Maternal environmental enrichment protects neonatal brains from hypoxic-ischemic challenge by mitigating brain energetic dysfunction and modulating glial cell responses.

There is evidence that maternal milieu and changes in environmental factors during the prenatal period may exert a lasting impact on the brain health of the newborn, even in case of neonatal brain hypoxia-ischemia (HI). The present study aimed to investigate the effects of maternal environmental enrichment (EE) on HI-induced energetic and metabolic failure, along with subsequent neural cell responses in the early postnatal period. Male Wistar pups born to dams exposed to maternal EE or standard conditions (SC) were randomly divided into Sham-SC, HI-SC, Sham-EE, and HI-EE groups. Neonatal HI was induced on postnatal day (PND) 3. The Na+,K+-ATPase activity, mitochondrial function and neuroinflammatory related-proteins were assessed at 24 h and 48 h after HI. MicroPET-FDG scans were used to measure glucose uptake at three time points: 24 h post-HI, PND18, and PND24. Moreover, neuronal preservation and glial cell responses were evaluated at PND18. After HI, animals exposed to maternal EE showed an increase in Na+,K+-ATPase activity, preservation of mitochondrial potential/mass ratio, and a reduction in mitochondrial swelling. Glucose uptake was preserved in HI-EE animals from PND18 onwards. Maternal EE attenuated HI-induced cell degeneration, white matter injury, and reduced astrocyte immunofluorescence. Moreover, the HI-EE group exhibited elevated levels of IL-10 and a reduction in Iba-1 positive cells. Data suggested that the regulation of AKT/ERK1/2 signaling pathways could be involved in the effects of maternal EE. This study evidenced that antenatal environmental stimuli could promote bioenergetic and neural resilience in the offspring against early HI damage, supporting the translational value of pregnancy-focused environmental treatments.

Effect of environmental enrichment on behavioral and morphological outcomes following neonatal hypoxia-ischemia in rodent models: A systematic review and meta-analysis.

Neonatal hypoxia-ischemia (HI) is a major cause of mortality and morbidity in newborns and, despite recent advances in neonatal intensive care, there is no definitive treatment for this pathology. Once preclinical studies have shown that environmental enrichment (EE) seems to be a promising therapy for children with HI, the present study conducts a systematic review and meta-analysis of articles with EE in HI rodent models focusing on neurodevelopmental reflexes, motor and cognitive function as well as brain damage. The protocol was registered a priori at PROSPERO. The search was conducted in PubMed, Embase and PsycINFO databases, resulting in the inclusion of 22 articles. Interestingly, EE showed a beneficial impact on neurodevelopmental reflexes (SMD= -0.73, CI= [-0.98; -0.47], p< 0.001, I2= 0.0%), motor function (SMD= -0.55, CI= [-0.81; -0.28], p< 0.001, I2= 62.6%), cognitive function (SMD= -0.93, CI= [-1.14; -0.72], p< 0.001, I2= 27.8%) and brain damage (SMD= -0.80, CI= [-1.03; -0.58], p< 0.001, I2= 10.7%). The main factors that potentiate EE positive effects were enhanced study quality, earlier age at injury as well as earlier start and longer duration of EE exposure. Overall, EE was able to counteract the behavioral and histological damage induced by the lesion, being a promising therapeutic strategy for HI.

Differential glucose and beta-hydroxybutyrate metabolism confers an intrinsic neuroprotection to the immature brain in a rat model of neonatal hypoxia ischemia.

Neonatal hypoxia ischemia (HI) is the main cause of newborn mortality and morbidity. Preclinical studies have shown that the immature rat brain is more resilient to HI injury, suggesting innate mechanisms of neuroprotection. During neonatal period brain metabolism experience changes that might greatly affect the outcome of HI injury. Therefore, the aim of the present study was to investigate how changes in brain metabolism interfere with HI outcome in different stages of CNS development. For this purpose, animals were divided into 6 groups: HIP3, HIP7 and HIP11 (HI performed at postnatal days 3, 7 and 11, respectively), and their respective shams. In vivo [18F]FDG micro positron emission tomography (microPET) imaging was performed 24 and 72 h after HI, as well as ex-vivo assessments of glucose and beta-hydroxybutyrate (BHB) oxidation. At adulthood behavioral tests and histology were performed. Behavioral and histological analysis showed greater impairments in HIP11 animals, while HIP3 rats were not affected. Changes in [18F]FDG metabolism were found only in the lesion area of HIP11, where a substantial hypometabolism was detected. Furthermore, [18F]FDG hypometabolism predicted impaired cognition and worst histological outcomes at adulthood. Finally, substrate oxidation assessments showed that glucose oxidation remained unaltered and higher level of BHB oxidation found in P3 animals, suggesting a more resilient metabolism. Overall, present results show [18F]FDG microPET predicts long-term injury outcome and suggests that higher BHB utilization is one of the mechanisms that confer the intrinsic neuroprotection to the immature brain and should be explored as a therapeutic target for treatment of HI.

Tissue Injury and Astrocytic Reaction, But Not Cognitive Deficits, Are Dependent on Hypoxia Duration in Very Immature Rats Undergoing Neonatal Hypoxia-Ischemia.

Preterm birth and hypoxia-ischemia (HI) are major causes of neonatal death and neurological disabilities in newborns. The widely used preclinical HI model combines carotid occlusion with hypoxia exposure; however, the relationship between different hypoxia exposure periods with brain tissue loss, astrocyte reactivity and behavioral impairments following HI is lacking. Present study evaluated HI-induced behavioral and morphological consequences in rats exposed to different periods of hypoxia at postnatal day 3. Wistar rats of both sexes were assigned into four groups: control group, HI-120 min, HI-180 min and HI-210 min. Neurodevelopmental reflexes, exploratory abilities and cognitive function were assessed. At adulthood, tissue damage and reactive astrogliosis were measured. Animals exposed to HI-180 and HI-210 min had delayed neurodevelopmental reflexes compared to control group. Histological assessment showed tissue loss that was restricted to the ipsilateral hemisphere in lower periods of hypoxia exposure (120 and 180 min) but affected both hemispheres when 210 min was used. Reactive astrogliosis was increased only after 210 min of hypoxia. Interestingly, cognitive deficits were induced regardless the duration of hypoxia and there were correlations between behavioral parameters and cortex, hippocampus and corpus callosum volumes. These results show the duration of hypoxia has a close relationship with astrocytic response and tissue damage progression. Furthermore, the long-lasting cognitive memory deficit and its association with brain structures beyond the hippocampus suggests that complex anatomical changes should be involved in functional alterations taking place as hypoxia duration is increased, even when the cognitive impairment limit is achieved.

Arundic acid administration protects astrocytes, recovers histological damage and memory deficits induced by neonatal hypoxia ischemia in rats.

INTRODUCTION: Perinatal hypoxia-ischemia (HI) is one of the main causes of mortality and chronic neurological morbidity in infants and children. Astrocytes play a key role in HI progression, becoming reactive in response to the injury, releasing S100 calcium binding protein B (S100B). Since S100B inhibition seems to have neuroprotective effects on central nervous system injury models, here we evaluated the neuroprotective effects of an S100B inhibitor, arundic acid (AA) in a HI model. METHODS: On the 7th postnatal day, animals were submitted to the combination of common carotid artery occlusion and hypoxic atmosphere (8% O2) for 60 min. Three experiments were performed in order to: (1) define AA dose (0.1, 1 or 10 mg/kg, pre-hypoxia i.p. injection), (2) test if repeated AA administrations (10 mg/kg at 3 time points: Pre-hypoxia, 24 h and 48 h after HI) would improve the response and (3) investigate biochemical mechanisms involved in AA protection two days after HI. RESULTS: AA at a dose of 10 mg/kg applied before and after hypoxia, was the only treatment protocol that was able to improve HI-induced memory deficits, to reduce tissue damage, to promote astrocytic survival in the hippocampus and to reduced extracellular release of S100B in the cerebrospinal fluid. CONCLUSION: Overall, AA treatment showed beneficial effects on memory deficits, tissue damage, promoting astrocyte survival likely by reducing S100B release. Protection aided to astrocytes by AA treatment against HI lesion may lead to development of new therapeutic strategies that target these particular cells.

Preventive and therapeutic effects of environmental enrichment in Wistar rats submitted to neonatal hypoxia-ischemia.

Environmental enrichment (EE) at early stages of neurodevelopment attenuates HI-induced behavioral, histological and cellular damage. However, the effects of EE exposure during gestational or early postnatal period and the possible influence of sexual dimorphism on EE protection are not fully understood. Present study evaluated the effects of pre-natal and postnatal EE, as well as their combination, in male and female rats submitted to neonatal HI at postnatal day (PND) 3. Wistar rats were housed in EE or in standard condition (SC) during all pregnancy. At PND1, the litters were randomly allocated to the same prenatal environment during lactation (SC + SC or EE + EE) or housed in a new environment until weaning (SC + EE or EE + SC). Behavioral tasks were performed from PND 60-75. Then, animals were euthanized for biochemical and histological analysis. Prenatal and early postnatal EE alone improved performance of HI males in the Water Maze spatial memory task, while HI females were most benefited from early postnatal stimulation. Moreover, EE attenuated HI-induced lower anxiety-like behavior in rats of both sexes and decreased hyperlocomotion in HI females. Hippocampus tissue preservation and higher VEGF and TrkB levels were observed in all HI groups exposed to EE. Interestingly, HI males exposed to prenatal or postnatal EE alone exhibited higher GFAP levels and additional tissue preservation. Therefore, both prenatal and early postnatal environmental enrichment cause attenuation of HI-induced impairments, revealing their preventive and therapeutic actions, possibly due to VEGF and astrocyte activity; some of these effects are sex-specific.

Prenatal and Early Postnatal Environmental Enrichment Reduce Acute Cell Death and Prevent Neurodevelopment and Memory Impairments in Rats Submitted to Neonatal Hypoxia Ischemia.

Environmental enrichment (EE) is an experimental strategy to attenuate the negative effects of different neurological conditions including neonatal hypoxia ischemia encephalopathy (HIE). The aim of the present study was to investigate the influence of prenatal and early postnatal EE in animals submitted to neonatal HIE model at postnatal day (PND) 3. Wistar rats were housed in EE or standard conditions (SC) during pregnancy and lactation periods. Pups of both sexes were assigned to one of four experimental groups, considering the early environmental conditions and the injury: SC-Sham, SC-HIE, EE-sham, and EE-HIE. The offspring were euthanized at two different time points: 48 h after HIE for biochemical analyses or at PND 67 for histological analyses. Behavioral tests were performed at PND 7, 14, 21, and 60. Offspring from EE mothers had better performance in neurodevelopmental and spatial memory tests when compared to the SC groups. HIE animals showed a reduction of IGF-1 and VEGF in the parietal cortex, but no differences in BDNF and TrkB levels were found. EE-HIE animals showed reduction in cell death, lower astrocyte reactivity, and an increase in AKTp levels in the hippocampus and parietal cortex. In addition, the EE was also able to prevent the hippocampus tissue loss. Altogether, present findings point to the protective potential of the prenatal and early postnatal EE in attenuating molecular and histological damage, as well as the neurodevelopmental impairments and the cognitive deficit, caused by HIE insult at PND 3.

Galantamine administration reduces reactive astrogliosis and upregulates the anti-oxidant enzyme catalase in rats submitted to neonatal hypoxia ischemia.

Neonatal hypoxia ischemia (HI) plays a role in the etiology of several neurological pathologies and causes severe sequelae. Acetylcholine is a neurotransmitter in the central nervous system and cholinesterase inhibitors have demonstrated a positive action over HI induced deficits. In order to evaluate the effects of pre and post-hypoxia administrations of galantamine, a cholinesterase inhibitor, in a model of perinatal HI, Wistar rats in the post-natal day 7 (PND7) were subjected to a combination of unilateral occlusion of the right carotid artery with the exposure to a 1h hypoxia. Intraperitoneal injections of galantamine were administered in two different protocols: one pre and other post-hypoxia. The analysis of brain structures volume at PND45 showed that pre-hypoxia galantamine treatment prevented tissue injury to the ipsilesional hippocampus. Also, immunofluorescence showed HI-induced increase in the number of astrocytes that was prevented by pre-hypoxia treatment. Biochemical analysis was performed in the ipsilesional hippocampus at PND8 and revealed that pre-hypoxia galantamine treatment: 1) prevented the neuronal loss induced by HI; 2) reduced the HI-induced hypertrophy of astrocytes; and 3) caused an increase in the activity of the anti-oxidant enzyme catalase. Overall, treatment with galantamine was able to prevent the brain damage, increase the survival of neurons, reduce astrocytic reaction and increase the activity of the anti-oxidant enzyme catalase in rats submitted to neonatal hypoxia ischemia.

Administration of Huperzia quadrifariata Extract, a Cholinesterase Inhibitory Alkaloid Mixture, has Neuroprotective Effects in a Rat Model of Cerebral Hypoxia-Ischemia.

Neonatal hypoxia-ischemia (HI) is an etiologic component of several neurologic pathologies associated to cognitive impairment. The mechanisms involved in HI-induced tissue damage start immediately after HI and extend for days. Acetylcholine is an important neurotransmitter in the central nervous system and exerts a protector effect on tissue damage by modulating inflammation, and cholinesterase inhibitors have shown neuroprotective properties and their action are often attributed to inhibition of the immune response. The administration of Huperzia quadrifariata alkaloid extract (HqAE), with potent and selective cholinesterase inhibitor properties, will reduce the HI induced behavioral deficits and tissue damage. A total of 84 newborn Wistar rat pups at post natal day 7 (PND7) were subjected to right carotid occlusion followed by 1 h of hypoxia (8% of O2) and i.p. injections of saline, vehicle or HqAE (10 mg/kg). Morris Water Maze and inhibitory avoidance tests were used to assess the cognitive function. Flow cytometry was performed at PND11. Histological analysis was performed at PND45. HqAE treatment was able to prevent the HI induced cognitive deficits in both tests and, at PND45, histological analysis showed that HqAE treatment reduced hippocampus tissue damage. Flow cytometry of the injured hippocampus revealed that the treatment was able to reduce cellular death and the number of infiltrating T cells. Altogether, these results show the therapeutic potential of the Huperzia quadrifariata alkaloid extract to prevent cognitive deficits and histological damage caused by neonatal hypoxia-ischemia, probably by reducing cellular death and T cell mobilization.