Neuroprotective Effect Of Ischemic Postconditioning Research Articles

Restoration of the Attenuated Neuroprotective Effect of Ischemic Postconditioning in Diabetic Mice by SGLT Inhibitor Phlorizin.

The study has been commenced to discover the potential of sodium dependent glucose co-transporters (SGLT) in neuroprotective mechanism of ischemic postconditioning (iPoCo) in diabetic and non-diabetic mice. Cerebral ischemic injury in mice was induced by bilateral carotid artery occlusion (BCAO) for 12 min followed by reperfusion for 24 hr. For iPoCo, three episodes of carotid artery reperfusion and occlusion of 10 sec each were instituted immediately after BCAO, followed by 24 hr reperfusion. Learning and memory were evaluated using the Morris water maze test. Motor coordination was assessed using rotarod test, inclined beam walking test, neurological severity score (NSS), and lateral push response. Glutathione and Thiobarbituric acid reactive species level was quantified to evaluate the oxidative stress; the cholinergic activity of the brain was estimated in terms of acetylcholinestrase activity, and the levels of myeloperoxidase were measured as inflammation marker. Cerebral infarct size was evaluated using triphenyltetrazolium chloride staining. Fasting blood glucose levels of animals were taken before and 6 hr after the surgical procedure. BCAO resulted in impairment of memory and motor coordination and biochemical alterations along with a marked rise in cerebral infarct size and NSS. iPoCo diminished the deadly effect of BCAO in non-diabetic mice; however, it failed to abolish the deleterious effects of ischemia- reperfusion injury in diabetic mice. Pretreatment of Phlorizin (SGLT-inhibitor) potentiated the neuroprotective effects of iPoCo in non-diabetics and restored the protective effect of iPoCo in diabetic mice. It may be concluded that the neuroprotective effect of iPoCo is abolished in diabetic mice, and SGLT plays an important role in neuroprotection.

Overview of Experimental and Clinical Findings regarding the Neuroprotective Effects of Cerebral Ischemic Postconditioning.

Research on attenuating the structural and functional deficits observed following ischemia-reperfusion has become increasingly focused on the therapeutic potential of ischemic postconditioning. In recent years, various methods and animal models of ischemic postconditioning have been utilized. The results of these numerous studies have indicated that the mechanisms underlying the neuroprotective effects of ischemic postconditioning may involve reductions in the generation of free radicals and inhibition of calcium overload, as well as the release of endogenous active substances, alterations in membrane channel function, and activation of protein kinases. Here we review the novel discovery, mechanism, key factors, and clinical application of ischemic postconditioning and discuss its implications for future research and problem of clinical practice.

The role of AMPA-receptors in mechanisms of neuroprotective effect of cerebral ischemic postconditioning

Objective. The purpose of the study is to explore the role of AMPA receptors in the neuroprotective effect of ischemic postconditioning (IРost) in the model of reversible global forebrain ischemia in Mongolian gerbils. Design and methods. Reversible global cerebral ischemia was induced by bilateral occlusion of common carotid arteries in Mongolian gerbils for 7 minutes. IРost was induced by three 15-s episodes of reperfusion/reocclusion. Antagonist of AMPA receptors NBQX was administered intraperitoneally at 2 nd minute of ischemia at a dose of 30 mg/kg. In the early and delayed reperfusion period, the number of viable neurons in the CA1, CA2, CA, and CA4 fields of the hippocampus was estimated. Results. Reversible 7 minutes ischemia followed by 48 hours of reperfusion resulted in a significant reduction in the number of viable neurons in the fields CA1 and СА3 of the hippocampus, while in the late reperfusion period a significant reduction in the number of viable neurons was observed in fields CA1, СА3 and CA4. Application of IРost has led to an increase in the number of viable neurons in fields CA1 and CA3 in the early reperfusion period and in fields CA1, CA3 and CA4 — in delayed period. AMPA receptor antagonist NBQX attenuated neuroprotective effect of IPost to a level comparable with the effect of a separate application of NBQX for neurons in the CA1 and CA3 fields of the hippocampus. Conclusions . Along with other mechanisms, the activation of AMPA receptors might be implicated in the mechanisms of cerebral IPost.

Postconditioning mitigates cell death following oxygen and glucose deprivation in PC12 cells and forebrain reperfusion injury in rats.

Postconditioning mitigates ischemia-induced cellular damage via a modified reperfusion procedure. Mitochondrial permeability transition (MPT) is an important pathophysiological change in reperfusion injury. This study explores the role of MPT modulation underlying hypoxic postconditioning (HPoC) in PC12 cells and studies the neuroprotective effects of ischemic postconditioning (IPoC) on rats. Oxygen-glucose deprivation (OGD) was performed for 10 hr on PC12 cells. HPoC was induced by three cycles of 10-min reoxygenation/10-min rehypoxia after OGD. The MPT inhibitor N-methyl-4-isoleucine cyclosporine (NIM811) and the MPT inducer carboxyatractyloside (CATR) were administered to selective groups before OGD. Cellular death was evaluated by flow cytometry and Western blot analysis. JC-1 fluorescence signal was used to estimate the mitochondrial membrane potential (△Ψm ). Transient global cerebral ischemia (tGCI) was induced via the two-vessel occlusion and hypotension method in male Sprague Dawley rats. IPoC was induced by three cycles of 10-sec reperfusion/10-sec reocclusion after index ischemia. HPoC and NIM811 administration attenuated cell death, cytochrome c release, and caspase-3 activity and maintained △Ψm of PC12 cells after OGD. The addition of CATR negated the protection conferred by HPoC. IPoC reduced neuronal degeneration and cytochrome c release and cleaved caspase-9 expression of hippocampal CA1 neurons in rats after tGCI. HPoC protected PC12 cells against OGD by modulating the MPT. IPoC attenuated degeneration of hippocampal neurons after cerebral ischemia.

Tadalafil enhances the neuroprotective effects of ischemic postconditioning in mice, probably in a nitric oxide associated manner.

This study investigates the modulatory effect of tadalafil, a selective phosphodiesterase (PDE-5) inhibitor, on the neuroprotective effects of ischemic postconditioning (iPoCo) in mice. Bilateral carotid artery occlusion (BCAO) for 12 min followed by reperfusion for 24 h was employed to produce ischemia and reperfusion induced cerebral injury. Cerebral infarct size was measured using TTC staining. Memory was assessed using the Morris water maze test. Degree of motor incoordination was evaluated using inclined beam-walking, rota-rod, and lateral push tests. Brain nitrite/nitrate, acetylcholinesterase activity, TBARS, and glutathione levels were also estimated. BCAO followed by reperfusion produced a significant increase in cerebral infarct size, brain nitrite/nitrate and TBARS levels, and acetylcholinesterase activity along with a reduction in glutathione. Marked impairment of memory and motor coordination was also noted. iPoCo consisting of 3 episodes of 10 s carotid artery occlusion and reperfusion instituted immediately after BCAO significantly decreased infarct size, memory impairment, motor incoordination, and altered biochemistry. Pretreatment with tadalafil mimicked the neuroprotective effects of iPoCo. The tadalafil-induced neuroprotective effects were significantly attenuated by l-NAME, a nonselective NOS inhibitor. We concluded that tadalafil mimics the neuroprotective effects of iPoCo, probably through a nitric oxide dependent pathway, and PDE-5 could be a target of interest with respect to the neuroprotective mechanism of iPoCo.

Neuroprotection of Ischemic Postconditioning by Downregulating the Postsynaptic Signaling Mediated by Kainate Receptors

Ischemic postconditioning, a brief episode of ischemia after a prolonged ischemic insult, has been found to reduce the delayed neuronal loss after stroke. However, the mechanisms underlying such endogenous neuroprotective strategy remain obscure. In this study, we try to explore the excitatory postsynaptic signal events associated with neuroprotective effect of ischemic postconditioning. Global cerebral ischemia was induced for 15 minutes by the 4-vessel occlusion method in male Sprague-Dawley rats. Ischemic postconditioning was conducted 10 minutes later by a single reocclusion for 3 minutes. A severe global cerebral ischemia after 5 days of reperfusion destroyed almost all hippocampal CA1 pyramidal neurons. A brief ischemic postconditioning robustly reduced the neuronal loss after ischemia. Preadministration of phosphoinositide 3-kinase inhibitor LY294002 blocked the neuroprotection of postconditioning, whereas mitogen-activated protein kinase kinase 1 inhibitor PD98059 had no effect. Ischemic postconditioning significantly increased the Akt phosphorylation (Ser473). In addition, postconditioning not only perturbed the binding of postsynaptic density protein-95 with glutamatergic kainate receptor subunit 2 and mixed lineage kinase 3 but also suppressed the downstream activation of mixed lineage kinase 3, mitogen-activated protein kinase kinase 7, and c-Jun N-terminal kinase 3. LY294002, but not PD98059, abolished the postconditioning-induced decreases in the assembly of glutamatergic kainate receptor subunit 2-postsynaptic density protein-95-mixed lineage kinase 3 complex and in the mixed lineage kinase 3-c-Jun N-terminal kinase 3 signaling. Akt inhibitor IV, a specific Akt inhibitor, showed the same effects as LY294002. Ischemic postconditioning protects neurons against stroke by attenuating the postsynaptic glutamatergic kainate receptor subunit 2-postsynaptic density protein-95-mixed lineage kinase 3-c-Jun N-terminal kinase 3 signal cascade via phosphoinositide 3-kinase-Akt pathway.

Abstract WP117: Ischemic Post-conditioning Partially Reverses Cell Cycle Re-entry Following Transient Ischemia/Reperfusion Injury

Background and purpose —The neuroprotective effects of ischemic postconditioning (IPostC) are well established but the underlying mechanism is still unknown. We began to explore this critical gap in the field by genomic comparison of ischemic rat cortex following transient middle cerebral artery occlusion (tMCAO) alone or tMCAO+IPostC. Methods and Results —Microarray analysis revealed that tMCAO-induced transcriptional changes in 40 cell cycle regulators were ameliorated by IPostC, suggesting that IPostC reversed neuronal cell cycle re-entry. RT-PCR, immunoblotting, and immunofluorescence were subsequently used to quantify or localize the cell proliferation marker proliferating cell nuclear antigen (PCNA), positive and negative cell cycle regulators, and related signaling molecules. As expected, IPostC reversed the rise in mRNA levels of positive cell cycle regulators cdcb1, cdk1, cdca2, cadca3, and cdca7. Elevations in cyclin D1 and neuronal cyclin A2 were similarly inhibited. tMCAO-induced phosphorylation of extracellular signal-regulated kinase (p-ERK), glycogen synthase kinase-3β (p-GS3K3β), and cAMP response element binding protein (p-CREB) were also all depressed by IPostC. Furthermore, p-ERK colocalized with neuronal cyclin A2. Conclusions —The present study demonstrates the potent inhibitory effect of IPostC treatment on tMCAO-induced cell-cycle reentry and on ERK/CREB and GSK3β/CREB signaling. Because neuronal cell cycle re-entry is pro-apoptotic, these findings lend insight into potential mechanisms underlying neuroprotection with IPostC.

The neuroprotective effect of ischemic postconditioning and citicoline against cerebral ischemia-reperfusion injury in mongolian gerbils

Background. Ischemic postconditioning (IPostC) - a phenomenon of cerebral infarct volume reduction after a series of short ischemic stimuli in the early reperfusion period after prolonged ischemia. The mechanisms of IPostC remain poorly studied; in addition, the effects of combined application of pharmacological neuroprotection and IPostC are not investigated that precludes the translation of IPostC to the clinical practice. Objective. To assess the neuroprotective effect of IPostC and neuroprotective agent citicoline, and the effect of their combined application in the experimental model of global cerebral ischemia-reperfusion. Materials and methods. Transient global cerebral ischemia was induced by bilateral occlusion of the common carotid arteries in Mongolian gerbils for 7 minutes. IPostC was induced by three 15-s episodes of reperfusion/reocclusion. Citicoline was administered intraperitoneally at a dose of 500 mg/kg 2 min after the start of ischemia. In a separate series of experiments, the combined application of IPostC and citicoline was used. The number of viable neurons in the CA1 hippocampus was used as an and-point of neuroprotection. Results. In the experimental groups using IPostC, citicoline administration at a dose of 500 mg/kg, as well as their combined use a significant (p < 0,05) increase in the number of viable pyramidal neurons in the CA1 hippocampus was observed when compared with control group. The combined use of citicoline and IPostC did not provide an additional increase in the number of viable neurons. Conclusions. The combined use of citicoline and IPostC resulted in the expression of a significant neuroprotective phenotype, but its magnitude was not different from that in the groups of isolated citicoline and IPostC use.

Postconditioning by mild hypoxic exposures reduces rat brain injury caused by severe hypoxia

A potent neuroprotective effect of ischemic postconditioning has previously been described using cerebral artery occlusion but this is not a practical therapeutic option. The present study has been performed to determine whether postconditioning by mild episodes of hypobaric hypoxia (hypoxic postconditioning, HP) can reduce post-hypoxic brain injury in rats. Male Wistar rats were submitted to severe hypobaric hypoxia (180Torr, 3h) followed by HP (360Torr, 2h, 3 trials spaced at 24h) starting either 3h (early HP) or 24h (delayed HP) after severe hypoxia. The structural and functional brain injury was assessed by a complex of histological techniques, behavioral methods, and by testing the functions of the hypothalamic–pituitary–adrenal axis (HPA). It was found that early and delayed HP considerably attenuated post-hypoxic injury, reducing pyknosis, hyperchromatosis, and interstitial brain edema, as well as the rates of neuronal loss in hippocampus and neocortex. Delayed HP produced prominent anxiolytic effect on rat behavior, preventing development of post-hypoxic anxiety. Both modes of HP had beneficial effect on the functioning of HPA, but only delayed HP normalized completely the baseline HPA activity and its reactivity to stress. The results obtained demonstrate that postconditioning by using repetitive episodes of mild hypobaric hypoxia may provide a powerful neuroprotective procedure that can be easily adopted for clinical practice and recommended as a research tool for identification of endogenous mechanisms involved in post-ischemic neuroprotection.

Inhibition of mitochondrial permeability transition pore opening contributes to the neuroprotective effects of ischemic postconditioning in rats

Ischemic postconditioning (IPost) has been shown to attenuate cerebral ischemia–reperfusion injury. However, the mechanism remains elusive. Because opening of the mitochondrial permeability transition pore (MPTP) is a crucial determinant of cell death after ischemia–reperfusion, we hypothesized that the neuroprotective effect of IPost may be associated with inhibition of MPTP opening. In part 1 of this study, pentobarbital-anesthetized rats subjected to middle cerebral artery occlusion for 90min, followed by reperfusion for 72h, were assigned to receive one of the following treatments: three cycles of IPost (15s each), intracerebroventricular injection of saline (control), administration of the MPTP inhibitor cyclosporin A (CsA) (2μmol/L, 15μL) or its vehicle alcohol, administration of the MPTP opener atractyloside (Atr) (2mmol/L, 15μL), or IPost plus CsA/Atr treatment. Neurological deficit scores (NDS) and infarct volumes were assessed. Mitochondrial ultrastructure and swelling were also examined after reperfusion. In part 2, control and IPost groups underwent ischemia (90min) and reperfusion (15min). CsA and Atr groups were treated as described in part 1. Brain mitochondria were isolated after reperfusion and MPTP activity was evaluated. IPost or CsA treatment significantly improved NDS and reduced infarction volume, while Atr reversed the neuroprotective effects of IPost, and attenuated the decrease in mitochondrial swelling induced by IPost or CsA. Thus, inhibiting MPTP opening may play a crucial role in the neuroprotective effects of IPost, which may have potential clinical value against cerebral ischemia–reperfusion injury.

Modulation of Neuroprotective Effect of Ischemic Post-Conditioning by Dichlorobenzamil a Na+/Ca2+ Exchanger Inhibitor in Mice

The present study has been designed to investigate the modulatory role of dichlorobenzamil a Na(+)/Ca(2+) exchanger inhibitor in ischemic post-conditioning-induced neuroprotection in mice. Bilateral carotid artery occlusion followed by reperfusion, produced a significant rise in cerebral infarct size along with impairment of memory and motor coordination. Ischemic post-conditioning involving three episodes of 10 s carotid artery occlusion with intermittent reperfusion of 10 s proceeding prolonged ischemic insult of 12 min, produced a significant decrease in cerebral infarct size along with reversal of ischemia-reperfusion-induced impairment of memory and motor coordination. Ischemic post-conditioning-induced neuroprotective effects were significantly abolished by pretreatment with dichlorobenzamil (DBZ 0.5, 1 mg/kg, intraperitoneal) a Na(+)/Ca(2+) exchange inhibitor. It may be concluded that Na(+)/Ca(2+) exchanger possibly plays an important role in ischemic post-conditioning-induced neuroprotection.

The enhanced protective effect of postconditioning by 17β‐estradiol after transient focal cerebral ischemia in mice

The aim of this study was to explore the enhancement of 17β‐estradiol (E2) in the neuroprotective effect of ischemic postconditioning (PostC) following transient focal cerebral ischemia in mice. Adult female mice were ovariectomied two weeks before 30 min transient focal cerebral ischemia induced by right middle cerebral artery occlusion. E2 or vehicle (sesame oil) was administered following the reperfusion immediately. PostC consisted of 3 cycles of 15 s reperfusion/15 s occlusion on right common carotid artery. Brains were obtained 24 h, 2 d, 3 d and 5 d after permanent reperfusion. Nissl staining was used to measure the infarct sizes and neurological scores were evaluated. Levels of TNF‐α, IL‐1β and IL‐6, and expressions of caspase 3 and Bcl‐2 in brain homogenates were analyzed. We found that E2 enhanced the infarct‐limiting effect and improvement of neurological score by PostC. Levels of TNF‐α, IL‐1β and IL‐6 in brain homogenates from E2 treated postconditioned mice were significantly lower than those in vehicle treated group. Expression of caspase 3 and Bcl‐2 in brains of postconditioned mice with E2 treatment showed lower levels than that of vehicle treated group. These results provide first evidence that E2 enhances the neuroprotective effect of ischemic PostC at least via antagonizing inflammation and apoptosis induced by cerebral ischemia and reperfusion.