroprotective effect in the in vivo rat model of stroke. Local treatment with L-theanine at reperfusion prevents glutamate receptor agonist-mediated brain injury.

Intralipid, a Clinically Safe Compound, Protects the Heart Against Ischemia-Reperfusion Injury More Efficiently Than Cyclosporine-A

We have recently shown that postischemic administration of intralipid protects the heart against ischemia-reperfusion injury. Here we compared the cardioprotective effects of intralipid with cyclosporine-A, a potent inhibitor of the mitochondrial permeability transition pore opening. In vivo rat hearts or isolated Langendorff-perfused mouse hearts were subjected to ischemia followed by reperfusion with intralipid (0.5%, 1% and 2% ex-vivo, and 20% in vivo), cyclosporine-A (0.2 μM, 0.8 μM, and 1.5 μM ex- vivo and 10 mg/kg in vivo), or vehicle. The hemodynamic function, infarct size, calcium retention capacity, mitochondrial superoxide production, and phosphorylation levels of protein kinase B (Akt)/glycogen synthase kinase-3β (GSK-3β) were measured. The values are mean ± SEM. Administration of intralipid at reperfusion significantly reduced myocardial infarct size compared with cyclosporine-A in vivo (infarct size/area at risk)%: 22.9 ± 2.5% vs. 35.2 ± 3.5%; P = 0.030, n = 7/group). Postischemic administration of intralipid at its optimal dose (1%) was more effective than cyclosporine-A (0.8 μM) in protecting the ex vivo heart against ischemia-reperfusion injury, as the rate pressure product at the end of reperfusion was significantly higher (mmHg · beats/min: 12,740 ± 675 [n = 7] vs. 9,203 ± 10,781 [n = 5], P = 0.024), and the infarct size was markedly smaller (17.3 ± 2.9 [n = 7] vs. 29.2 ± 2.7 [n = 5], P = 0.014). Intralipid was as efficient as cyclosporine-A in inhibiting the mitochondrial permeability transition pore opening (calcium retention capacity = 280 ± 8.2 vs. 260.3 ± 2.9 nmol/mg mitochondria protein in cyclosporine-A, P = 0.454, n = 6) and in reducing cardiac mitochondrial superoxide production. Unlike intralipid, which increased phosphorylation of Akt (6-fold) and GSK-3β (5-fold), cyclosporine-A had no effect on the activation of these prosurvival kinases. Although intralipid inhibits the opening of the mitochondrial permeability transition pore as efficiently as cyclosporine-A, intralipid is more effective in reducing the infarct size and improving the cardiac functional recovery.

Resistance to reperfusion injury following short term postischemic administration of natural honey in globally ischemic isolated rat heart.

RESULTS of our previous study revealed that preischemic perfusion of honey before zero flow global ischemia had cardioprotective effects in rat. The present study investigated potential resistance to reperfusion injury following short term postischemic administration of natural honey in globally ischemic isolated rat heart. Male Wistar rats were divided into five groups (n=10-13). The rat hearts were isolated, mounted on a Langendorff apparatus, allowed to equilibrate for 30 min then subjected to 30 min global ischemia. In the control group, the hearts were reperfused with drug free normal Krebs-Henseleit (K/H) solution before ischemia and during 120 min reperfusion. In the treatment groups, reperfusion was initiated with K/H solution containing different concentration of honey (0.25, 0.5, 1 and 2%) for 15 min and was resumed until the end of 120 min with normal K/H solution. In the control group, VEBs number was 784±199, while in honey concentration of 0.25, 0.5, 1 and 2%, it decreased to 83±23 (P<0.001), 138±48 (P<0.01), 142±37 (P<0.001) and 157±40 (P<0.01), respectively. Number and duration of VT and time spent in reversible VF were also reduced by honey. In the control group, the infarct size was 54.1±7.8%, however; honey (0.25, 0.5, 1 and 2%) markedly lowered the value to 12.4±2.4, 12.7±3.3, 11.3±2.6 and 7.9±1.7 (P<0.001), respectively. Postischemic administration of natural honey in global ischemia showed protective effects against ischemia/reperfusion (I/R) injuries in isolated rat heart. Antioxidant and radical scavenging activity, lipoperoxidation inhibition, reduction of necrotized tissue, presence of rich energy sources, various type of vitamins, minerals and enzymes and formation of NO-contain metabolites may probably involve in those cardioprotective effects.

Evaluation of the Neuroprotective Effect of Minocycline in a Rabbit Spinal Cord Ischemia Model

To investigate whether postischemic administration of minocycline attenuates hind-limb motor dysfunction and gray and white matter injuries after spinal cord ischemia. A prospective, randomized, laboratory investigation. Laboratory in university, single institution. Male New Zealand White rabbits. Spinal cord ischemia was induced by an occlusion of the infrarenal aorta for 15 minutes. The groups were administered minocycline 1 hour after reperfusion (M-1; n = 8), minocycline 3 hours after reperfusion (M-3; n = 8), saline 1 hour after reperfusion (control [C]; n = 8), or saline and no occlusion (sham; n = 4). Minocycline was administered intravenously at 10 mg/kg 6 times at 12-hour intervals until 60 hours after the initial administration. Hind-limb motor function was assessed using the Tarlov score. For histologic assessments, gray and white matter injuries were evaluated 72 hours after reperfusion using the number of normal neurons and the percentage of areas of vacuolation, respectively. Motor function 72 hours after reperfusion was significantly better in group M-1 than in group C. The number of neurons in the anterior horn was significantly larger in group M-1 than in groups M-3 or C but did not differ significantly between groups M-3 and C. No significant difference was noted in the percentage of areas of vacuolation among the ischemia groups. Minocycline administration beginning at 1 hour after reperfusion improved hind-limb motor dysfunction and attenuated gray matter injury in a rabbit spinal cord ischemia model.

Reduction of Ischemia/Reperfusion Injury With Bendavia, a Mitochondria‐Targeting Cytoprotective Peptide

BackgroundManifestations of reperfusion injury include myocyte death leading to infarction, contractile dysfunction, and vascular injury characterized by the “no-reflow” phenomenon. Mitochondria-produced reactive oxygen species are believed to be centrally involved in each of these aspects of reperfusion injury, although currently no therapies reduce reperfusion injury by targeting mitochondria specifically.Methods and ResultsWe investigated the cardioprotective effects of a mitochondria-targeted peptide, Bendavia (Stealth Peptides), across a spectrum of experimental cardiac ischemia/reperfusion models. Postischemic administration of Bendavia reduced infarct size in an in vivo sheep model by 15% (P=0.02) and in an ex vivo guinea pig model by 38% to 42% (P<0.05). In an in vivo rabbit model, the extent of coronary no-reflow was assessed with Thioflavin S staining and was significantly smaller in the Bendavia group for any given ischemic risk area than in the control group (P=0.0085). Myocardial uptake of Bendavia was ≈25% per minute, and uptake remained consistent throughout reperfusion. Postischemic recovery of cardiac hemodynamics was not influenced by Bendavia in any of the models studied. Isolated myocytes exposed to hypoxia/reoxygenation showed improved survival when treated with Bendavia. This protection appeared to be mediated by lowered reactive oxygen species–mediated cell death during reoxygenation, associated with sustainment of mitochondrial membrane potential in Bendavia-treated myocytes.ConclusionsPostischemic administration of Bendavia protected against reperfusion injury in several distinct models of injury. These data suggest that Bendavia is a mitochondria-targeted therapy that reduces reperfusion injury by maintaining mitochondrial energetics and suppressing cellular reactive oxygen species levels. (J Am Heart Assoc. 2012;1:e001644 doi: 10.1161/JAHA.112.001644.)

Delayed post-ischaemic administration of xenon reduces brain damage in a rat model of global ischaemia

Cerebral ischaemia is among the leading causes of death, disability and economic expense in the world. Xenon has been shown to be neuroprotective both in vivo and in vitro, predominantly when administered as a preconditioning agent. We have used a rat model of global ischaemia to investigate whether xenon-induced neuroprotection is observed following an ischaemic insult.

Effects of MS-31-038, a novel Na(+)-H+ exchange inhibitor, on the myocardial infarct size in rats after postischemic administration.

The abilities of 2-(2-methylphenyl)-5,7-dimethoxy-4-quinolyl carbonylguanidine dihydrochloride (CAS 181048-29-3, MS-31-050) and 2-phenyl-8-(2-methoxyethoxy)-4-quinolyl carbonylguanidine bismethanesulfonate (CAS 181048-36-2, MS-31-038) in inhibiting Na(+)-H+ exchange, ischemia- and reperfusion-induced injury were determined and compared with those of 4-isopropyl-3-methylsulfonylbenzoyl guanidine methanesulfonate (CAS 159138-81-5, IMGM), a selective inhibitor of Na(+)-H+ exchange. MS-31-050 and IMGM exhibited comparable inhibitory effects on Na(+)-dependent pH recovery and antiarrhythmic effects during ischemia in anesthetized rats. In rats subjected to ischemia and reperfusion, MS-31-050 (10 mg/kg i.v.) significantly reduced the infarct size when given prior to the onset of ischemia. However, postischemic treatment with either MS-31-050 or IMGM failed to protect reperfused hearts. In contrast, MS-31-038 reduced the infarct size dramatically from 65.4 +/- 7.4% in control to 29.9 +/- 11.6% at 3 mg/kg and 9.8 +/- 3.4% at 10 mg/kg even when administered before the onset of reperfusion. These results suggest the beneficial effects of Na(+)-H+ exchange inhibitors on myocardial ischemia/reperfusion injury.

Abstract P296: Intralipid Protects the Heart Against Ischemia/Reperfusion Injury More Efficiently than Cyclosporine-A

It has been shown that intralipid protects the heart against ischemia reperfusion (I/R) injury. Here we compared the cardioprotective effect of Intralipid (ILP) with cyclosporine-A (CsA), a potent inhibitor of the mitochondria permeability transition pore (mPTP) opening. Isolated hearts (Langendorff) from male mice were subjected to 20 minutes of global ischemia followed by reperfusion with Krebs Henseleit buffer (CTRL), additional 1% ILP (ILP group) or 1.5µM CsA (CsA group). The reperfusion duration was 40 min for measurement of cardiac function and infarct size, 10 min for measurement of mitochondrial calcium retention capacity and 5 min for superoxide production measurements using electron spin resonance. Post-ischemic administration of ILP improved the heart functional recovery even better than CsA as Rate pressure product (RPP) at the end of 40 min was significantly higher in ILP group compared to CsA group (13676±611 mmHg*beats/min (n=6) in ILP group vs. 7426±1158 mmHg*beats/min (n=3) in CsA, P&lt;0.01). Consistent with the heart function, the infarct size was also significantly smaller in ILP compared to CsA group (18.3±2.4% (n=9) in ILP vs. 29.9±2.0% (n=3) in CsA, P&lt;0.01). The heart functional recoveries were significantly higher and the infarct sizes were significantly smaller in both groups compared to CTRL (RPP=2999±863 mmHg*beats/min (n=6), the infarct size=54.8±2.9% (n=10)). ILP was as efficient as CSA in inhibiting mPTP opening as calcium retention capacity (CRC) was not significantly different between the mitochondria isolated from the heart which were reperfused with ILP or CSA (274.3±8.4 nM/mg-mitochondria protein (n=6) in ILP vs. 260.3±2.9 nM/mg-mitochondria protein (n=6) in CsA). The CRC in both groups were higher than CTRL (168.6±9.6 nM/mg mitochondria protein (n=7)). ILP prevented superoxide production in mitochondria similarly as CSA (0.344±0.1104 (n=4) in ILP vs. 0.5744±0.1445 (n=4) in CsA) which were significantly lower than CTRL (1±0.1197 (n=4), normalized to CTRL). In conclusion, although ILP inhibits the opening of the mPTP as efficiently as CsA, ILP is more effective than CsA in reducing the infarct size and improving the heart functional recovery.

Abstract 272: Intralipid Protects Late Pregnant Mouse Heart Against Ischemia/Reperfusion Injury More Efficiently than Cyclosporine-A

Introduction Recently, we found that hearts of late pregnant (LP) mice have higher vulnerability to ischemia/reperfusion (I/R) injury compared to hearts of non-pregnant mice. We also found intralipid (ITLD), a clinically safe compound protects the male rodent hearts against I/R injury if administrated at reperfusion. Here we explored whether ITLD can protect the function of LP mouse hearts against I/R injury, and compared its action with cyclosporine-A (CsA), a potent inhibitor of mithochondrial permeability transition pore (mPTP) opening. Methods Isolated LP mouse hearts were subjected to 20 min ischemia followed by reperfusion with i) Krebs Henseleit buffer (CTRL), ii) additional 1% ITLD or iii) 1.5µM CsA. The duration of reperfusion was 40 min for heart functional measurements and infarct size assessment and 10 min for measurements of mitochondrial calcium retention capacity (CRC). Two way ANOVA was used for statistical analysis. P&lt;0.05 were considered statistically significant. Values are expressed as mean± SEM. Results The heart functional recovery of LP mice after ischemia was very poor. Postischemic administration of ITLD significantly improved the cardiac function of LP mice; the rate pressure product (RPP) at the end of reperfusion was improved from 1614±438 mmHg*beats/min (n=4) to 11556±784 mmHg*beats/min (n=3) with addition of ITLD. The infarct size was also significantly reduced by ITLD to 17±2% (n=3) from 59±5% (n=4) in CTRL (P&lt;0.01). Interestingly, ITLD was even more efficient than CsA in protecting the hearts of LP mice against ischemia reperfusion injury as in CsA group the RPP was significantly lower at the end of reperfusion (5960±1332, n=4) and the infarct size was larger (25±3%, n=4) compared to ITLD(P&lt;0.05). ITLD-induced cardioprotection of LP mouse hearts against I/R was associated with a delay in the opening of the mPTP similar to CsA. The calcium load required for triggering the opening of the mPTP increased from 180±7 nM/mg (n=6) protein in CTRL to 290±12 nM/mg (n=6) protein in ITLD (P&lt;0.01), which was not significantly different than 276±20 nM/mg protein in CsA group. Conclusion ITLD protects late pregnant mouse hearts against I/R injury more efficiently than CsA by inhibiting the mPTP opening.

Intraperitoneal aminoguanidine improves sciatic nerve ischemia-reperfusion injury in male sprague-dawley rats.

The present work was designed to investigate the potential protective effects of post-ischemic treatment with aminoguanidine (AG) on sciatic nerve ischemia/reperfusion (I/R) injury in rat. Seventy-two rats were divided into 12 groups (n = 6). We used ischemia model in these groups by occluding the right common iliac and femoral arteries for 3 h with a silk suture 6-0 using slipknot technique. Treatment groups (2, 4, 6, 8, 10, and 12) received 150 mg/kg AG intraperitoneally 24 h after induction of ischemia. After certain time intervals of reperfusion (2, 4, 7, 14, and 28 days), the function of the hind limb was assessed using behavioral scores based on gait, racing reflex, toe spread, pinch sensitivity, paw position, and grasp. After euthanasia, sciatic nerves were removed at the end of reperfusion times and sections were cut at 5 μm, then were stained for light microscopy studies and graded for ischemic fiber degeneration (IFD), edema, and apoptosis. Maximal behavioral deficit occurred at 7 days of reperfusion. The comparison of behavioral score pertaining to the control and AG groups revealed significant differences and showed also a better time course in recovery (P < 0.05). Other than 3 and 4 groups, the amount of edema in AG treatment groups showed significant differences compared with control groups (P < 0.05). IFD was also significantly decreased in the AG treatment groups than controls. Most importantly, I/R-induced apoptosis were improved significantly on the 4th, 7(th), and 14th days of reperfusion in AG-treated groups compared to controls. In conclusion, our findings suggest that post-ischemic administration of AG exhibits protective effect against sciatic nerve I/R injury.

Phosphorylation of GSK-3b is Required for Intralipid to Protect the Heart Against Ischemia/Reperfusion Injury

Recently we found that administration of Intralipid (ILP) during reperfusion significantly improves post-ischemic cardiac function and reduces the myocardial infarct size by ∼70% , both in the isolated mouse heart and in-vivo rat heart. Here we investigated whether ILP-induced cardioprotection is mediated through the inhibition of GSK-3b. Wild type (WT) C57BL/6 male mice and GSK-3b Knockin (KI) were used. The isolated hearts were subjected to 20 min of global normothermic ischemia followed by reperfusion with 1% ILP (40 min for heart function and infarct size and 10 min for calcium retention capacity (CRC) experiments). The left ventricular (LV) systolic pressure, LV end-diastolic pressure (LVDP), heart rate, maximum velocity of contraction (dP/dt max) and maximum velocity of relaxation (dP/dt min) were recorded. Myocardial necrosis was assessed using TTC staining. Mitochondria were isolated to measure CRC by calculating the number of pulses required to trigger the opening of the mitochondrial transition permeability pore as a result of calcium overload. Before ischemia, the baseline RPP, LVDP, dP/dtmax and dP/dtmin in GSK-3b KI mice were similar to WT. However, the functional recovery during reperfusion was very poor in GSK-3b KI mice. At the end of 40 min of reperfusion, the RPP was 1990±499 in GSK-3b KI mice vs. 15405±1011mmHg∗beats/min in WT, the LV dP/dtmax was 239.7±17.8 in GSK-3b KI vs. 2703±145 mmHg/s in WT and the LV dP/dtmin was 219±14 in GSK-3b KI vs. 1683±66 mmHg/s in WT. The infarct size was significantly larger compared to WT (45.3±10.3 vs. 16.7±2.33% in WT, P<0.001). Postischemic administration of ILP in GSK-3b KI mice demonstrated lower CRC than WT (1.3±0.1 vs. 2.7±0.06 µM/mg-mitochondrial protein in WT). In conclusions, these data demonstrate that phosphoryltaion of GSK-3b is required for the cardioprotective action of ILP.

Postischemic administration of liposome-encapsulated luteolin prevents against ischemia-reperfusion injury in a rat middle cerebral artery occlusion model

Oxidative stress-induced neuronal cell death has been implicated in neurodegenerative diseases; one such disease is ischemic stroke. Using reactive oxygen species (ROS)-insulted primary neurons, we screened neuroprotectants with clinical potential and then, using ischemia/reperfusion (I/R) model, investigated the anti-ischemic potential of candidate neuroprotectants. Here, we showed that luteolin, isolated from the ripe fruit of Perilla frutescens (L.) Britt, exhibited a neuroprotective action upon the in vitro platform, thus serving as candidate for in vivo pharmacological evaluation. Liposome-encapsulated luteolin produced dramatic preventing effects on I/R-induced behavioral and histological injuries after a 13-day post-ischemic treatment. Furthermore, this phytochemical not only lowered the increased level of mitochondrial ROS but also substantially up-regulated the decreased activity of catalase and glutathione in I/R rat brains. Collectively, luteolin as a neuroprotectant acts by anti-ischemic activity likely through a rebalancing of pro-oxidant/antioxidant status. Its multitarget mechanisms implicate potential effectiveness for clinically treating ischemia stroke.

Reduction of the infarct size by simultaneous administration of l-histidine and diphenhydramine in ischaemic rat brains

Aims While diphenhydramine is a histamine H 1 receptor antagonist, the agent has been shown to inhibit histamine- N-methyltransferase, a histamine inactivating enzyme in the brain. Since an increase in the brain concentration of histamine ameliorates reperfusion injury after cerebral ischaemia, effects of postischaemic administration of diphenhydramine were evaluated in rats treated with l-histidine, a precursor of histamine. Methods The right middle cerebral artery was occluded for 2 h, and the infarct size was determined 24 h after reperfusion of cerebral blood flow. Brain oedema was evaluated by comparing the area of the right hemisphere to that of the left hemisphere. Results Focal cerebral ischaemia provoked marked damage in saline-treated control rats, and infarct volumes in the striatum and cerebral cortex were 56 (49–63) mm 3 and 110 (72–148) mm 3, respectively (means and 95% confidence intervals, n = 6). Administration of l-histidine (1000 mg/kg, intraperitoneal) immediately after reperfusion did not affect the infarct size. Simultaneous administration of diphenhydramine (20 mg/kg, intraperitoneal) with l-histidine reduced the infarct size to 25% and 21% of that in the control group, respectively. The combination therapy completely reduced ischaemia-induced brain oedema. Conclusion Because histamine H 1 action does not influence ischaemic brain damage, elevation of the central histamine concentration by blockade of histamine- N-methyltransferase may be a likely mechanism responsible for the alleviation.

Roles of Cyclooxygenase 2 in Sevoflurane- and Olprinone-Induced Early Phase of Preconditioning and Postconditioning Against Myocardial Infarction in Rat Hearts

It is known that selective cyclooxygenase 2(COX-2) inhibitors increase mortality in patients with previous myocardial infarction, and it has been suggested that COX-2 plays an important role in cardioprotection against ischemia. The current study was carried out to determine whether COX-2 is involved in the mechanisms of sevoflurane- and olprinone-induced early-phase preconditioning (E-PreC) and postconditioning (PostC) in rat hearts. Male Sprague-Dawley rats were anesthetized with sodium pentobarbital. After opening the chest, all rats underwent 30-minute occlusion of left anterior descending coronary artery followed by 2-hour reperfusion, and the infarct size was measured after the reperfusion. The rats were randomly assigned to groups with pre- and postischemic exposure to sevoflurane and administration of olprinone with or without a selective COX-2 inhibitor, NS-398. The infarct size in the control group was 42% ± 6% of the area at risk. Infarct size was significantly reduced by pre- and postischemic administration of sevoflurane (16% ± 7% and 17% ± 6%, respectively), as well as by olprinone (14% ± 4% and 15% ± 10%, respectively). NS-398 prevented the protective effects of both pre- and postischemic exposure to sevoflurane (35% ± 8% and 42% ± 10%, respectively), whereas the protective effect of both pre- and postischemic administration of olprinone was not influenced by NS-398 (12% ± 5% and 19% ± 7%, respectively). Cyclooxygenase 2 could be a critical mediator of sevoflurane-induced but not olprinone-induced E-PreC or PostC in rat hearts.

Post-ischemic administration of nimodipine following focal cerebral ischemic-reperfusion injury in rats alleviated excitotoxicity, neurobehavioural alterations and partially the bioenergetics

The present study focuses on the temporal calcium significance in middle cerebral artery occluded (2h ischemia)–reperfused (70h reperfusion) rats treated with nimodipine (NM) through concurrent measurements of excitotoxicity, bioenergetics and neurobehavioural paradigms. Further, the suitable therapeutic time window of calcium channel antagonism in stroke was also ascertained. NM (5mg/kg, i.p.) was administered at pre (30min before the induction of ischemia), during (1h following occlusion of MCA) and post-ischemic (3h after begin of reperfusion) states. The magnitude of neuroprotection in terms of excitotoxicity (glutamate, glutamine synthetase, Na+K+ATPase), bioenergetics (ATP, NAD+) and neurobehavioural paradigms (neurological score and open field exploratory behaviour) were measured and compared to ensure the therapeutic time-window of NM in stroke. Middle cerebral artery occlusion-reperfusion (MCAO/R) was found to elevate glutamate, glutamine synthetase levels and deplete Na+K+ATPase activity in the vehicle treated group (IR group). Significant decrease in bioenergetics such as ATP and NAD+ levels was also observed. Further, IR group demonstrated grievous oxidative stress (increase in lipid peroxidation, protein carbonyl content, nitrite/nitrate levels and decrease in superoxide dismutase and glutathione levels) along with anxiogenic behaviour, neurological deficits and neuronal damage and decreased nuclear to cytoplasm ratio in CA1 hippocampal region. Post-ischemic NM administration reversed the excitotoxicity, neurobehavioural and histopathological alterations significantly, but it restored bioenergetics level in MCAO/R rats only partially.These findings were further confirmed with the combination treatment (CT) of post-ischemic NM and pre-ischemic memantine (MN) administration, since MN showed protective effect in the pre-ischemic administration (Babu and Ramanathan, 2009). The failure of NM to forefend the neurodegeneration on pre- and during-ischemic administration suggests that the initial phase damages in ischemic-reperfusion (IR) might be mediated through other mechanism(s) such as glutamergic overstimulation or reverse operation of glutamate transporters. From the present study, it is concluded that calcium plays a crucial role in post-ischemic status and the suitable therapeutic time window of calcium antagonism is the post-ischemic state.

Comparison of the post-embolization effects of tissue-plasminogen activator and simvastatin on neurological outcome in a clinically relevant rat model of acute ischemic stroke

Data has emerged, largely from non-thromboembolic animal models of stroke, that suggests that statins, which have efficacy in preventing strokes when given pre-ischemically, may have a positive effect on stroke even when given post-ischemically, possibly through pleitropic cerebrovascular effects. The goal of this study was to characterize the effects of IV tPA in a clinically relevant model of stroke utilizing a vascular occlusion with a freshly formed clot, and evaluate the effects of post-ischemic administration of simvastatin on stroke outcome in this model. Neurological deficit, clot burden, and lesion volume were assessed after treatment with tPA in one experiment, and after treatment with simvastatin in another. In the tPA experiment, treatment with 10 mg/kg of tPA IV (with 20% given as an initial bolus, and 80% given as an infusion over the remaining 30 min), starting within an hour after stroke, resulted in significant reductions, compared with control animals, in neurological deficit (mean ± SD neuroscores of 21.5 ± 21.1 and 30 ± 29.3, respectively, p = 0.005), clot burden ( p = 0.010) and lesion volume ( p = 0.049) at 24 h. In the simvastatin experiment on the other hand, treatment with a 20 mg/kg of simvastatin as a single intraperitoneal dose within an hour after stroke resulted in no salutary effects on neurological deficit, clot burden or lesion volume compared with controls at 24 h. These results suggest that more research needs to be done to fully ascertain the therapeutic potential and optimal dosing paradigm of a post-ischemic treatment with a statin.

Telmisartan, a partial agonist of peroxisome proliferator-activated receptor γ, improves impairment of spatial memory and hippocampal apoptosis in rats treated with repeated cerebral ischemia

Telmisartan, an angiotensin type 1 receptor blocker (ARB), is used for hypertension to control blood pressure and has been shown to have a partial agonistic effect on peroxisome proliferator-activated receptor γ (PPARγ). Recently, the ligand of PPARγ has been implicated in cerebroprotection due to its anti-inflammatory effect. In this study, we investigated whether telmisartan has a cerebroprotective effect on memory impairment and neuronal cell death induced by repeated cerebral ischemia. Repeated cerebral ischemia (RI: 10 min × 2) significantly induced impairment of spatial memory and hippocampal apoptosis in rats. Fourteen-day pre- and post-ischemic administration of telmisartan (0.3, 1, 3 mg/kg/day, p.o.) increased the number of correct choices and reduced the number of errors made in the eight-arm radial maze task in a dose-dependent manner in RI treated rats. TUNEL-positive cells in the hippocampus CA1 areas were also reduced following 14-day administration of telmisartan (3 mg/kg/day, p.o.). Seven-day post-ischemic administration of telmisartan improved spatial memory and reduced TUNEL-positive cells while 7-day pre-ischemic administration of telmisartan did not. These effects of telmisartan were inhibited by the PPARγ antagonist, GW9662. On further experiment, 7-day post-ischemic administration of telmisartan reduced the expression of caspase-3 in the hippocampus, and this effect was also inhibited by GW9662. These results suggest that telmisartan improves memory impairment and reduces neuronal apoptosis via a PPARγ-dependent caspase-3 inhibiting mechanism. Telmisartan, which has the unique character of having both ARB and PPARγ agonistic effect, will be useful for preventing memory impairment after cerebrovascular disease.

Posttreatment but Not Pretreatment with Selective β-Adrenoreceptor 1 Antagonists Provides Neuroprotection in the Hippocampus in Rats Subjected to Transient Forebrain Ischemia

beta-Adrenoreceptor antagonists provide neuroprotection against focal cerebral ischemia, but the effects of these antagonists on experimental global cerebral ischemia are unknown. That is, the effect of beta-adrenoreceptor antagonism in vulnerable brain regions after ischemic insult has not been examined. Therefore, we investigated the neuroprotective effects of preischemic or postischemic administration of propranolol (a nonselective beta-adrenoreceptor antagonist), esmolol, and landiolol (selective beta-adrenoreceptor 1 antagonists) against forebrain ischemia in rats. IV administration of saline 10 microL . h(-1), propranolol 100 microg . kg(-1) . min(-1), esmolol 200 microg . kg(-1) . min(-1), or landiolol 50 microg . kg(-1) . min(-1) in male Sprague-Dawley rats was started 30 minutes before or 60 minutes after 8-minute bilateral carotid artery occlusion combined with hypotension (35 mm Hg) under isoflurane (1.5%) anesthesia. All drugs were administered continuously until 5 days after reperfusion, and the animals were evaluated neurologically and histologically after this 5-day period. Preischemic treatment with propranolol, esmolol, or landiolol failed to provide neuroprotection against forebrain ischemia in the hippocampus. Rats treated with propranolol tended to have a worse score for motor activity and a higher mortality rate (up to 64%), but the differences with other groups were not statistically significant. Postischemic treatment with esmolol and landiolol, but not with propranolol, reduced neuronal injury after forebrain ischemia. However, motor activity did not differ among rats treated postischemically with any of the beta-adrenoreceptor antagonists or saline. Postischemic treatment with esmolol and landiolol provided neuroprotection in the hippocampus in rats subjected to bilateral carotid artery occlusion combined with hemorrhagic shock, whereas treatment with propranolol failed to show neuroprotection. We suggest that concomitant beta-blockade and shock might work as a systemic depressant, rather than a neuroprotectant, resulting in exacerbation of cerebral ischemia.

Postischemic poly (ADP-ribose) polymerase (PARP) inhibition reduces ischemia reperfusion injury in a hind-limb ischemia model

Several experiments were designed to determine whether the systemic, postischemic administration of PJ34,which is a poly-adenosine diphosphate (ADP)-ribose polymerase inhibitor, decreased tissue injury and inflammation after hind-limb ischemia reperfusion (I/R). C57BL6 mouse limbs were subjected to 1.5 h ischemia followed by 24-h reperfusion. The treatment group (PJ) received intraperitoneal PJ34 (30 mg/kg) immediately before reperfusion, as well as 15 min and 2 h into reperfusion. The control group (CG) received lactated Ringer's alone at the same time intervals as PJ34 administration. The skeletal muscle levels of adenosine triphosphate (ATP), macrophage inflammatory protein-2 (MIP-2), keratinocyte derived chemokine (KC), and myeloperoxidase (MPO) were measured. Quantitative measurement of skeletal muscle tissue injury was assessed by microscopic analysis of fiber injury. ATP levels were higher in limbs of PJ versus CG mice (absolute ATP: 4.7 +/- 0.35 vs 2.3 +/- 0.15-ng/mg tissue, P = .002). The levels of MIP-2, KC, and MPO were lower in PJ versus CG mice (MIP-2: 1.4 +/- 0.34 vs 3.67 +/- 0.67-pg/mg protein, P = .014; KC: 4.97 +/- 0.97 vs 12.65 +/- 3.05-pg/mg protein, P = .037; MPO: 46.27 +/- 10.53 vs 107.34 +/- 13.58-ng/mg protein, P = .008). Muscle fiber injury was markedly reduced in PJ versus CG mice (4.25 +/- 1.9% vs 22.68 +/- 3.0% total fibers, P = .0004). Systemic postischemic administration of PJ34 preserved skeletal muscle energy levels, decreased inflammatory markers, and preserved tissue viability post-I/R. These results support PARP inhibition as a viable treatment for skeletal muscle I/R in a clinically relevant post hoc scenario.

Intralipid Induces Cardioprotection against Ischemia-Reperfusion Injury by Inhibiting the Mitochondrial Permeability Transition Pore Opening Via the PI3K/AKT Pathway

Acute myocardial infarction is a major cause of mortality, and the best hope of salvaging viable myocardium is by rapid cardiac reperfusion. A novel cardioprotective drug which could be applied at the time of reperfusion after acute infarction would be ideal. Here we tested the hypothesis that administration of Intralipid at the onset of reperfusion protects the heart against ischemia reperfusion (I/R) injury. Isolated hearts (Langendorff) from male mice were subjected to 20 minutes of global normothermic (37°C) ischemia followed by 40 minutes of reperfusion with Krebs Henseleit buffer (CTRL) or with additional 1% Intralipid (ILP). Postischemic treatment with Intralipid significantly improved the cardiac function; the rate pressure product (RPP) was increased from 3432±334mmHg∗beats/min in CTRL to 15405±1011mmHg∗beats/min in ILP. Consistent with the higher functional recovery in ILP, the infarct size was markedly smaller in ILP (19±3%,n=8) compared to CTRL (55.6±3.4%,n=10). The inhibition of mitochondria permeability transition pore (mPTP) opening during reperfusion has been shown to induce cardioprotection. To investigate whether intralipid-induced cardioprotection occurs by inhibition of the mPTP opening, we compared the viability of isolated mitochondria by calcium overload. Postischemic administration of Intralipid inhibited the opening of the mPTP as calcium retention capacity was higher in the ILP group compared to control (2.7±0.06 vs. 1.5±0.11 μM/mg-mitochondrial protein, p<0.05). To identifiy the key signaling molecules involved in regulating mPTP opening, Western Blot analyis of heart lysates was performed. The activity of AKT/ERK1/GSK were respectively 2.3, 5 and 2.7 fold higher in ILP compared to CTRL. The involvement of PI3K/AKT pathway was further investigated by LY294002, a specific inhibitor of PI3K. The Intralipid-induced cardioprotection was fully abolished in the presence of LY294002, indicating the cardioprotective action of Intralipid is mediated via PI3K/AKT pathway.