Blood-brain Barrier Preservation Research Articles

Sestrins: A New Kid for Stroke Treatment?

The sestrin family includes several conserved stress-induced proteins that contribute to the maintenance of homeostasis, DNA stability and cell viability in response to various types of injuries. It is well established that the protective functions of AMP-dependent protein kinase (AMPK) and/or mammalian target of rapamycin (mTOR) are regulated by sestrins. Additionally, it has been revealed that sestrins are able to protect cells from oxidative stress by scavenging reactive oxygen species (ROS). The essential involvement of sestrins in mTORC1 inhibition and ROS scavenging signaling pathways, which modulate metabolism homeostasis and regulate autophagy, indicates that sestrins may serve as a potential agent for cell growth, development, metabolism, and neurodegenerative disorders. However, the potential role of sestrins in stroke has not been discussed and summarized. Based on the current understanding of sestrins, it is believed that sestrins are one of the potential endogenous protective molecules/mechanisms following cerebral stroke, which are associated with neuronal protection, neuroinflammation suppression, and blood brain barrier preservation.

Exercise Does Not Protect against Peripheral and Central Effects of a High Cholesterol Diet Given Ad libitum in Old ApoE−/− Mice

Aim: Advanced atherosclerosis increases inflammation and stroke risk in the cerebral vasculature. Exercise is known to improve cardio-metabolic profiles when associated with a caloric restriction, but it remains debated whether it is still beneficial without the dietary control. The aim of this study was to determine both the peripheral and central effects of exercise training combined with a cholesterol-rich diet given ad libitum in old ApoE−/− mice.Methods: Forty-five-weeks old obese ApoE−/− mice fed with a high cholesterol diet ad libitum were divided into Exercise-trained (EX; running wheel free access) and Sedentary (SED) groups. Insulin tolerance and brain imaging were performed before and after the twelve-weeks training. Tissue insulin resistance, oxidative stress, and inflammation markers in plasma, aorta, and brain were then assessed.Results: In EX ApoE−/− mice, no beneficial effect of exercise was observed on weight, abdominal fat, metabolic parameters, oxidative stress, or inflammation compared to SED. Despite the regular exercise training in ApoE−/− EX mice (mean of 12.5 km/week during 12 weeks), brain inflammation imaging score was significantly associated with increased blood brain barrier (BBB) leakage evaluated by imaging follow-up (r2 = 0.87; p = 0.049) with a faster evolution compared to SED ApoE−/−mice.Conclusion: We conclude that in a context of high cardio-metabolic risk, exercise does not provide any protective effect in old ApoE−/− animals under high cholesterol diet given ad libitum. Peripheral (insulin sensitivity and oxidative/inflammatory status) but also central features (BBB preservation and protection against inflammation) did not show any benefits of exercise. Indeed, there was a fast induction of irreversible brain damage that was more pronounced in exercise-trained ApoE−/− mice.

Artesunate Protected Blood-Brain Barrier via Sphingosine 1 Phosphate Receptor 1/Phosphatidylinositol 3 Kinase Pathway After Subarachnoid Hemorrhage in Rats.

Blood-brain barrier preservation plays an important role in attenuating vasogenic brain edema after subarachnoid hemorrhage (SAH). This study was designed to investigate the protective effect and mechanism of artesunate, a traditional anti-malaria drug, on blood-brain barrier after SAH. Three hundred and seventy-seven (377) male Sprague-Dawley rats were subjected to endovascular perforation model for SAH. The rats received artesunate alone or in combination with Sphingosine-1-phosphate receptor-1 (S1P1) small interfering RNA (siRNA), antagonist VPC23019, or phosphatidylinositol 3-kinase inhibitor wortmannin after SAH. Modified Garcia score, SAH grades, brain water content, Evans blue leakage, transmission electron microscope, immunohistochemistry staining, Western blot, and cultured endothelial cells were used to investigate the optimum concentration and the therapeutic mechanism of artesunate. We found that artesunate (200mg/kg) could do better in raising modified Garcia score, reducing brain water content and Evans blue leakage than other groups after SAH. Moreover, artesunate elevated S1P1 expression, enhanced phosphatidylinositol 3-kinase activation, lowered GSK-3β activation, stabilized β-catenin, and improved the expression of Claudin-3 and Claudin-5 after SAH in rats. These effects were eliminated by S1P1 siRNA, VPC23019, and wortmannin. This study revealed that artesunate could preserve blood-brain barrier integrity and improve neurological outcome after SAH, possibly through activating S1P1, enhancing phosphatidylinositol 3-kinase activation, stabilizing β-catenin via GSK-3β inhibition, and then effectively raising the expression of Claudin-3 and Claudin-5. Therefore, artesunate may be favorable for the blood-brain barrier (BBB) protection after SAH and become a potential candidate for the treatment of SAH patients.

Novel Approaches in Astrocyte Protection: from Experimental Methods to Computational Approaches.

Astrocytes are important for normal brain functioning. Astrocytes are metabolic regulators of the brain that exert many functions such as the preservation of blood-brain barrier (BBB) function, clearance of toxic substances, and generation of antioxidant molecules and growth factors. These functions are fundamental to sustain the function and survival of neurons and other brain cells. For these reasons, the protection of astrocytes has become relevant for the prevention of neuronal death during brain pathologies such as Parkinson's disease, Alzheimer's disease, stroke, and other neurodegenerative conditions. Currently, different strategies are being used to protect the main astrocytic functions during neurological diseases, including the use of growth factors, steroid derivatives, mesenchymal stem cell paracrine factors, nicotine derivatives, and computational biology tools. Moreover, the combined use of experimental approaches with bioinformatics tools such as the ones obtained through system biology has allowed a broader knowledge in astrocytic protection both in normal and pathological conditions. In the present review, we highlight some of these recent paradigms in assessing astrocyte protection using experimental and computational approaches and discuss how they could be used for the study of restorative therapies for the brain in pathological conditions.

Efficacy of prophylactic flavan-3-ol in permanent focal ischemia in 12-mo-old mice.

The consumption of flavan-3-ol-containing foods, including (-)-epicatechin (EC), has been linked to lower incidence of cardiovascular disease and stroke. We previously demonstrated nuclear transcription factor erythroid 2p45-related factor-2 (Nrf2) -dependent EC efficacy in reducing stroke-induced deficits in 2-mo-old mice; yet stroke is primarily a disease of the elderly. Because neuroinflammation, oxidative stress, and vascular dysfunction are hallmarks of aging, we tested whether Nrf2 mediates EC efficacy in aging mice through modulation of glial responses and blood brain barrier permeability. First, we compared anastomosis in naïve wild-type and C57BL/6 Nrf2(-/-) mice to identify potential differences in cerebrovascular architecture. Data showed no significant differences in the number of anastomoses or mean intersection points, indicating similar gross vascular physiology. To assess efficacy and mechanisms of protection, wild-type or Nrf2(-/-) mice were administered the minimum effective EC dose established in our previous studies before the permanent distal middle cerebral artery occlusion. Similar to previous results with young mice, 12-mo-old wild types also showed significant reductions in infarct volume (41.01 ± 29.57%) and improved performance in removing adhesive tape relative to vehicle-treated controls, whereas a trend toward protection was observed in Nrf2(-/-). However, EC did not reduce immunoreactivity for the microglia/macrophage marker anti-ionized calcium-binding adapter molecule 1, suggesting that dampened activation/recruitment did not account for EC protection. Furthermore, there were no differences in mouse IgG extravasation or spontaneous hemorrhage between EC-treated groups. These data demonstrate that EC protection occurs independent of microglia/macrophage modulation or blood brain barrier preservation, suggesting that the glial cell responses in young mice are compensatory to another, and potentially novel, protective mechanism.

Investigation of iron's neurotoxicity during cerebral maturation in the neonatal rat model of haemolysis.

Haemolytic disease of newborns due to rhesus and AB0 incompatibility is encountered frequently in neonatal clinics and may lead to severe haemolysis. In this study, it is suggested that important amounts of iron released with haemolysis may have a toxic effect on the brain parenchymal tissue, and the severity of the toxic effect can be correlated with the maturation of the brain barrier systems. To demonstrate the accumulation and the neuro-toxic effects of free iron (Fe) in the brain an experimental haemolysis model with various maturation phases was performed. The study was composed of 48 Wistar rats with the following ages: five days old (Group A), 10 days old (Group B), and 19 days old (Group C). Each group was divided into three experimental subgroups and three control groups. Experimental groups were treated with intraperitoneal 75 mg/kg/day phenyl hydrazine hydrochloride for haemolysis. We demonstrated that the blood brain barrier (BBB) is permeable in five-day-old newborn rats and is mature in 10- and 19-day-old rats. Iron staining and neuronal damage were detected in group A and group B rats. No damage was detected in the brain tissue of group C animals. The presence of iron staining and neuronal damage in group B with mature BBB may suggest the existence of other incomplete barrier systems different from BBB that lead to iron accumulation in the brain. Blood brain barrier has a partial role in Fe transport, and the alternative barrier systems may also be involved. It could be supposed that after maturation of all barrier systems, excessive Fe penetration to the brain cannot occur. Our findings showed that the toxic amounts of iron may penetrate into the brain parenchyma of newborns despite the BBB preservation and cause neuronal damage in newborns, but the mature brain is not affected by the same magnitude blood levels.

Effects of progesterone vs. dexamethasone on brain oedema and inflammatory responses following experimental brain resection

Background: Dexamethasone (DEXA) is commonly used to reduce brain swelling during neurosurgical procedures. DEXA, however, has many side-effects that can increase the risks of post-operative complications. In contrast, progesterone (PRO) has fewer side-effects and has been found to be neuroprotective on traumatic brain injury (TBI). Whether PRO may be used as an alternative to DEXA during routine procedures has not been fully explored.Object: To compare the effects of DEXA and PRO on surgical brain injury (SBI).Methods: Seventy-five adult male Sprague Dawley rats were randomized into five groups: (1) SBI + drug vehicle (peanut oil, 1 ml kg−1); (2) SBI + DEXA (1 mg kg−1); (3) SBI + low-dose PRO (10 mg kg−1); (4) SBI + high-dose PRO (20 mg kg−1); and (5) sham SBI + drug vehicle. Magnetic resonance imaging study and assessments of brain water content (BWC), blood–brain barrier (BBB) permeability, cellular inflammatory responses and matrix metalloproteinase 9 (MMP-9) expression were conducted.Results: This model consistently resulted in increased BWC and BBB disruption. PRO reduced astrocyte and microglia responses and attenuated brain oedema with preservation of BBB. A significant down-regulation of MMP-9 expression occurred in the PRO 20 group.Conclusions: PRO is as effective as DEXA in reducing brain oedema and inflammation following SBI; 10 mg kg−1 of PRO was demonstrated to be more effective in relieving acute cellular inflammatory responses.

Neuroprotective effects of erythropoietin pretreatment in a rodent model of transient middle cerebral artery occlusion

OBJECT.: There is an unmet clinical need to develop neuroprotective agents for neurosurgical and endovascular procedures that require transient cerebral artery occlusion. The aim in this study was to explore the effects of a single dose of recombinant human erythropoietin (rhEPO) before middle cerebral artery (MCA) occlusion in a focal cerebral ischemia/reperfusion model. Twenty-eight adult male Wistar rats were subjected to right MCA occlusion via the intraluminal thread technique for 60 minutes under continuous cortical perfusion monitoring by laser Doppler flowmetry. Rats were divided into 2 groups: control and treatment. In the treated group, rhEPO (1000 IU/kg intravenously) was administered 10 minutes before the onset of the MCA ischemia. At 24-hour reperfusion, animals were examined for neurological deficits, blood samples were collected, and animals were killed. The following parameters were evaluated: brain infarct volume, ipsilateral hemispheric edema, neuron-specific enolase plasma levels, parenchyma histological features (H & E staining), Fluoro-Jade-positive neurons, p-Akt and total Akt expression by Western blot analysis, and p-Akt-positive nuclei by immunohistochemical investigation. Infarct volume and Fluoro-Jade staining of degenerating neurons in the infarct area did not vary between groups. The severity of neurological deficit (p < 0.001), amount of brain edema (78% reduction in treatment group, p < 0.001), and neuron-specific enolase plasma levels (p < 0.001) were reduced in the treatment group. Perivascular edema was histologically less marked in the treatment group. No variations in the expression or localization of p-Akt were seen. Administration of rhEPO before the onset of 60-minute transient MCA ischemia protected the brain from this insult. It is unlikely that rhEPO pretreatment leads to direct neuronal antiapoptotic effects, as supported by the lack of Akt activation, and its benefits are most probably related to an indirect effect on brain edema as a consequence of blood-brain barrier preservation. Although research on EPO derivatives is increasing, rhEPO acts through distinct neuroprotective pathways and its clinical safety profile is well known. Clinically available rhEPO is a potential therapy for prevention of neuronal injury induced by transitory artery occlusion during neurovascular procedures.

PHA-543613 Preserves Blood–Brain Barrier Integrity After Intracerebral Hemorrhage in Mice

Blood-brain barrier disruption and consequent vasogenic edema formation codetermine the clinical course of intracerebral hemorrhage (ICH). This study examined the effect of PHA-543613, a novel α7 nicotinic acetylcholine receptor agonist, on blood-brain barrier preservation after ICH. Male CD-1 mice, subjected to intrastriatal blood infusion, received PHA-543613 alone or in combination with α7 nicotinic acetylcholine receptor antagonist methyllycaconitine or phosphatidylinositol 3-kinase inhibitor wortmannin. PHA-543613 alone, but not in combination with methyllycaconitine or wortmannin, inhibited glycogen synthase kinase-3β, thus, stabilizing β-catenin and tight junction proteins, which was paralleled by improved blood-brain barrier stability and ameliorated neurofunctional deficits in ICH animals. PHA-543613 preserved blood-brain barrier integrity after ICH, possibly through phosphatidylinositol 3-kinase-Akt-induced inhibition of glycogen synthase kinase-3β and β-catenin stabilization.

Abstract WMP33: Synergistic Effect Of Dehydroascorbic Acid, Glutathione And Melatonin On Brain Endothelium Protection In Ischemic Rats

Introduction: Stroke (cerebral ischemia) is the second leading cause of death worldwide. Antioxidant strategies have been used successfully to diminish ischemic cerebral tissue damage in animals but the utility of a pharmacological agent as a clinically relevant therapeutic strategy may depend, in part, on its ability to cross the BBB. To optimally improve the therapeutic window this present study investigated whether the intravenous administration of dehydroascorbic acid (DHA), glutathione (GSH) and melatonin potentially elevates the level of cerebroprotection. Methods: Male rats were subjected to 60 min middle carotid artery occlusion (MCAO) using an intraluminal filament. Single dose of DHA (100mg/kg/ml), glutathione (500 mg/kg/ml) and melatonin (10 mg/kg/ml) was administered intravenously immediately after MCAO at a time interval of 5 min each. Western blotting was performed to delineate the effects of antioxidant treatment on the proliferation and neurogenesis in the ischemic region. We also examined the neuroprotective dosing regimen of triple antioxidants treatment and its effects on the functional outcomes following cerebral ischemia. Results: Triple antioxidant treatment significantly reduced the infarct volume and improved behavioral performance evidenced by TTC staining and neurological test score results. The significant increase in the claudin-5 and occludine expressions in the antioxidant treatment group depicts the preservation of blood-brain barrier (BBB) integrity via a reduction in the enormous amount of stroke-induced free radical production. Moreover, treatment with triple antioxidants after stroke dramatically enhanced endogenous neurogenesis (doublecortin positive) and cell proliferation (ki67 positive) in the peri-infarct regions. These outcomes were in the favor of rescuing the damaged neurons at the injury site defining the neuroprotective effects of triple antioxidants treatment following experimental stroke. Conclusion: The results from this study demonstrate that triple antioxidant treatment prevents brain edema, BBB hyperpermeability, and tight junction proteins disruption, possibly through its antioxidant effects thereby offering optimal neuroprotection in rat MCAO model.

Blood-Brain Barrier Integrity and Glial Support: Mechanisms that can be Targeted for Novel Therapeutic Approaches in Stroke

The blood-brain barrier (BBB) is a critical regulator of brain homeostasis. Additionally, the BBB is the most significant obstacle to effective CNS drug delivery. It possesses specific charcteristics (i.e., tight junction protein complexes, influx and efflux transporters) that control permeation of circulating solutes including therapeutic agents. In order to form this "barrier," brain microvascular endothelial cells require support of adjacent astrocytes and microglia. This intricate relationship also occurs between endothelial cells and other cell types and structures of the CNS (i.e., pericytes, neurons, extracellular matrix), which implies existence of a "neurovascular unit." Ischemic stroke can disrupt the neurovascular unit at both the structural and functional level, which leads to an increase in leak across the BBB. Recent studies have identified several pathophysiological mechanisms (i.e., oxidative stress, activation of cytokine-mediated intracellular signaling systems) that mediate changes in the neurovascular unit during ischemic stroke. This review summarizes current knowledge in this area and emphasizes pathways (i.e., oxidative stress, cytokine-mediated intracellular signaling, glial-expressed receptors/targets) that can be manipulated pharmacologically for i) preservation of BBB and glial integrity during ischemic stroke and ii) control of drug permeation and/or transport across the BBB. Targeting these pathways present a novel opportunity for optimization of CNS delivery of therapeutics in the setting of ischemic stroke.

Role of the pituitary–adrenal axis in granulocyte-colony stimulating factor-induced neuroprotection against hypoxia–ischemia in neonatal rats

Several reports indicate that the activity of the hypothalamic–pituitary–adrenal axis (HPA) is increased after a brain insult and that its down-regulation can improve detrimental outcomes associated with ischemic brain injuries. Granulocyte-colony stimulating factor (G-CSF) is a neuroprotective drug shown in the naïve rat to regulate hormones of the HPA axis. In this study we investigate whether G-CSF confers its neuroprotective properties by influencing the HPA response after neonatal hypoxia–ischemia (HI). Following the Rice–Vannucci model, seven day old rats (P7) were subjected to unilateral carotid ligation followed by 2.5h of hypoxia. To test our hypothesis, metyrapone was administered to inhibit the release of rodent specific glucocorticoid, corticosterone, at the adrenal level. Dexamethasone, a synthetic glucocorticoid, was administered to agonize the effects of corticosterone. Our results show that both G-CSF and metyrapone significantly reduced infarct volume while dexamethasone treatment did not reduce infarct size even when combined with G-CSF. The protective effects of G-CSF do not include blood brain barrier preservation as suggested by the brain edema results. G-CSF did not affect the pituitary released adrenocorticotropic hormone (ACTH) levels in the blood plasma at 4h, but suppressed the increase of corticosterone in the blood. The administration of G-CSF and metyrapone increased weight gain, and significantly reduced the Bax/Bcl-2 ratio in the brain while dexamethasone reversed the effects of G-CSF. The combination of G-CSF and metyrapone significantly decreased caspase-3 protein levels in the brain, and the effect was antagonized by dexamethasone. We report that G-CSF is neuroprotective in neonatal HI by reducing infarct volume, by suppressing the HI-induced increase of the Bax/Bcl-2 ratio, and by decreasing corticosterone in the blood. Metyrapone was able to confer similar neuroprotection as G-CSF while dexamethasone reversed the effects of G-CSF. In conclusion, we show that decreasing HPA axis activity is neuroprotective after neonatal HI, which can be conferred by administering G-CSF.

Role of c‐Jun N‐terminal kinase in early brain injury after subarachnoid hemorrhage

The c-Jun N-terminal kinase (JNK) is induced by cerebral ischemia and injurious blood components acutely after subarachnoid hemorrhage (SAH). We hypothesized that inhibition of JNK will prevent damage to the neurovascular unit in the early brain injury period after SAH. Ninety-nine male SD rats (300-350 g) were randomly assigned to sham, SAH, and SAH treated with JNK inhibitor groups. SAH was induced by endovascular perforation. The JNK inhibitor SP600125 was administered intraperitoneally at 1 hr before and 6 hr after SAH. At 24 hr after SAH, we observed increased phosphorylation of JNK and c-Jun. Signs of neurovascular damage were observed in the hemorrhagic brains; these included the increases of aquaporin (AQP)-1 expression and brain water content as well as enhanced matrix metalloproteinase (MMP)-9 activity, vascular collagen IV loss, increased VEGF tissue level, and Evans blue extravasation. The appearances of cleaved caspase-3 expression, TUNEL-positive cells, and apoptotic morphology in cerebral tissues were associated with neurological deficit after SAH. JNK inhibition prevented c-Jun phosphorylation and suppressed AQP1, MMP-9, VEGF, and caspase-3 activation, with concomitant diminution of neuronal injury, blood-brain barrier preservation, reduced brain swelling, and improved neurological deficit in rats after SAH. This study demonstrates a multitude of beneficial effects of JNK inhibition, including protection of the neurovascular unit in early brain injury after SAH.

Reduction in levels of matrix metalloproteinases and increased expression of tissue inhibitor of metalloproteinase—2 in response to mild hypothermia therapy in experimental stroke

Mild hypothermia is a robust neuroprotectant, and the results of prospective clinical trials have indicated that it may improve neurological outcome in certain instances. One aspect of this protection has been associated with the prevention of blood-brain barrier (BBB) disruption. Matrix metalloproteinases (MMPs) have been implicated in BBB disruption because they can degrade the extracellular matrix. In this study the authors explored the relationship between hypothermia and MMPs and whether BBB preservation resulting from mild hypothermia therapy is due to alterations in MMP expression. Rats were subjected to middle cerebral artery occlusion for 2 hours; the animals were maintained in a state of normothermia or mild hypothermia (33 degrees C) immediately after the onset of ischemia. The animals' brains were collected 2, 6, and 24 hours after ischemia began. Contrast-enhanced T1-weighted magnetic resonance imaging was performed at 24 hours to assess the extent of BBB disruption. Consistent with prior reports, areas of BBB disruption detected on T1-weighted images were smaller in the brains of rats maintained in a state of hypothermia (normothermia group 8.6 +/- 3% of the brain; hypothermia group 0.2 +/-0.1% of the brain; p < 0.01). Expression of both MMP-2 and MMP-9 at the transcriptional and translational levels was reduced in hypothermic brains at 6 hours and 24 hours after ischemic injury. Matrix metalloproteinase-9 was primarily localized to cells of monocytic origin but was also observed in neurons and astrocytes. Matrix metalloproteinase-2 was found in some neurons and astrocytes but not in inflammatory cells. In addition, hypothermia increased the levels of the endogenous MMP inhibitor, tissue inhibitor of metalloproteinases-2. The authors conclude that mild hypothermia attenuates BBB disruption, decreases MMP expression, and suppresses MMP activity.

Blood-brain barrier preservation in the in vitro isolated guinea pig brain preparation.

The morphofunctional preservation of the blood-brain barrier (BBB) was evaluated in the isolated guinea pig brain maintained in vitro by arterial perfusion. Electron microscopy evaluation after 5 hr in vitro demonstrated that cerebral capillaries and BBB specializations in this preparation retain features compatible with structural integrity. BBB-impermeable and -permeable atropine derivatives arterially perfused to antagonize carbachol-induced fast oscillatory activity confirmed the functional preservation of the BBB in vitro. To study BBB function further, changes in extracellular K+ concentration during arterial perfusion of a high-K+ solution were measured with K+-sensitive electrodes positioned in the cortex and, as control, at the brain venous outlet, where the solution perfused through the brain arterial system was collected. After 5 hr in vitro, the [K+](o) values measured during high-K+ perfusion in the piriform and entorhinal cortices were 5.02 +/- 0.17 mM (mean +/- SE) and 5.2 +/- 0.21 mM, respectively (n = 6). Coperfusion of the high-K+ solution with the Na+/K+ pump blocker ouabain (10 microM; n = 4) induced consistently spreading depression preceded by a rise in [K+](o). Finally, sporadic, isolated spots of extravasation of the fluorescent marker fluorescein isothiocyanate (FITC)-dextran preferentially circumscribed to deep cortical layers was observed in brains perfused with FITC-dextran after 5 hr in vitro. The study demonstrates that the in vitro isolated guinea pig brain is viable for studying cerebrovascular interactions and BBB permeability of compounds active in the central nervous system.

Vertical osteotomy for extending the mandibular rami in vertical posterior insufficiency

Several reports indicate that the activity of the hypothalamic–pituitary–adrenal axis (HPA) is increased after a brain insult and that its down-regulation can improve detrimental outcomes associated with ischemic brain injuries. Granulocyte-colony stimulating factor (G-CSF) is a neuroprotective drug shown in the naïve rat to regulate hormones of the HPA axis. In this study we investigate whether G-CSF confers its neuroprotective properties by influencing the HPA response after neonatal hypoxia–ischemia (HI). Following the Rice–Vannucci model, seven day old rats (P7) were subjected to unilateral carotid ligation followed by 2.5 h of hypoxia. To test our hypothesis, metyrapone was administered to inhibit the release of rodent specific glucocorticoid, corticosterone, at the adrenal level. Dexamethasone, a synthetic glucocorticoid, was administered to agonize the effects of corticosterone. Our results show that both G-CSF and metyrapone significantly reduced infarct volume while dexamethasone treatment did not reduce infarct size even when combined with G-CSF. The protective effects of G-CSF do not include blood brain barrier preservation as suggested by the brain edema results. G-CSF did not affect the pituitary released adrenocorticotropic hormone (ACTH) levels in the blood plasma at 4 h, but suppressed the increase of corticosterone in the blood. The administration of G-CSF and metyrapone increased weight gain, and significantly reduced the Bax/Bcl-2 ratio in the brain while dexamethasone reversed the effects of G-CSF. The combination of G-CSF and metyrapone significantly decreased caspase-3 protein levels in the brain, and the effect was antagonized by dexamethasone. We report that G-CSF is neuroprotective in neonatal HI by reducing infarct volume, by suppressing the HI-induced increase of the Bax/Bcl-2 ratio, and by decreasing corticosterone in the blood. Metyrapone was able to confer similar neuroprotection as G-CSF while dexamethasone reversed the effects of G-CSF. In conclusion, we show that decreasing HPA axis activity is neuroprotective after neonatal HI, which can be conferred by administering G-CSF.

The influence of total body hyperthermia on brain haemodynamics and blood-brain barrier in dogs.

This study was designed to examine the influence of total body hyperthermia (TBHT) using an extracorporeal circuit with a heat exchanger on the cerebral blood flow (CBF), intracranial pressure (ICP), brain tissue pH, cerebral autoregulation and blood-brain barrier (BBB) permeability in dogs. The rectal temperature of the dow was raised to 41.5 degrees C, maintained at 41.5-42.0 degrees C for 2 hours (HT period) and then reduced to normothermia by cooling. Regional CBF was measured by the hydrogen clearance method before heating, during the HT period and after cooling. ICP and brain tissue pH were monitored during the TBHT treatment. Autoregulation of the CBF during the HT period was assessed by measuring the regional CBF and the ICP in a state of induced hypo- or hypertension. The influence of TBHT on BBB permeability was examined using an immunohistochemical technique. The regional CBF increased from 38.1 +/- 6.5 (mean +/- SD) to 49.1 +/- 9.8 ml/100 g/min and the ICP from 10.3 +/- 4.2 to 16.8 +/- 3.4 mmHg when TBHT was raised. These returned to normal values after cooling. The regional CBF and the ICP changed in parallel with drug-induced changes of mean arterial blood pressure during the HT period. These changes suggest that autoregulation of the CBF is paralysed during the HT period. Brain tissue pH decreased rapidly when the rectal temperature exceeded 41.0 degrees C. The pH was 7.18 +/- 0.05 during the HT period and was relatively stable. The pH returned to a normal value after cooling. Immunopositive stain for albumin was not observed in heated brain tissue except for the normally leaky pineal gland and the choroid plexus, indicating preservation of BBB during TBHT. These results suggest that brain oedema may occur easily due to paralysed cerebral autoregulation when the arterial blood pressure fluctuates excessively, so arterial blood pressure must be controlled strictly during TBHT.

Effects of intracarotid hyperosmolar mannitol in triethyl tin (TET)-induced rat brain edema — preservation of blood-brain barrier (BBB) in TET edema

The effect of intracarotid hyperosmolar mannitol on the blood-brain barrier (BBB) in triethyl tin (TET)-induced rat brain edema was examined by using intravenous Evans blue (EB, MW 68,000) and Adriamycin (ADM, MW 580) as tracers. Three ml of 1.4 M mannitol solution were administered through the right carotid artery during 45 s for BBB opening. The barrier was opened for 60–120 minutes and then re-established. The reversibility was preserved in TET-treated ratsand controls. In fact, the intravenous injection of EB stained both TET-treated and non-treated cerebral hemispheres with mannitol-induced transient BBB disruption, but not without BBB disruption. BBB was resistant to both high and low molecular weight substances in TET-induced edema. The importance of this hyperosmotic studies provides the evidence for normal BBB function in TET-induced brain edema.