Cerebral Ischemic Neuronal Damage Research Articles

Experience with therapeutic hypothermia in the treatment of ischemic stroke

Patients and methods. Craniocerebral hypothermia (CCH) was carried out in 25 patients (18 men and 7 women aged 53—82 years) in the acute period (first 72 hours) of ischemic stroke verified by brain computed tomography or magnetic resonance imaging. An ATG-01 Russian apparatus was used for CCH, which can long maintain scalp temperature at +3±2 °С. Results. There was regression of neurological deficit apparently due to reduced brain edema, lowered intracranial pressure, as well as blood flow improvement in the great vessels of the affected hemisphere, which was indirectly suggestive of better blood supply to the penumbral area and decreased lesion volume. By and large, lower brain temperature is able to restrict the development of all basic pathogenetic mechanisms of neuronal damage in cerebral ischemia. It is stated that further investigations are required to prove the efficiency of CCH.

RETRACTED ARTICLE: Honokiol suppresses the development of post-ischemic glucose intolerance and neuronal damage in mice

Honokiol, a constituent of Magnolia obovata, has various pharmacological effects, including protection against cerebral ischemia. However, few studies have been conducted to evaluate the possible neuroprotective effects of honokiol against cerebral ischemia. We recently reported that cerebral ischemic neuronal damage could be triggered by glucose intolerance that develops after the onset of ischemic stress (i.e., post-ischemic glucose intolerance). In addition, suppression of post-ischemic glucose intolerance significantly ameliorated ischemic neuronal damage. Here, we investigated the effects of honokiol on the development of post-ischemic glucose intolerance and neuronal damage. Mice were subjected to middle cerebral artery occlusion (MCAO) for 2h. The development of post-ischemic glucose intolerance on day 1 and neuronal damage on day 3 after MCAO were significantly reduced by intraperitoneal administration of honokiol (10mg/kg) compared with the vehicle-treated group. Honokiol did not affect serum insulin or adiponectin levels. However, honokiol significantly decreased the expression of phosphoenolpyruvate carboxykinase and increased the expression of 5'-AMP-activated protein kinase (AMPK) on day 1 after MCAO, compared with the vehicle-treated MCAO group. The results of this study suggest that honokiol could prevent post-ischemic glucose intolerance in an AMPK-dependent manner, which may be involved in the neuroprotective effects of honokiol against cerebral ischemia.

ChemInform Abstract: Effectiveness of Metformin in Prevention of Development of Hyperglycemia and Neuronal Damage Caused by Ischemic Stress

AbstractReview: 13 refs.

Involvement of Matrix Metalloproteinase-Mediated Proteolysis of Neural Cell Adhesion Molecule in the Development of Cerebral Ischemic Neuronal Damage

Neural cell adhesion molecule (NCAM) is a membrane protein abundantly expressed in the central nervous system. Recently, it has been reported that dysfunction of NCAM is linked to human brain disorders. Furthermore, NCAM is one of the proteolysis targets of matrix metalloproteinase (MMP), whose activation is implicated in neuronal damage. The aim of this study was to elucidate the involvement of MMP-mediated proteolysis of NCAM in the development of ischemic neuronal damage. Male ddY and MMP-9 knockout (KO) C57BL/6J mice were subjected to 2 h of middle cerebral artery occlusion (MCAO). In MCAO model mice, development of infarction and behavioral abnormality were clearly observed on days 1 and 3 after MCAO. Protein levels of MMP-2 and MMP-9 were significantly increased on days 1 and 3 after MCAO. In addition, full-length NCAM (180 kDa) was significantly decreased, but its metabolite levels increased on day 1 by ischemic stress per se. NCAM small interfering RNA significantly increased the neuronal damage induced by MCAO. MMP inhibition or MMP-9 gene KO attenuated the infarction, behavioral abnormalities, and decrease of NCAM (180 kDa) observed after MCAO in mice. The present findings clearly suggest that MMP-2/MMP-9-mediated NCAM proteolysis is implicated in the exacerbation of ischemic neuronal damage.

Effectiveness of Metformin in Prevention of Development of Hyperglycemia and Neuronal Damage Caused by Ischemic Stress

Hyperglycemia is a known exacerbating factor in ischemic stroke. It has been reported that hyperglycemia and post-ischemic glucose intolerance can develop after stroke and be involved in the development of neuronal damage, as we described previously. Here, we focus on the effectiveness of metformin, a hypoglycemic drug, in preventing the development of neuronal damage using the middle cerebral artery occlusion (MCAO) model mice. 5'-AMP-activated protein kinase (AMPK) is a serine/threonine kinase that plays a key role in the hypoglycemic effects of metformin. Recently, it has been reported that centrally activated AMPK is involved in the development of ischemic neuronal damage, while the effect of peripherally activated AMPK on ischemic neuronal damage is not known. In the liver, AMPK activity was not affected by MCAO, while the administration of intraperitoneal metformin (250 mg/kg), an AMPK activator, significantly activated hepatic AMPK and suppressed both the development of post-ischemic glucose intolerance and ischemic neuronal damage without alteration of central AMPK activity. On the other hand, the administration of intracerebroventricular metformin (100 µg/mouse) significantly exacerbated the development of neuronal damage observed on day 1 after MCAO. These effects were significantly blocked by compound C, a specific AMPK inhibitor. These results suggest that central AMPK is activated by ischemic stress per se, although peripheral AMPK is not altered. Furthermore, the regulation of post-ischemic glucose intolerance by metformin-induced activation of peripheral AMPK contributes to the reduction of cerebral ischemic neuronal damage.

Thrombin potently enhances swelling‐sensitive glutamate efflux from cultured astrocytes

High concentrations of thrombin (Thr) have been linked to neuronal damage in cerebral ischemia and traumatic brain injury. In the present study we found that Thr markedly enhanced swelling-activated efflux of (3)H-glutamate from cultured astrocytes exposed to hyposmotic medium. Thr (0.5-5 U/mL) elicited small (3)H-glutamate efflux under isosmotic conditions and increased the hyposmotic glutamate efflux by 5- to 10-fold, the maximum effect being observed at 15% osmolarity reduction. These Thr effects involve its protease activity and are fully mimicked by SFFLRN, the synthetic peptide activating protease-activated receptor-1. Thr potentiation of (3)H-glutamate efflux was largely dependent on a Thr-elicited increases in cytosolic Ca(2+) (Ca(2+) (i)) concentration ([Ca(2+)](i)). Preventing Ca(2+) (i) rise by treatment with EGTA-AM or with the phospholipase C blocker U73122 reduced the Thr-increased glutamate efflux by 68%. The protein kinase C blockers Go6976 or chelerythrine reduced the Thr effect by 19%-22%, while Ca/calmodulin blocker W7 caused a 63% inhibition. In addition to this Ca(2+)-sensitive pathway, Thr effect on glutamate efflux also involved activation of phosphoinositide-3 kinase (PI3K), since it was reduced by the PI3K inhibitor wortmannin (51% inhibition). Treating cells with EGTA-AM plus wortmannin essentially abolished Thr-dependent glutamate efflux. Thr-activated glutamate release was potently inhibited by the blockers of the volume-sensitive anion permeability pathway, NPPB (IC(50) 15.8 microM), DCPIB (IC(50) 4.2 microM), and tamoxifen (IC(50) 6.6 microM. These results suggest that Thr may contribute to the excitotoxic neuronal injury by elevating extracellular glutamate release from glial cells. Therefore, this work may aid in search of neuroprotective strategies for treating cerebral ischemia and brain trauma.

Reduction of Cerebral Infarction in Stroke-Prone Spontaneously Hypertensive Rats by Statins Associated With Amelioration of Oxidative Stress

This study aimed to clarify the effect of statins on spontaneous stroke and to examine the antioxidative effect in artificial transient middle cerebral artery occlusion (tMCAO). Stroke-prone spontaneous hypertensive rats (SHR-SP) were treated with pitavastatin, atorvastatin, simvastatin, or vehicle for 4 weeks. Physiological parameters, serum lipids, and infarct volumes were examined. The markers for oxidative stresses on lipids and DNA were immunohistochemically detected in vehicle-treated or simvastatin-treated SHR-SP with tMCAO. Atorvastatin and simvastatin decreased infarct volumes, with simvastatin most effective. Simvastatin significantly reduced immunoreactivities for oxidative stress markers for lipids and DNA in neurons after tMCAO. The results suggest that the antioxidative properties of statins may be implicated in their beneficial effects against neuronal damage in cerebral ischemia.

Sevoflurane-induced preconditioning protects against cerebral ischemic neuronal damage in rats

In the present study, we tested the ability of sevoflurane to induce early and late preconditioning against ischemic neuronal injury using an in vivo model of global cerebral ischemia in the rat. Seven-minute global ischemia was induced by cardiac arrest, followed by resuscitation and recovery for 7 days. Hippocampal slices were then prepared and the amplitude of extracellularly recorded, orthodromically evoked, CA1 population spikes (neuronal function) was quantified. Rats were preconditioned for 30 min with 1.0 minimum alveolar concentration (MAC) of sevoflurane once or on 4 consecutive days, 15 min (single exposure, early) or 24 h (four exposures, late preconditoning) prior to cardiac arrest. After early or late preconditioning, sevoflurane reduced ischemic neuronal damage from 43 ± 3% [sham rats, (mean ± SEM)] to 30 ± 3% and 35 ± 4%, respectively. Histopathology demonstrated a preserved morphology of the CA1 region of preconditioned rats, whereas pyknosis was present in control and sham-treated rats. Sevoflurane-induced preconditioning confers neuroprotection during an early as well as late time window.

Neuroprotective effect of geranylgeranylacetone, a noninvasive heat shock protein inducer, on cerebral infarction in rats

The present study evaluated the neuroprotective effect of geranylgeranylacetone (GGA), which is known as an antiulcer agent and more recently as a heat shock protein (HSP) inducer, against cerebral infarction induced by permanent left middle cerebral artery (MCA) occlusion. GGA was given orally in various regimens prior to MCA occlusion in rats. Pretreatment with a single oral GGA dose (800 mg/kg) 48 h before ischemia significantly attenuated cerebral infarction volume (81.7 ± 18.4 mm 3 versus 369.1 ± 70.2 mm 3; P < .01). A significant increase in HSP70 immunoreactivity was found in the neocortex in GGA-treated animals with or without ischemia. Pretreatment with a single oral dose of GGA provides an important tool for exploring the mechanisms of neuroprotection against cerebral ischemic neuronal damage.

Astrocytic gap junctions composed of connexin 43 reduce apoptotic neuronal damage in cerebral ischemia.

Astrocytes may play a vital role in neuroprotection by providing energy substrates to neurons and regulating the concentration of K+ and neurotransmitters through gap junctions. Connexin 43 (Cx43) is one of the major gap junction proteins in astrocytes. We have shown that, after focal stroke, heterozygote Cx43 null (Cx43+/-) mice exhibited larger infarction volumes than wild-type (Cx43+/+) mice. We explored the underlying mechanism by which gap junctional intercellular communication influences astrocytic activation and neuroprotection in ischemia. Both Cx43+/- and Cx43+/+ mice underwent right side permanent middle cerebral artery occlusion (MCAO). Mice were prepared by transcardial perfusion, and at 24 hours and 4 days after surgery, brains were prepared for immunohistochemistry or Western blot analysis. Four days after MCAO, Cx43+/- mice showed severe apoptosis in the penumbral lesion compared with Cx43+/+ mice. The level of caspase-3 was significantly higher in the stroke lesion of Cx43+/- mice than in Cx43+/+ mice. Four days after MCAO, Cx43+/- mice showed a significantly larger infarct volume but a smaller area of astrogliosis than did Cx43+/+ mice. The penumbra of Cx43+/- mice showed an increased level of Cx30 compared with Cx43+/+ mice. Gap junctions may play an important role in astrocytic activation. Reactive astrocytes may reduce neuronal apoptosis under ischemia by regulating extracellular conditions through their gap junction.

Lactate, glucose and energy metabolism in the ischemic brain (Review)

For many decades, lactate was considered to be an end product of anaerobic glycolysis in mammalian tissues with no other function in metabolism. As determination of lactate level became a routine in hospital blood work, fluctuations in its levels were associated with situations other than oxygen lack. It was just a matter of time before elevated lactate blood levels emerged as a 'red flag' for potential malaise and frequently was blamed as the of it. Lactate and its accompanying acidosis are still considered today to be major contributors to selective neuronal damage in cerebral ischemia despite the emergence of alternative, more compelling postulates as to the causes of this damage. The attitude, especially among clinicians, has been difficult to change despite recent findings that strongly indicate lactate to be a possible beneficial intermediate in brain energy metabolism. This review revisits briefly the annals that brought about the lingering negative attitude toward lactate and expands on the more recent findings and debates that have illuminated this monocarboxylate in a different, more positive light.

Na+-K+-Cl- cotransporter in rat focal cerebral ischemia.

In cultured neurons, the authors previously demonstrated that the Na+-K+-Cl- cotransporter is significantly stimulated by elevated extracellular potassium and glutamate, which are important factors in cerebral ischemic damage. These findings led the authors to hypothesize that stimulation of the cotransporter after ischemia might result in Na+, K+, and Cl- influx, and might contribute to neuron damage. In the current study, the authors investigated such a role of the Na+-K+-Cl- cotransporter in focal cerebral ischemia. Cerebral ischemia was induced by 2-hour occlusion of the left middle cerebral artery (MCA) and 24-hour reperfusion in male spontaneously hypertensive rats (SHRs). Immunocytochemical staining and immunoblotting revealed an up-regulation of expression of the cotransporter protein in neurons in cortex at 24 hours of reperfusion. Artificial cerebral spinal fluid (aCSF) or 100 micromol/L bumetanide (a cotransporter inhibitor) in aCSF were continuously microdialyzed through a microdialysis probe into left cortices throughout 2-hour MCA occlusion and 24-hour reperfusion. Compared with the aCSF-treated group, infarction volume was significantly reduced in the bumetanide-treated group (25%, P < 0.05). In addition, brain water content in the bumetanide-treated brains was decreased by 70% (P < 0.05). These results strongly suggest that the Na+-K+-Cl- cotransporter may play an important role in cerebral ischemic neuronal damage.

Gene therapy for cerebrovascular disease.

To review the principles of and the experimental and clinical results of gene therapy for cerebrovascular disease. Literature review. Vectors for gene transfer into the brain or into the cerebral vasculature include naked plasmid deoxyribonucleic acid, cationic liposomes, and viruses such as adenovirus, retrovirus, adeno-associated virus, and herpes simplex virus. Experiments using these vectors showed that intra- or perivascular application to systemic arteries can lead to transfection and expression of a foreign transgene in the adventitia and the endothelium. Intrathecal administration can lead to transfection and foreign transgene expression in leptomeningeal cells as well as in fibroblasts of blood vessel adventitia. Biological effects demonstrated thus far include increased nitric oxide production by transfection of cerebral arterial adventitia with adenovirus expressing nitric oxide synthase. Adenoviruses carrying foreign genes have been used to decrease neuronal damage in cerebral ischemia and to decrease blood pressure in spontaneously hypertensive rats. Vectors and therapeutic applications for gene therapy are evolving rapidly. Gene therapy for cerebrovascular disease is likely to have clinical application in the near future and will have a major impact on neurosurgery. Neurosurgeons will need to be aware of the literature in this area.

Albumin enhances superoxide production in cultured microglia.

Microglial activation and disruption of blood-brain barrier (BBB) are known to occur and contribute to neuronal damages in cerebral ischemic conditions and in some neurodegenerative diseases. To investigate whether a serum factor leaked out from circulation enhances microglial activation, we examined the effect of normal rat serum on superoxide (O2-) production by cultured microglia. Microglia cultured from neonatal rat brains were studied on their O2- production induced by the addition of phorbol myristate acetate by a method of acetyl-cytochrome c reduction. The O2- production was significantly increased by the addition of 0.01% rat serum, and the maximal enhancement was observed at about 0.1% rat serum. After the serum was fractionated using a molecular sieve membrane, we observed the enhancing effect only in a greater molecular weight fraction (>50 kDa). Furthermore, three kinds of bovine serum albumin (BSA) with different purity, and human serum and plasma albumins, also enhanced O2- production to a similar extent to that by rat serum. However, other proteins tested showed no significant effect. The enhancement of O2- production by BSA was observed dose-dependently, and the effect of 50 microg/ml of purified BSA was equivalent to that of 0.1% rat serum, suggesting that albumin itself enhances O2- production by microglia. These results imply that albumin leaked out through impaired BBB may activate microglia and that the potentiation of O2- production by albumin results in the pathogenesis of neuronal damage in cerebral ischemia and some neurodegenerative diseases.

Acetyl-L-carnitine attenuates neuronal damage in gerbils with transient forebrain ischemia only when given before the insult.

The underlying mechanisms leading to neuronal damage in cerebral ischemia are multifactoral. In this study, we evaluated the neuroprotective effects of acetyl-L-carnitine, a medication that may enhance metabolic recovery after cerebral ischemia. The 5-minute transient forebrain ischemia model in gerbils was used. Acetyl-L-carnitine was given 30 minutes before the insult in one set of animals and 30 minutes after the insult in a second set of animals with histological evaluation at 7 days (Group A) and 28 days (Group B). Damage assessment was done using a 4-point damage score and Mann-Whitney U test was used for statistical analysis. Compared to the controls, there was significant protection in the cerebral cortex, hippocampus and the striatum in animals treated with the medication before the insult in Group A and Group B. Post-ischemic therapy showed little evidence of neuronal protection in either group. Behavioral tests in the Group B animals showed no significant differences between the treated or the saline controls. Our study shows, that pre-ischemic treatment with acetyl-L-carnitine results in neuronal protection. This may have clinical significance in situations (such as bypass surgery) where treatment could be initiated prior to the insult.

Immunocytochemical and electron microscopic evidence of neuronal damage in cerebral ischemia

Immunocytochemical and electron microscopic evidence of neuronal damage in cerebral ischemia

The glutamate antagonist MK‐801 reduces focal ischemic brain damage in the rat

Excessive activation of the N-methyl-D-aspartate (NMDA) subtype of glutamate receptor has been implicated in the sequence of neurochemical events that results in irreversible neuronal damage in cerebral ischemia. The effects of the NMDA antagonist (+)-5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5,10-imine maleate (MK-801) upon the amount of ischemic brain damage has been assessed quantitatively in the lightly anesthetized rat. Focal cerebral ischemia was produced by the permanent occlusion of one middle cerebral artery (MCA), and the animals were killed 3 hours after the arterial occlusion. MK-801 (0.5 mg/kg) was administered intravenously either 30 minutes prior to MCA occlusion or 30 minutes after the induction of ischemia. Pretreatment with MK-801 reduced the volume of ischemic damage both in the cerebral cortex (by 38% compared with untreated rats with MCA occlusion; p less than 0.01) and in the caudate nucleus (by 18% compared with controls; p less than 0.05). Treatment with MK-801, initiated 30 minutes after MCA occlusion, reduced the volume of ischemic damage in the cerebral cortex (by 52% compared with controls; p less than 0.01). The volume of ischemic damage in the caudate nucleus was minimally influenced by MK-801 treatment initiated after MCA occlusion. The antiischemic effects of MK-801 were readily demonstrable despite the hypotension that MK-801 induced in rats anesthetized with halothane (0.5%), nitrous oxide (70%), and oxygen (30%). The potency of MK-801 in reducing ischemic brain damage, even when administered after the induction of ischemia, highlights the potential use of NMDA receptor antagonists for the treatment of focal cerebral ischemia in humans.