Effects Of Global Cerebral Ischemia Research Articles

Global cerebral ischemia in adolescent male Long Evans rats: Effects of vanillic acid supplementation on stress response, emotionality, and visuospatial memory

The developmental period is critical in delineating plastic response to internal and external events. However, neurobehavioural effects of global cerebral ischemia (GCI) in the maturing brain remain largely unknown. This study characterised the effects of GCI experienced at puberty on adulthood (1) hippocampus CA1 neuronal damage, (2) cognitive and emotional impairments, and (3) glucocorticoid receptor (GR) expression. Effects of adolescent exposure to the phenol vanillic acid (VA) on post-ischemic outcomes were also determined. Male Long Evans rats (n = 35) were supplemented for 21 consecutive days (postnatal days 33–53) with VA (91 mg/kg) or nut paste vehicle (control) prior to a 10-min GCI or sham surgery. As adults, rats were tested in the Open Field Test (OFT), Elevated-Plus Maze (EPM), and Barnes Maze (BM). GR expression was determined in the basolateral amygdala (BLA), CA1, and paraventricular nucleus (PVN), and brain injury assessed via CA1 neuronal density. Adolescent GCI exposure induced extensive hippocampal CA1 injury, which was not prevented by VA supplementation. Behaviourally, GCI increased EPM exploration while having no impact on spatial memory. VA intake increased OFT peripheral exploration. Notably, while no delayed changes in CA1 and PVN GR immunoreactivity were noted, both treatments separately increased BLA GR expression when compared with sham-nut paste rats. Age at GCI occurrence plays a critical role on post-ischemic impairments. The observation of minimal functional impairments despite important CA1 neuronal damage supports use of compensatory mechanisms. Our findings also show daily VA supplementation during adolescence to have no protective effects on post-ischemic outcomes, contrasting adult intake.

Heterogeneity in brain distribution of activated microglia and astrocytes in a rat ischemic model of Alzheimer's disease after 2 years of survival.

The present study was designed to follow neuroinflammation after ischemic brain injury in the long-term survival rat model. Immunohistochemistry was performed 2 years after 10 min global brain ischemia due to cardiac arrest. For the visualization of the cellular inflammatory reaction microglial marker Iba1 and astrocyte marker GFAP were used. In post-ischemic animals our study revealed significant activation of astrocytes in all tested brain regions (hippocampal CA1 and CA3 areas and dentate gyrus, motor and somatosensory cortex, striatum and thalamus), while microglial activation was only found in CA1 and CA3 areas, and the motor cortex. In the specifically sensitive brain areas microglia and astrocytes showed simultaneously significant activation, while in the resistant brain areas only astrocytes were activated. Thus, there was clear evidence of less intensive neuroinflammation in brain areas resistant to ischemia. Such neuroinflammatory processes are backed by microglia and astrocytes activity even up to 2 years after ischemia-reperfusion brain injury. Our study thus revealed a chronic effect of global cerebral ischemia on the neuroinflammatory reaction in the rat brain even 2 years after the insult.

Stearic acid methyl ester affords neuroprotection and improves functional outcomes after cardiac arrest

Cardiac arrest causes neuronal damage and functional impairments that can result in learning/memory dysfunction after ischemia. We previously identified a saturated fatty acid (stearic acid methyl ester, SAME) that was released from the superior cervical ganglion (sympathetic ganglion). The function of stearic acid methyl ester is currently unknown. Here, we show that SAME can inhibit the detrimental effects of global cerebral ischemia (i.e. cardiac arrest). Treatment with SAME in the presence of asphyxial cardiac arrest (ACA) revived learning and working memory deficits. Similarly, SAME-treated hippocampal slices after oxygen-glucose deprivation inhibited neuronal cell death. Moreover, SAME afforded neuroprotection against ACA in the CA1 region of the hippocampus, reduced ionized calcium-binding adapter molecule 1 expression and inflammatory cytokines/chemokines, with restoration in mitochondria respiration. Altogether, we describe a unique and uncharted role of saturated fatty acids in the brain that may have important implications against cerebral ischemia.

Cdh1 overexpression improves emotion and cognitive-related behaviors via regulating hippocampal neuroplasticity in global cerebral ischemia rats

Post-stroke survivors exhibited cognitive deficits and performed emotional impairment. However, the effect of global cerebral ischemia on standard behavioral measures of emotionality and underlying mechanism remain largely unknown. Our previous work identified that down-regulation of Cdh1 contributed to ischemic neuronal death in rat, thus we hypothesized that Cdh1 exerts a role in emotionality after cerebral ischemia, and we investigated the effect of Cdh1 overexpression on neurogenic behaviors and possible mechanisms in transient global cerebral ischemia reperfusion (tGCI/R) rats. A series of behavioral tests were used to evaluate emotion and cognitive related behaviors, and molecular biological techniques were employed to investigate hippocampal neuroplasticity. The results showed that tGCI/R rats displayed anxiety- and depression-like behaviors and a certain degree of cognitive impairment, and these abnormal behaviors accompanied with a loss of hippocampal synapses and dendritic spines, disruption of dendrite arborization and decline in the level of GAP-43, synaptophysin, synapsin and PSD-95. However, Cdh1 overexpression improved negative emotionality, ameliorated cognitive deficits, rescued hippocampal synapses loss, prevented dendritic network disorganization, and increased the level of synaptic-associated proteins after tGCI/R. Taken together, these findings suggest that Cdh1 overexpression exerts a neuroprotective effect by regulating hippocampal neuroplasticity thus improving negative emotionality and cognitive deficits after tGCI/R.

Mild hypothermia inhibits the Notch 3 and Notch 4 activation and seizure after stroke in the rat model

Ischemic brain injury is an important cause for seizure. Mild hypothermia of the brain or the whole body is an effective way to remit the post-stroke seizure. Our previous study revealed an implication of Notch 1 and 2 in the post-stroke seizure. This study further investigated the involvement of Notch 3 and 4 in post-stroke seizure and the effect of mild hypothermia on these two factors. A global cerebral ischemia (GCI) model was conducted in Sprague Dawley rats. Seizure activity was evaluated by the frequency of seizure attacks, seizure severity scores, and seizure discharges. Seizures were frequently occurred in the first and the second 24 h after GCI, however active whole-body cooling (mild hypothermia) and DAPT (Notch inhibitor) injection into the hippocampus, alone or in combination, alleviated seizure activity after GCI. Immunohistochemistry and Western blot assays revealed the up-regulation of Notch intracellular domain (NICD) 3 and 4 in the cerebral cortex and hippocampus following GCI, but mild hypothermia and DAPT inhibited the up-regulation of NICD 3 and 4. NF-κB, PPARα, PPARγ, cyclin D1, Sox2 and Pax6 are associated with the pathogenesis of diverse type of seizures. GCI induced NF-κB, cyclin D1, and Pax6 activity, but suppressed PPARγ. These effects of GCI were abolished by both mild hypothermia and DAPT treatment. This indicated the implication of Notch signaling in the effects of GCI. Collectively, mild hypothermia inhibits Notch 3 and Notch 4 activation and seizure after stroke in the rat model. This study adds to the further understanding of the pathogenesis of post-stroke seizures and the protective mechanism of mild hypothermia.

Calcium uptake and cytochrome c release from normal and ischemic brain mitochondria

At abnormally elevated levels of intracellular Ca2+, mitochondrial Ca2+ uptake may compromise mitochondrial electron transport activities and trigger membrane permeability changes that allow for release of cytochrome c and other mitochondrial apoptotic proteins into the cytosol. In this study, a clinically relevant canine cardiac arrest model was used to assess the effects of global cerebral ischemia and reperfusion on mitochondrial Ca2+ uptake capacity, Ca2+ uptake-mediated inhibition of respiration, and Ca2+-induced cytochrome c release, as measured in vitro in a K+-based medium in the presence of Mg2+, ATP, and NADH-linked oxidizable substrates. Maximum Ca2+ uptake by frontal cortex mitochondria was significantly lower following 10 min cardiac arrest compared to non-ischemic controls. Mitochondria from ischemic brains were also more sensitive to the respiratory inhibition associated with accumulation of large levels of Ca2+. Cytochrome c was released from brain mitochondria in vitro in a Ca2+-dose-dependent manner and was more pronounced following both 10 min of ischemia alone and following 24 h reperfusion, in comparison to mitochondria from non-ischemic Shams. These effects of ischemia and reperfusion on brain mitochondria could compromise intracellular Ca2+ homeostasis, decrease aerobic and increase anaerobic cerebral energy metabolism, and potentiate the cytochrome c-dependent induction of apoptosis, when re-oxygenated mitochondria are exposed to abnormally high levels of intracellular Ca2+.

Time-dependent effects of global cerebral ischemia on anxiety, locomotion, and habituation in rats

Although changes in emotionality represent common features of post-ischemic recovery in humans, little is known about the effects of global cerebral ischemia on standard behavioral measures of emotionality in rodents. The present study investigated anxiety, locomotor activity, and habituation in test-naïve ischemic (subjected to 10 min global ischemia) and sham-operated rats tested 1, 5, 15, and 30 days post-reperfusion in the elevated plus-maze and the open-field. Although rats tested on day 1 post-reperfusion showed increased anxiety relative to sham-operated controls, they demonstrated decreased anxiety on day 5. Anxiety levels were normal on days 15 and 30 following ischemia. Similarly, time-dependent changes in locomotor activity were observed with ischemic rats showing increased activity level on days 1, 5, and 30 post-reperfusion. Surprisingly, locomotor activity was suppressed at day 15. Habituation deficits in the open-field were apparent only on day 1 despite the lack of CA1 neuronal degeneration at this time interval. These findings suggest that both the nature and extent of the effects of global ischemia on behavioral measures of emotionality, locomotion, and habituation in rats are time-dependent.

Cerebral vasospasm after subarachnoid hemorrhage: the emerging revolution

Cerebral vasospasm is the classic cause of delayed neurological deterioration after aneurysmal subarachnoid hemorrhage, leading to cerebral ischemia and infarction, and thus to poor outcome and occasionally death. Advances in diagnosis and treatment-principally the use of nimodipine, intensive care management, hemodynamic manipulations and endovascular neuroradiology procedures-have improved the prospects for these patients, but outcomes remain disappointing. Recent clinical trials have demonstrated marked prevention of vasospasm with the endothelin receptor antagonist clazosentan, yet patient outcome was not improved. This Review considers possible explanations for this result and proposes alternative causes of neurological deterioration and poor outcome after subarachnoid hemorrhage, including delayed effects of global cerebral ischemia, thromboembolism, microcirculatory dysfunction and cortical spreading depression.

Proliferation of neural and neuronal progenitors after global brain ischemia in young adult macaque monkeys

To investigate the effect of global cerebral ischemia on brain cell proliferation in young adult macaques, we infused 5-bromo-2′-deoxyuridine (BrdU), a DNA replication indicator, into monkeys subjected to ischemia or sham-operated. Subsequent quantification by BrdU immunohistochemistry revealed a significant postischemic increase in the number of BrdU-labeled cells in the hippocampal dentate gyrus, subventricular zone of the temporal horn of the lateral ventricle, and temporal neocortex. In all animals, 20–40% of the newly generated cells in the dentate gyrus and subventricular zone expressed the neural progenitor cell markers Musashi1 or Nestin. A few BrdU-positive cells in postischemic monkeys were double-stained for markers of neuronal progenitors (class III β-tubulin, TUC4, doublecortin, or Hu), neurons (NeuN), or glia (S100β or GFAP). Our results suggest that ischemia activates endogenous neuronal and glial precursors residing in diverse locations of the adult primate central nervous system.

Effects of global cerebral ischemia and preconditioning on heat shock protein 27 immunocontent and phosphorylation in rat hippocampus

Global cerebral ischemia, with or without preconditioning, leads to an increase in heat shock protein 27 (HSP27) immunocontent and alterations in HSP27 phosphorylation in CA1 and dentate gyrus areas of the hippocampus. We studied different times of reperfusion (1, 4, 7, 14, 21 and 30 days) using 2 min, 10 min or 2+10 min of ischemia. The results showed an increase in HSP27 immunocontent of about 300% after 10 min of ischemia in CA1 and dentate gyrus. CA1, a hippocampal vulnerable area, showed an increase in HSP27 phosphorylation, parallel with immunocontent. In dentate gyrus, a resistant area, the increase in HSP phosphorylation was lower than immunocontent. After preconditioned ischemia (2+10 min), when CA1 neurons are protected to a lethal, 10 min insult, we observed an increase in HSP immunocontent and a decrease in phosphorylation in both regions of the hippocampus, suggesting that, when there is no neuronal death, HSP27 in a vulnerable area responds similarly to the resistant area. When dephosphorylated, HSP27 acts as a chaperone, protecting other proteins from denaturation. As it is markedly expressed in astrocytes, we suggest that HSP27 could be protecting hippocampal astrocytes, which could then be helping neurons to resist to the insult, maintaining tissue normal homeostasis.

Cyclic AMP Response Element Binding Protein (CREB) and CREB Binding Protein (CBP) in Global Cerebral Ischemia

Cyclic AMP (cAMP) response element binding protein (CREB) is a transcription factor that has been implicated in neuronal responses to ischemia. We examined the effect of global cerebral ischemia in the rat on the expression of CREB, its transcriptionally active phosphorylated form (pCREB), and the nuclear adaptor protein, CREB binding protein (CBP). Global ischemia induced the expression of pCREB and CBP in vulnerable neurons of the hippocampal CA1 sector. In primary cultures of murine cortical neurons subjected to hypoxia, CBP was selectively expressed in cells with morphologically intact cell nuclei, and not in cells with condensed or fragmented nuclei indicative of irreversibly damaged neurons. These results support a role for transcriptional activation by CREB and CBP in neuronal cell-survival programs following cerebral ischemia.

Activity-dependent regulation of the noradrenergic transmission in the central visual structures of rat

To investigate the effect of global cerebral ischemia on brain cell proliferation in young adult macaques, we infused 5-bromo-2′-deoxyuridine (BrdU), a DNA replication indicator, into monkeys subjected to ischemia or sham-operated. Subsequent quantification by BrdU immunohistochemistry revealed a significant postischemic increase in the number of BrdU-labeled cells in the hippocampal dentate gyrus, subventricular zone of the temporal horn of the lateral ventricle, and temporal neocortex. In all animals, 20–40% of the newly generated cells in the dentate gyrus and subventricular zone expressed the neural progenitor cell markers Musashi1 or Nestin. A few BrdU-positive cells in postischemic monkeys were double-stained for markers of neuronal progenitors (class III β-tubulin, TUC4, doublecortin, or Hu), neurons (NeuN), or glia (S100β or GFAP). Our results suggest that ischemia activates endogenous neuronal and glial precursors residing in diverse locations of the adult primate central nervous system.

Non-NMDA glutamate receptor binding in canine brain after global cerebral ischemia and reperfusion.

We employed a canine model to test the effects of global cerebral ischemia and reperfusion on binding to alpha-amino-3-hydroxy-5-methyl- 4-isoxazole proprionate (AMPA), kainate (KA), and metabotropic glutamate receptors. Ischemia was induced by 10 min of cardiac arrest, followed by restoration of spontaneous circulation for periods of 0, 0.5, 2, 4, and 24 h. Frozen sections were prepared from parietal and temporal cortex, hippocampus, and striatum, and in vitro autoradiography was performed with one of three radioligands: [3H]AMPA, [3H]KA, or [3H] glutamate (using conditions allowing specific labeling of the metabotropic binding site). In striatum, metabotropic binding was unchanged, whereas AMPA and KA binding decreased by 20-30% at 30 min postischemia, remaining depressed through 24 h. In cortex, AMPA and metabotropic binding were decreased at several time-points after ischemia and recirculation, particularly in parietal cortex, whereas KA binding was unaffected in this tissue. Binding to hippocampal regions was largely unchanged, except for a decrease in KA binding at 2 and 4 h postischemia. These findings contrast with results from parallel studies showing increased striatal binding to NMDA receptors following ischemia. Decreased binding to non-NMDA glutamate receptors in striatum and parietal cortex may serve to protect against damage mediated through these receptors.

Acute changes in intracranial pressure and pressure-volume index after forebrain ischemia in normoglycemic and hyperglycemic rats.

Hyperglycemia enhances the deleterious effect of global cerebral ischemia. One possible explanation is that increased anaerobic glycolysis leads to exaggeration of intracellular acidosis and increased postischemic edema. To examine the importance of this edema on postischemic cerebral perfusion dynamics, we measured acute changes in intracranial pressure (ICP), specific gravity, and the pressure-volume index (PVI) after forebrain ischemia in normoglycemic and hyperglycemic rats. Rats underwent 15 minutes of forebrain ischemia and 90 minutes of reperfusion. ICP and mean arterial pressure were continuously monitored. Before ischemia, rats received either saline or glucose intravenously. Ninety minutes after ischemia, the specific gravity of the neocortex was measured. In a second experiment, the PVI was measured at 20 and 60 minutes after ischemia. Preischemic ICP (mean+/-SD) was 7 +/- 1 mm Hg in both groups. A peak ICP (approximately 11 mm Hg) occurred within 15 to 20 minutes after ischemia in both groups. Between 25 and 80 minutes after ischemia, ICP was significantly but only slightly greater in hyperglycemic than in normoglycemic rats. Cerebral perfusion pressure was similar between groups and remained greater than 100 mm Hg. Specific gravity was also similar for both groups but was less than normal values. The PVI in hyperglycemic rats was lower than in normoglycemic rats, indicating reduced compliance. These findings indicate that hyperglycemia-augmented intraischemic tissue acidosis does not contribute to worsened outcome by means of compromised cerebral perfusion pressure during the early stages of recovery. Nevertheless, evidence was found for decreased cerebral compliance, indicating an effect of hyperglycemia on intracranial volume compartments other than cortical parenchyma.

Brain ischemia decreases phosphatidylcholine-phospholipase D but not phosphatidylinositol-phospholipase C in rats.

Phosphatidylcholine (PC)-phospholipase D (PLD) is an important intracellular signaling pathway in response to a variety of agonists, but little is known about the effects of brain ischemia on the PC-PLD system. We thus have examined the effects of global cerebral ischemia on PLD in rats. We have examined the effects of global ischemia (decapitation or four-vessel occlusion) on PLD and PLC activity in the membrane fraction of rat brains. We measured the PLD and PLC activity in detergent-mixed micelle assay systems using 3H-labeled exogenous substrate. The results demonstrate that basal PLD activity showed a gradual decrease with increased duration (5 to 30 minutes) of ischemia by decapitation in the hippocampus; after 30 minutes of ischemia, PLD activity was significantly decreased compared with the control. Lineweaver-Burk plots showed that the apparent Vmax value of PLD in ischemia was one half of that in the control without changes in Km value. Ischemia by decapitation significantly decreased PLD activity in the brain stem as well as the hippocampus, whereas in four-vessel occlusion study, ischemia significantly decreased PLD activity in the hippocampus but not in the brain stem. Lowered temperature (30 degrees C and 22 degrees C) during ischemic incubation did not reverse the ischemia-induced PLD activity decrease. In contrast to PLD, ischemia by decapitation had no effect on basal phosphatidylinositol-phospholipase C activity or the amount of phospholipase C beta 1 in the membrane fractions from 30-minute ischemic hippocampus by immunoblots probed with the antibody. These results suggest that PC-PLD is one of the target enzymes of ischemia; its decrease may cause a perturbation of PC hydrolysis and/or disorders of intracellular transduction of signals or choline metabolism for acetylcholine formation in brain.

Ischemia/reperfusion-induced changes in membrane fluidity characteristics of brain capillary endothelial cells and its prevention by liposomal-incorporated superoxide dismutase

The effect of global cerebral ischemia and reperfusion on cerebral capillary endothelial cell membrane fluidity was examined using electron paramagnetic resonance techniques following 8 minutes of global ischemia and 15 minutes of blood reperfusion. The luminal surface of the cerebral vasculature was perfused with a series of doxyl stearic acid reporters (5-, 12-, 16-doxyl stearic acid) which differ in the site of attachment of the nitroxide free radical on the fatty acid chain. Each doxyl stearic acid reports on membrane fluidity characteristics from different depths within the membrane. Ischemia/reperfusion produced a membrane ordering that was markedly dependent on intramembrane location, and was consistent with changes previously associated with lipid peroxidation. The effect of ischemia/reperfusion on membrane fluidity was maximal in the membrane environment reported by 12-doxyl stearic acid (12-DS). The utilization of a liposomal system was shown to enhance superoxide dismutase delivery to cerebral tissues as well as attenuating the change in membrane order seen following reperfusion-induced lipid peroxidation.