alpha-Phenyl-n-tert-butyl-nitrone attenuates hypoxic-ischemic white matter injury in the neonatal rat brain

White matter of the neonatal brain is highly sensitive to hypoxic-ischemic insult. The susceptibility of premature oligodendrocytes (OLs) to free radicals (FRs) produced during hypoxia-ischemia (HI) has been proposed as one of the mechanisms involved. To test this hypothesis, and to further investigate if the FR scavenger alpha-phenyl-N-tert-butyl-nitrone (PBN) attenuates hypoxic-ischemic white matter damage (WMD), postnatal day 4 (P4) SD rats were subjected to bilateral common carotid artery ligation (BCAL), followed by 8% oxygen exposure for 20 min. Pathological changes were evaluated on P6 and P9, 2 and 5 days after the HI insult. HI caused severe WMD including rarefaction, necrosis and cavity formation in the corpus callosum, external and internal capsule areas. OL injury was evidenced by degeneration of O4 positive OLs on P6. Disrupted myelination was verified by decreased immunostaining of myelin basic protein (MBP) on P9. Axonal injury was demonstrated by increased amyloid precursor protein (APP) immunostaining on both P6 and P9. Two lipid peroxidation end products, malondialdehyde (MDA) and 4-hydroxynonenal (4-HNE), showed a one-fold elevation within 1-24 h following HI. 4-HNE immunostaining was found to specifically localize in the white matter area. Furthermore, pyknotic O4+ OLs were double-labeled with 4-HNE. These findings suggest that FRs are involved in the pathogenesis of neonatal WMD. PBN (100 mg/kg, i.p.) treatment alleviated the pathological changes of WMD following HI. It improved the survival of O4 positive OLs, attenuated hypomyelination and reduced axonal damage. PBN treatment also decreased the brain concentration of MDA/4-HNE and positive 4-HNE staining in the white matter area. These findings indicate that in the current WMD model, PBN protects both OLs and axons, the two main components in the white matter, from neonatal HI insult. FR scavenging appears to be the primary mechanism underlying its neuroprotective effect.     

A rat model of severe neonatal hypoxic-ischemic brain injury.

BACKGROUND AND PURPOSE: Perinatal hypoxic-ischemic brain injury is a common problem with severe neurological sequelae. In this report we describe in detail a simple model of hypoxia-ischemia in the neonatal rat that gives rise to severe neocortical infarction and to selective hippocampal neuronal necrosis. METHODS: Seven-day-old Simonsen Wistar rat pups underwent bilateral carotid artery ligation under methoxyflurane anesthesia and, after a 4 to 6-hour recovery, were exposed to 60 minutes of hypoxia (6.5% O2); they were perfusion-fixed 3 days later for histological study. Brain temperature was monitored throughout this treatment. RESULTS: We found that 64 +/- 3% of neocortex above the rhinal sulcus was infarcted; this infarction was evenly distributed through the cerebral hemispheres. In the hippocampus, neuronal necrosis was selective for the internal (hilar) layers of granule cells of the dentate gyrus, with relative sparing of CA1 pyramidal cells. In addition, brain temperature was tightly controlled throughout the experimental manipulations. CONCLUSIONS: The present model is easy and sensitive and provides an infarct of sufficient severity and homogeneity to make it well suited for pharmacological and biochemical studies directed toward therapeutic amelioration and mechanisms of hypoxic-ischemic brain damage, respectively. In addition, the pattern of damage in the hippocampus is quite different from that seen in the adult brain, which should be helpful in studying the ontogeny of selective vulnerability.     

New oligodendrocytes are generated after neonatal hypoxic-ischemic brain injury in rodents

Neonatal hypoxic-ischemic (HI) white matter injury is a major contributor to chronic neurological dysfunction. Immature oligodendrocytes (OLGs) are highly vulnerable to HI injury. As little is known about in vivo OLG repair mechanisms in neonates, we studied whether new OLGs are generated after HI injury in P7 rats. Rats received daily BrdU injections at P12-14 or P21-22 and sacrificed at P14 to study the level of cell proliferation or at P35 to permit dividing OLG precursors to differentiate. In P14 HI-injured animals, the number of BrdU+ cells in the injured hemisphere is consistently greater than controls. At P35, sections were double-labeled for BrdU and markers for OLGs, astrocytes, and microglia. Double-labeled BrdU+/myelin basic protein+ and BrdU+/carbonic anhydrase+ OLGs are abundant in the injured striatum, corpus callosum, and the infarct core. Quantitative studies show four times as many OLGs are generated from P21-35 in HI corpora callosa than controls. Surprisingly, the infarct core contains many newly generated OLGs in addition to hypertrophied astrocytes and activated microglia. These glia and non-CNS cells may stimulate OLG progenitor proliferation or induce their migration. At P35, astrogliosis and microgliosis are dramatic ipsilaterally but only a few microglia and some astrocytes are BrdU+. This finding indicates microglial and astrocytic hyperplasia occurs shortly after HI but before the P21 BrdU injections. Although the neonatal brain undergoes massive cell death and atrophy the first week after injury, it retains the potential to generate new OLGs up to 4 weeks after injury within and surrounding the infarct.     

Erythropoietin exerts neuroprotective effect in neonatal rat model of hypoxic-ischemic brain injury

Hypoxic-ischemic encephalopathy seen in survivors of perinatal asphyxia is a frequently encountered and a major clinical problem for which there is currently no effective treatment. Hematopoietic neuroprotective agents, such as erythropoietin (EPO) may rescue neurons from cell death in this setting. EPO is a cytokine hormone that has neuroprotective effect in vitro and in vivo. In this study, we evaluated the effect of posthypoxic EPO administration in an animal model of neonatal hypoxic-ischemic injury. Our results show that a single intracerebroventricular injection of EPO immediately after hypoxic-ischemic insult in neonatal rat model of hypoxic-ischemia reduced the extent of hypoxic-ischemic brain damage. The mean infarct volume assessed 7 days after hypoxia was significantly smaller in EPO-treated group than in the control group. These findings suggest that EPO may provide benefit after hypoxic-ischemic events in the developing brain, a major contributor to static encephalopathy and cerebral palsy.     

Neuroprotective effects of sodium orthovanadate after hypoxic-ischemic brain injury in neonatal rats

Sodium orthovanadate (SOV), a competitive inhibitor of protein tyrosine phosphatases, is neuroprotective in adult animals following an ischemic event. The present study evaluated whether SOV might be protective in a rat pup hypoxic-ischemic (HI) model. Seven-day-old rat pups had the right carotid artery permanently ligated followed by 140 min of hypoxia (8% oxygen). SOV 1.15, 2.3, 4.6, 9.2 or 18.4 mg/kg and vehicle were administered by i.p. injection at 5 min after reoxygenation. Brain damage was evaluated by weight loss of the right hemisphere at 22 days after hypoxia and by gross and microscopic morphology. SOV lowered blood glucose at doses of 1.15, 2.3 and 4.6 mg/kg and induced toxic effects at 9.2 mg/kg. The doses of 2.3 and 4.6 mg/kg of SOV significantly reduced brain weight loss (p < 0.05), but treatment with 1.15 or 9.2 mg/kg did not. SOV 4.6 mg/kg also improved the histopathologic score and diminished the HI induced reduction of Akt and ERK-1/2 phosphorylation in the cortex (p < 0.05) and increased the density of BrdU-positive cells in the subventricular zone (p < 0.01). In conclusion, SOV has neuroprotective effects in the neonatal rat HI model partially mediated by activating Akt and ERK-1/2 pathways.     

Minocycline markedly protects the neonatal brain against hypoxic-ischemic injury

Hypoxic-ischemic brain injury in the perinatal period is a major cause of morbidity and mortality. Presently, there are no proven effective therapies with which to safeguard the human neonatal brain against this type of injury. Minocycline, a semisynthetic tetracycline, has been shown to be neuroprotective in certain adult ischemic injury/stroke and neurodegenerative disease models. However, minocycline's neuroprotective effects have not been assessed after insults to the neonatal brain. We now report that minocycline administered either immediately before or immediately after a hypoxic-ischemic insult substantially blocks tissue damage in a rodent model of neonatal hypoxic-ischemic brain injury. Minocycline treatment prevents the formation of activated caspase-3, a known effector of apoptosis, as well as the appearance of a calpain cleaved substrate, a marker of excitotoxic/necrotic cell death. To our knowledge, this is the first report of a systemic treatment that can be administered after a hypoxic-ischemic insult, which provides robust, nearly complete neuroprotection to the developing brain. Our data suggest that minocycline or a related neuroprotective tetracycline may be a candidate to consider in human clinical trials to protect the developing brain against hypoxic-ischemic-induced damage.     

The pentose phosphate pathway and pyruvate carboxylation after neonatal hypoxic-ischemic brain injury

The neonatal brain is vulnerable to oxidative stress, and the pentose phosphate pathway (PPP) may be of particular importance to limit the injury. Furthermore, in the neonatal brain, neurons depend on de novo synthesis of neurotransmitters via pyruvate carboxylase (PC) in astrocytes to increase neurotransmitter pools. In the adult brain, PPP activity increases in response to various injuries while pyruvate carboxylation is reduced after ischemia. However, little is known about the response of these pathways after neonatal hypoxia-ischemia (HI). To this end, 7-day-old rats were subjected to unilateral carotid artery ligation followed by hypoxia. Animals were injected with [1,2-¹³C]glucose during the recovery phase and extracts of cerebral hemispheres ipsi- and contralateral to the operation were analyzed using ¹H- and ¹³C-NMR (nuclear magnetic resonance) spectroscopy and high-performance liquid chromatography (HPLC). After HI, glucose levels were increased and there was evidence of mitochondrial hypometabolism in both hemispheres. Moreover, metabolism via PPP was reduced bilaterally. Ipsilateral glucose metabolism via PC was reduced, but PC activity was relatively preserved compared with glucose metabolism via pyruvate dehydrogenase. The observed reduction in PPP activity after HI may contribute to the increased susceptibility of the neonatal brain to oxidative stress.     

HIF-1alpha inhibition ameliorates neonatal brain injury in a rat pup hypoxic-ischemic model

In this study, we tested if caspase-3 inhibition decreased ischemia-induced Aβ elevation by reducing β-secretase (BACE1) activity. Changes in caspase-3, Aβ and BACE1 levels were detected in rat striatum on different days after middle cerebral artery occlusion using immunostaining. We found that the positive labeled cells of activated caspase-3, Aβ, and BACE1 were significantly and time-dependently increased in the ipsilateral striatum. The results of Western blotting and RT-PCR showed that caspase-3 inhibitor Z-DEVD-FMK reduced BACE1 mRNA and protein levels, and inhibited its protease activity, thereby decreasing the amount of APP C99 and Aβ in ischemic brains. Moreover, Z-DEVD-FMK reduced BACE1 and GFAP double-labeled cells, but not GFAP protein levels or GFAP-labeled cells, in the ipsilateral striatum. Thus, we demonstrated that caspase-3 inhibition attenuated ischemia-induced Aβ formation by reducing BACE1 production and activity. This finding provides a therapeutic strategy for preventing Aβ accumulation and reducing the risk of neurodegeneration after stroke.     

Shenfu injection attenuates neonatal hypoxic-ischemic brain damage in rat

Perinatal hypoxia-ischemia remains the most important cause of brain injury in the newborn. However, there is still no effective cure for neonatal hypoxic-ischemic brain damage (HIBD). In the present study, we aimed to examine the neuroprotective effects of Shenfu injection (SFI) on HIBD of neonatal rat. Sprague–Dawley rats were divided randomly into three groups (n = 8): S group: the rats were sham operated; C group: the rats were operated for HIBD modeling and received intraperitoneal injection of saline; SFI group: the rats were operated for HIBD modeling and received intraperitoneal injection of SFI (10 ml/kg days) for 7 days. Flow cytometry analysis showed that apoptosis rate of neuron in hippocampal CAI region in SFI group was significantly less than in NC group at 3 and 7 days after HI insult (P < 0.05). Immunohistochemical staining demonstrated that Bcl-2 expression was markedly higher while Bax expression was significantly lower in SFI group than in the C group at 24, 72 h and 7 days after HI insult (P < 0.05). Our findings suggest that SFI exhibits neuroprotective effects for neonatal hypoxic-ischemic brain injury by preventing neuron apoptosis and has potential to be used in the clinical for the treatment of perinatal hypoxia-ischemia.     

Trace fear conditioning detects hypoxic-ischemic brain injury in neonatal mice

Trace fear conditioning is a well-established test for the assessment of learning deficits in rodents. The aim of this study was to determine whether hypoxia-ischemia (HI) on postnatal day 9 (P9) in mice prevents the acquisition and expression of cued and contextual fear learning in early adulthood. Brain injury was induced in mice on P9 by 30 min of HI. On P49 and P50, animals were tested for: (1) trace fear conditioning with a short delay (2 s) between a shock-paired tone plus light and shock, (2) trace fear conditioning with a longer delay (20 s) between a shock-paired tone and shock, and (3) trace fear conditioning with a 2-second delay between a shock-paired tone and shock with additional visual, olfactory and tactile contextual cues in the fear conditioning apparatus. Outcome was assessed as percent of time spent freezing during a 2-min test. Histological assessment of the hippocampus and amygdala was performed on P51 to determine the extent of HI injury. Both shock-paired tone plus light with a short delay and shock-paired tone with a short delay plus additional contextual cues enhanced tone-induced freezing behavior in a nonhandled control group, but not in the HI group. For trace fear conditioning with a 20-second delay between the tone and the shock, freezing behavior did not differ significantly between nonhandled control and HI animals. Dorsal hippocampal and amygdala volumes were smaller in the ischemic hemispheres of the HI mice that displayed impaired fear memory with shock-paired tone plus light. In summary, we have shown that trace fear conditioning is a sensitive method for detecting memory impairments in adolescent mice following mild HI injury during the neonatal period. Combining a discrete conditioned stimulus (shock-paired tone plus light) with a short trace delay was the most sensitive method for using the fear conditioning paradigm to detect mild HI damage to the hippocampus and amygdala.     

Neuroprotection of edaravone on hypoxic-ischemic brain injury in neonatal rats

Edaravone has an inhibitory effect on lipid peroxidation by scavenging free radicals and prevents vascular endothelial cell injury. We examined whether edaravone was effective on hypoxic-ischemic (HI) brain injury in immature brain or not using the Rice-Vannucci model. The initial dose, 3 mg/kg (0.05 ml) of edaravone, was injected intraperitoneally just before hypoxic exposure. Subsequently, the same dose was injected every 12 h until the animals were killed. Controls received saline injection as the same protocol. Macroscopic evaluation of brain injury revealed that the neuroprotective effect of edaravone on HI brain after 48 h post HI. TUNEL showed that edaravone injection decreased neurodegeneration. Quantitative analysis of cell death using H&E-stained 2.5 microm sections showed that there was a trend for both necrotic and apoptotic cells to decrease in edaravone injection group. Edaravone injection inhibited the release of cytochrome c from mitochondria to cytosol and caspase-3 activation in cortex and hippocampus between 24 and 168 h post HI. Our results suggest that edaravone is protective after HI insult in the immature brain by decreasing both apoptosis and necrosis and also by inhibiting mitochondrial injury.     

Minocycline worsens hypoxic-ischemic brain injury in a neonatal mouse model

Hypoxic-ischemic encephalopathy (HIE) is a leading cause of mortality and morbidity during the perinatal period, and currently no therapeutic drug is available. Minocycline, an antibiotic, has recently been shown to have neuroprotective effects distinct from its antimicrobial effect in several neurological disorders including ischemic brain injury. We examined the effect of minocycline on neonatal hypoxic-ischemic brain injury by using histologic scoring in both mouse and rat models. Mouse (C57Bl/6) and rat (SD) pups were exposed to a unilateral hypoxic-ischemic insult at 8 and 7 days of age, respectively. Minocycline hydrochloride was administered according to protocols that were reported to provide neuroprotection in adult or neonatal rats. Seven days after the insult, we examined brain injury in Nissl stained sections. Although minocycline ameliorated brain injury in the developing rat, it increased injury in the developing mouse. This detrimental effect in the mouse was consistent across different regions (cortex, striatum, and thalamus), with both single and multiple injection protocols and with both moderate and high-dose treatment (P < 0.05). The mechanism of the contrasting effects in mouse and rat is not clear and remains to be elucidated. Minocycline has been used as an antibiotic in the clinical setting for decades; therefore, it may be considered for use in infants with hypoxic-ischemic brain damage, based on prior reports of neuroprotection in the rat. However, it is important to examine this drug carefully before clinical use in human infants, taking our data in the mouse model into consideration.     

Posthypoxic glucose supplement reduces hypoxic-ischemic brain damage in the neonatal rat

We evaluated the effect of posthypoxic glucose supplement in a neonatal hypoxic-ischemic animal model. Seven-day-old rats underwent bilateral ligation of the carotid arteries, followed by exposure to an 8% oxygen atmosphere for 1 hour. The extent of hypoxic-ischemic brain damage was assessed histologically 72 hours later. Glucose load immediately after the end of the hypoxic exposure reduced the volume of neocortical infarction to 37% of the unsupplemented value, and attenuated ischemic damage in the striatum and the dentate gyrus. At the end of the hypoxic exposure, the brain level of glucose was 0.3 mmol/kg and the level of lactate 9 mmol/kg. Glucose supplement produced a rapid rise in brain glucose level to 3 to 5 mmol/kg over the next 2 hours. Lactate in both brain and plasma gradually fell toward the baseline level during the first hour of recovery. Posthypoxic glucose supplement slightly retarded lactate restitution. At any period of this neonatal model, brain lactate levels did not exceed the toxic level, which is postulated to be responsible for cerebral infarction in adult ischemic models. These results illustrate the important role of glucose in the development of neonatal hypoxic-ischemic encephalopathy and the fact that full cortical infarction can develop even if brain lactate levels are low.     

Pomegranate polyphenols and resveratrol protect the neonatal brain against hypoxic-ischemic injury

A previous study from our lab has shown that the polyphenol-rich pomegranate juice can protect the neonatal mouse brain against hypoxic-ischemic (H-I) injury when given to mothers in their drinking water. To test the hypothesis that this protection is due to the polyphenols in the juice, we studied the effects of the pomegranate polyphenol extract in the same neonatal H-I model. To further explore the role of a specific polyphenol in neonatal H-I we investigated the effects of resveratrol. The neuroprotective effects of resveratrol have been demonstrated in adult models of stroke, but had not previously been examined in neonates. We show that pomegranate polyphenols and resveratrol reduce caspase-3 activation following neonatal H-I. Resveratrol reduced caspase-3 activation when given before the injury but not when given 3 h after the injury. In addition to preventing caspase-3 activation, resveratrol also reduced calpain activation. Finally, we show that resveratrol can protect against tissue loss measured at 7 days after the injury. These and other recent findings suggest that polyphenols should be further investigated as a potential treatment to decrease brain injury due to neonatal H-I.     

Accumulation of PSA-NCAM marks nascent neurodegeneration in the dorsal hippocampus after neonatal hypoxic-ischemic brain injury in mice

Neonatal hypoxia-ischemia (nHI) disrupts hippocampal GABAergic development leading to memory deficits in mice. Polysialic-acid neural-cell adhesion molecule (PSA-NCAM) developmentally declines to trigger GABAergic maturation. We hypothesized that nHI changes PSA-NCAM abundance and cellular distribution, impairing GABAergic development, and marking nascent neurodegeneration. Cell degeneration, atrophy, and PSA-NCAM immunoreactivity (IR) were measured in CA1 of nHI-injured C57BL6 mice related to: (i) cellular subtype markers; (ii) GAD65/67 and synatophysin (SYP), pre-synaptic markers; (iii) phospho-Ser³⁹⁶Tau, cytoskeletal marker; and (iv) GAP43, axonalregeneration marker. PSA-NCAM IR was minimal in CA1 of shams at P11. After nHI, PSA-NCAM IR was increased in injured pyramidal cells (PCs), minimal in parvalbumin (PV)⁺INs, and absent in glia. PSA-NCAM IR correlated with injury severity and became prominent in perikaryal cytoplasm at P18. GAD65/67 and SYP IRs only weakly related to PSA-NCAM after nHI. Injured phospho-Ser³⁹⁶Tau⁺ PCs and PV⁺INs variably co-expressed PSA-NCAM at P40. While PCs with cytoplasmic marginalized PSA-NCAM had increased perisomatic GAP43, those with perikaryal cytoplasmic PSA-NCAM had minimal GAP43. PSA-NCAM increased in serum of nHI-injured mice. Increased PSA-NCAM is likely a generic acute response to nHI brain injury. PSA-NCAM aberrant cellular localization may aggravate neuronal degeneration. The significance of PSA-NCAM as a biomarker of recovery from nHI and nascent neurodegeneration needs further study.     

Oxypurinol administration fails to prevent hypoxic-ischemic brain injury in neonatal rats

The purpose of the present study was to determine whether oxypurinol, a xanthine oxidase inhibitor, reduces free radicals and brain injury in the rat pup hypoxic-ischemia (HI) model. Seven-day-old rat pups had right carotid arteries ligated followed by 2.5h of hypoxia (8% oxygen). Oxypurinol or vehicle was administered by i.p. injection at 5 min after reoxygenation and once daily for 3 days. Brain damage was evaluated by weight deficit of the right hemisphere at 22 days following hypoxia. Oxypurinol treatments did not reduce weight loss in the right hemisphere. Brain weight loss in the right hemisphere were -26.2+/-3.6, -15.2+/-6.9, -21.7+/-4.4, -15.8+/-5.1, and -16.7+/-3.4% in vehicle (n=33), 10 (n=17), 20 (n=16), 40 (n=15), and 135 mg/kg (n=13) oxypurinol-treated groups (p>0.05), respectively. Brain thiobarbituric acid-reacting substances (TBARS) were assessed 3 and 6h after reoxygenation. Concentrations of TBARS rose 1.5-fold due to HI. Oxypurinol did not significantly reduce an HI-induced increase in brain TBARS. Thus, xanthine oxidase may not be the primary source of oxy-radicals in pup brain and as such oxypurinol does not prevent free radical-mediated lipid peroxidation or protect against brain injury in the neonatal rat HI model.     

Impact of indolent inflammation on neonatal hypoxic-ischemic brain injury in mice

This report describes a new experimental model to evaluate the effect of a recurrent systemic inflammatory challenge, after cerebral hypoxia-ischemia in immature mice, on the progression of brain injury. Treatment with a low dose of lipopolysaccharide (E. coli O55:B5, 0.2mg/kg for 3 days, then 0.1mg/kg for 2 days) daily for 5 days after unilateral cerebral hypoxia-ischemia (right carotid ligation followed by 35min in 10% O2) in 10-day-old mice resulted in increased right forebrain tissue damage (35.6% reduction in right hemisphere volume compared to 20.6% reduction in saline-injected controls), in bilateral reductions in corpus callosum area (by 12%) and myelin basic protein immunostaining (by 19%), and in suppression of injury-related right subventricular zone cellular proliferation. The post-hypoxic-ischemic lipopolysaccharide regimen that amplified brain injury was not associated with increased mortality, nor with changes in body temperature, weight gain or blood glucose concentrations. The results of the present study demonstrate that systemic inflammation influences the evolution of tissue injury after neonatal cerebral hypoxia-ischemia and may also impair potential recovery mechanisms.     

新生小鼠缺血缺氧性脑损伤中TLR4-TRIF信号通路的表达

目的 建立新生小鼠缺血缺氧模型,探讨TLR4-TRIF信号通路中TLR4、IRF3的表达.方法 将60只7日龄C57BL/6小鼠分为假手术组、缺血缺氧1 d、2 d、3 d、4 d、7 d、建立新生小鼠缺血缺氧性脑病模型,比较鼠脑左右半球的质量,确定模型是否建立成功.以逆转录-聚合酶链反应（RT -PCR）检测大脑皮质和海马中TLR4 mRNA、IRF3m RNA的表达.结果 缺血缺氧1 d组右侧的脑质量明显重于左侧,说明缺氧缺血1 d后,右脑水肿明显.缺氧缺血组的TLR4m RNA、IRF3m RNA的表达水平均明显高于假手术组.结论 脑缺氧缺血损伤可激活TLR4,引起IRF3表达量增加.     

Preventive effects of dexamethasone on hypoxic-ischemic brain damage in the neonatal rat

To clarify the preventive effects of glucocorticoid on perinatal hypoxic-ischemic (HI) brain damage, an experiment was carried out on 4-day-old rats pretreated for 4 consecutive days with 3 different regimens; namely, a low dose dexamethasone (Dex) (0.1 mg/kg/day), a high dose Dex (0.5 mg/kg/day), and a saline administration. On the 7th postnatal day, after ligation of the left common carotid artery, the rats were exposed to 8% oxygen and decapitated on the 10th, 14th, 21st and 28th postnatal days. Ligated side brain damage was observed in 75, 7 and 3% of the rats in the saline, low and high dose Dex groups, respectively. However, a high mortality rate (42%) was noted in the high dose Dex group. The cumulative number of animals with poor outcome (death or brain damage) was 49 (80%), 13 (33%) and 24 (44%) in the saline, low and high dose Dex groups, respectively. On the 10th and 14th postnatal days, the rats in both the Dex groups showed delayed neuronal maturation and myelination in the non-ligated side motor cortex, however, these maturational differences disappeared on the 21st postnatal days. Otherwise, the number of cortical cells in both the Dex groups were significantly lower than that in the saline group on the 28th postnatal days (P < 0.05 in each). These findings suggest that the pretreatment with Dex protects the developing brain from HI injury through the suppression of the neuronal maturation. However, a decreased number of cortical cells may give rise to psychomotor retardation later.