Peripheral sTREM2-Related Inflammatory Activity Alterations in Early-Stage Alzheimer's Disease.

Alzheimer's disease (AD) has been linked to multiple immune system-related genetic variants. Triggering receptor expressed on myeloid cells 2 (TREM2) genetic variants are risk factors for AD and other neurodegenerative diseases. In addition, soluble TREM2 (sTREM2) isoform is elevated in cerebrospinal fluid in the early stages of AD and is associated with slower cognitive decline in a disease stage-dependent manner. Multiple studies have reported an altered peripheral immune response in AD. However, less is known about the relationship between peripheral sTREM2 and an altered peripheral immune response in AD. The objective of this study was to explore the relationship between human plasma sTREM2 and inflammatory activity in AD. The hypothesis of this exploratory study was that sTREM2-related inflammatory activity differs by AD stage. We observed different patterns of inflammatory activity across AD stages that implicate early-stage alterations in peripheral sTREM2-related inflammatory activity in AD. Notably, fractalkine showed a significant relationship with sTREM2 across different analyses in the control groups that was lost in later AD-related stages with high levels in mild cognitive impairment. Although multiple other inflammatory factors either differed significantly between groups or were significantly correlated with sTREM2 within specific groups, three inflammatory factors (fibroblast growth factor-2, GM-CSF, and IL-1ß) are notable because they exhibited both lower levels in AD, compared with mild cognitive impairment, and a change in the relationship with sTREM2. This evidence provides important support to the hypothesis that sTREM2-related inflammatory activity alterations are AD stage specific and provides critical information for therapeutic strategies focused on the immune response.

Lupus-prone B6.Nba2 male and female mice display anti-DWEYS reactivity and a neuropsychiatric phenotype.

OBJECTIVE: Neuropsychiatric lupus (NPSLE), a manifestation of the autoimmune disease systemic lupus erythematosus (SLE), is characterized by psychiatric symptoms including anxiety and depression and upregulated autoantibodies. The B6.Nba2 spontaneous mouse model develops SLE, but has not previously been tested for NPSLE. METHODS: We investigated the NPSLE phenotype in male and female B6.Nba2 mice (n = 12 each) and age- and sex-matched B6 controls (n = 10 each) via behavioral assessments for anxiety, depression, and memory deficits. Serum anti-dsDNA, anti-nRNP, anti-DWEYS peptide reactive IgG autoantibody levels and soluble TWEAK levels were determined by ELISA. Hippocampal regions were stained for activated microglia and neurons. RESULTS: Both male and female B6.Nba2 mice showed elevated anti-dsDNA IgG, anti-nRNP IgG and anti-DWEYS reactive antibodies, elevated serum soluble TWEAK levels, and a strong anxiety and depression phenotype (p < 0.05-0.0001). Male B6.Nba2 mice developed this phenotype at a slightly older age than females. Female B6.Nba2 mice displayed reduced numbers of neurons in the hippocampal region compared to female B6 controls (p < 0.05). CONCLUSION: The B6.Nba2 mouse model recapitulates many known NPSLE phenotypes, making it a promising model to investigate the development of NPSLE in the context of SLE.

Treg-specific IL-27Rα deletion uncovers a key role for IL-27 in Treg function to control autoimmunity.

Dysregulated Foxp3+ Treg functions result in uncontrolled immune activation and autoimmunity. Therefore, identifying cellular factors modulating Treg functions is an area of great importance. Here, using Treg-specific Il27ra-/- mice, we report that IL-27 signaling in Foxp3+ Tregs is essential for Tregs to control autoimmune inflammation in the central nervous system (CNS). Following experimental autoimmune encephalomyelitis (EAE) induction, Treg-specific Il27ra-/- mice develop more severe EAE. Consistent with the severe disease, the numbers of IFNγ- and IL-17-producing CD4 T cells infiltrating the CNS tissues are greater in these mice. Treg accumulation in the inflamed CNS tissues is not affected by the lack of IL-27 signaling in Tregs, suggesting a functional defect of Il27ra-/- Tregs. IL-10 production by conventional CD4 T cells and their CNS accumulation are rather elevated in Treg-specific Il27ra-/- mice. Analysis with Treg fate-mapping reporter mice further demonstrates that IL-27 signaling in Tregs may control stability of Foxp3 expression. Finally, systemic administration of recombinant IL-27 in Treg-specific Il27ra-/- mice fails to ameliorate the disease even in the presence of IL-27-responsive conventional CD4 T cells. These findings uncover a previously unknown role of IL-27 in regulating Treg function to control autoimmune inflammation.

Motor and Sensory Deficits in the teetering Mice Result from Mutation of the ESCRT Component HGS.

Neurons are particularly vulnerable to perturbations in endo-lysosomal transport, as several neurological disorders are caused by a primary deficit in this pathway. In this report, we used positional cloning to show that the spontaneously occurring neurological mutation teetering (tn) is a single nucleotide substitution in hepatocyte growth factor-regulated tyrosine kinase substrate (Hgs/Hrs), a component of the endosomal sorting complex required for transport (ESCRT). The tn mice exhibit hypokenesis, muscle weakness, reduced muscle size and early perinatal lethality by 5-weeks of age. Although HGS has been suggested to be essential for the sorting of ubiquitinated membrane proteins to the lysosome, there were no alterations in receptor tyrosine kinase levels in the central nervous system, and only a modest decrease in tropomyosin receptor kinase B (TrkB) in the sciatic nerves of the tn mice. Instead, loss of HGS resulted in structural alterations at the neuromuscular junction (NMJ), including swellings and ultra-terminal sprouting at motor axon terminals and an increase in the number of endosomes and multivesicular bodies. These structural changes were accompanied by a reduction in spontaneous and evoked release of acetylcholine, indicating a deficit in neurotransmitter release at the NMJ. These deficits in synaptic transmission were associated with elevated levels of ubiquitinated proteins in the synaptosome fraction. In addition to the deficits in neuronal function, mutation of Hgs resulted in both hypermyelinated and dysmyelinated axons in the tn mice, which supports a growing body of evidence that ESCRTs are required for proper myelination of peripheral nerves. Our results indicate that HGS has multiple roles in the nervous system and demonstrate a previously unanticipated requirement for ESCRTs in the maintenance of synaptic transmission.

Myocardial ischemia/reperfusion impairs neurogenesis and hippocampal-dependent learning and memory.

The incidence of cognitive impairment in cardiovascular disease (CVD) patients has increased, adversely impacting quality of life and imposing a significant economic burden. Brain imaging of CVD patients has detected changes in the hippocampus, a brain region critical for normal learning and memory. However, it is not clear whether adverse cardiac events or other associated co-morbidities impair cognition. Here, using a murine model of acute myocardial ischemia/reperfusion (I/R), where the coronary artery was occluded for 30min followed by reperfusion, we tested the hypothesis that acute myocardial infarction triggers impairment in cognitive function. Two months following cardiac I/R, behavioral assessments specific for hippocampal cognitive function were performed. Mice subjected to cardiac I/R performed worse in the fear-conditioning paradigm as well as the object location memory behavioral test compared to sham-operated mice. Reactive gliosis was apparent in the hippocampal subregions CA1, CA3, and dentate gyrus 72h post-cardiac I/R as compared with sham, which was sustained two months post-cardiac I/R. Consistent with the inflammatory response, the abundance of doublecortin positive newborn neurons was decreased in the dentate gyrus 72h and 2months post-cardiac I/R as compared with sham. Therefore, we conclude that following acute myocardial infarction, rapid inflammatory responses negatively affect neurogenesis, which may underlie long-term changes in learning and memory.

Inhibition of System Xc(-) Transporter Attenuates Autoimmune Inflammatory Demyelination.

T cell infiltration into the CNS is a significant underlying pathogenesis in autoimmune inflammatory demyelinating diseases. Several lines of evidence suggest that glutamate dysregulation in the CNS is an important consequence of immune cell infiltration in neuroinflammatory demyelinating diseases; yet, the causal link between inflammation and glutamate dysregulation is not well understood. A major source of glutamate release during oxidative stress is the system Xc(-) transporter; however, this mechanism has not been tested in animal models of autoimmune inflammatory demyelination. We find that pharmacological and genetic inhibition of system Xc(-) attenuates chronic and relapsing-remitting experimental autoimmune encephalomyelitis (EAE). Remarkably, pharmacological blockade of system Xc(-) 7 d after induction of EAE attenuated T cell infiltration into the CNS, but not T cell activation in the periphery. Mice harboring a Slc7a11 (xCT) mutation that inactivated system Xc(-) were resistant to EAE, corroborating a central role for system Xc(-) in mediating immune cell infiltration. We next examined the role of the system Xc(-) transporter in the CNS after immune cell infiltration. Pharmacological inhibitors of the system Xc(-) transporter administered during the first relapse in a SJL animal model of relapsing-remitting EAE abrogated clinical disease, inflammation, and myelin loss. Primary coculture studies demonstrate that myelin-specific CD4(+) Th1 cells provoke microglia to release glutamate via the system Xc(-) transporter, causing excitotoxic death to mature myelin-producing oligodendrocytes. Taken together, these studies support a novel role for the system Xc(-) transporter in mediating T cell infiltration into the CNS as well as promoting myelin destruction after immune cell infiltration in EAE.

LRRK2 inhibition attenuates microglial inflammatory responses.

Missense mutations in leucine-rich repeat kinase 2 (LRRK2) cause late-onset Parkinson's disease (PD), and common genetic variation in LRRK2 modifies susceptibility to Crohn's disease and leprosy. High levels of LRRK2 expression in peripheral monocytes and macrophages suggest a role for LRRK2 in these cells, yet little is known about LRRK2 expression and function in immune cells of the brain. Here, we demonstrate a role for LRRK2 in mediating microglial proinflammatory responses and morphology. In a murine model of neuroinflammation, we observe robust induction of LRRK2 in microglia. Experiments with toll-like receptor 4 (TLR4)-stimulated rat primary microglia show that inflammation increases LRRK2 activity and expression, while inhibition of LRRK2 kinase activity or knockdown of protein attenuates TNFα secretion and nitric oxide synthase (iNOS) induction. LRRK2 inhibition blocks TLR4 stimulated microglial process outgrowth and impairs ADP stimulated microglial chemotaxis. However, actin inhibitors that phenocopy inhibition of process outgrowth and chemotaxis fail to modify TLR4 stimulation of TNFα secretion and inducible iNOS induction, suggesting that LRRK2 acts upstream of cytoskeleton control as a stress-responsive kinase. These data demonstrate LRRK2 in regulating responses in immune cells of the brain and further implicate microglial involvement in late-onset PD.

Utility of the visual system to monitor neurodegeneration in multiple sclerosis.

Neurodegeneration occurs early in the multiple sclerosis (MS) disease course and is an important driver of permanent disability. Current immunomodulatory therapies do not directly target neuronal health; thus, there is a critical need to develop neuroprotective strategies in MS. Outcome measures in clinical trials primarily evaluate disease activity and clinical disability scores rather than measures of neurodegeneration. The visual system provides a noninvasive correlate of brain atrophy and neuronal function through structural and functional exams. Furthermore, optic nerve axons and their respective neuronal cell bodies in the retina, in addition to their synaptic input to the thalamus, provide a distinct anatomy to investigate neurodegenerative processes. This review discusses the utility of the visual system as an early output measure of neurodegeneration in MS as well as an important platform to evaluate neuroprotective strategies in preclinical models.

Neurodegeneration in multiple sclerosis.

Axonal loss in multiple sclerosis (MS) is a key component of disease progression and permanent neurologic disability. MS is a heterogeneous demyelinating and neurodegenerative disease of the central nervous system (CNS) with varying presentation, disease courses, and prognosis. Immunomodulatory therapies reduce the frequency and severity of inflammatory demyelinating events that are a hallmark of MS, but there is minimal therapy to treat progressive disease and there is no cure. Data from patients with MS, post-mortem histological analysis, and animal models of demyelinating disease have elucidated patterns of MS pathogenesis and underlying mechanisms of neurodegeneration. MRI and molecular biomarkers have been proposed to identify predictors of neurodegeneration and risk factors for disease progression. Early signs of axonal dysfunction have come to light including impaired mitochondrial trafficking, structural axonal changes, and synaptic alterations. With sustained inflammation as well as impaired remyelination, axons succumb to degeneration contributing to CNS atrophy and worsening of disease. These studies highlight the role of chronic demyelination in the CNS in perpetuating axonal loss, and the difficulty in promoting remyelination and repair amidst persistent inflammatory insult. Regenerative and neuroprotective strategies are essential to overcome this barrier, with early intervention being critical to rescue axonal integrity and function. The clinical and basic research studies discussed in this review have set the stage for identifying key propagators of neurodegeneration in MS, leading the way for neuroprotective therapeutic development. This article is categorized under: Immune System Diseases > Molecular and Cellular Physiology Neurological Diseases > Molecular and Cellular Physiology.

Endothelial-to-mesenchymal transition in multiple sclerosis: Good cells gone bad.

In cancer and cardiovascular disease, endothelial cells can transform into mesenchymal cells (EndoMT), affecting disease onset and progression. In this issue of Neuron, Sun et al. (2022) demonstrate how EndoMT triggers breakdown of the blood-CNS barrier in the pathogenesis of multiple sclerosis.

Remyelinating strategies: What can be learned from normal brain development.

Multiple sclerosis (MS) is a neuroinflammatory demyelinating and neurodegenerative disease of the central nervous system (CNS). Immunomodulatory therapies are effective in reducing relapses, however, there is no remedy for progressive disease emphasizing the need for regenerative strategies. Chronic demyelination causes axonal injury and loss which is a key component of neurodegeneration and permanent disability in MS. New oligodendrocyte progenitor cells (OPCs) proliferate in response to inflammatory demyelination representing the potential for remyelination to protect axons and preserve neuronal function. The majority of remyelinating therapies have targeted intrinsic signaling processes in oligodendrocytes to promote differentiation or utilized methods for transplantation of oligodendrocytes. Here, we discuss specific roles of microglia in contributing to normal myelin development and the significance of these functions for remyelinating strategies.

Microglia phagocytose oligodendrocyte progenitor cells and synapses during early postnatal development: implications for white versus gray matter maturation.

Emerging roles for microglia in modifying normal brain development continue to provide new perspectives on the functions of this resident immune cell in the brain. While the molecular underpinnings driving microglia's position in regulating developmental programs remain largely an unchartered territory, innate immune signaling lies at the forefront. At least three innate immune receptors expressed on microglia-fractalkine, complement, and triggering receptor expressed on microglia (TREM2)-modulate developmental synaptic pruning to refine brain circuitry. Our laboratory recently published that microglia with a unique amoeboid morphology invade the corpus callosum and engulf oligodendrocyte progenitor cells (OPCs) during early postnatal development before myelination in a fractalkine receptor (CX3CR1)-dependent manner to modulate ensheathment of axons. Amoeboid microglia are observed in the corpus callosum but not cerebral cortex, and lose their amoeboid shape at the commencement of myelination assuming a resting phenotype. Furthermore, OPCs contacted or engulfed by microglia do not express markers of cell death suggesting a novel homeostatic mechanism facilitating an appropriate OPC:axon ratio for proper myelin ensheathment. The unique morphology of microglia and the restricted window for phagocytic engulfment of OPCs suggest a critical period for OPC engulfment important for action potential propagation during development when activity-dependent mechanisms regulate synaptic pruning. In this review, we summarize the role of activity-dependent mechanisms in sculpting brain circuitry, how myelin ensheathment influences action potential propagation, the spatial and temporal relationship of microglia-dependent elimination of OPCs and synapses, and implications for the synergistic role of microglial phagocytosis in shaping the architecture for neuronal function.

Hyperoxia Induced Hypomyelination.

We asked whether hyperoxia might induce hypomyelination of the corpus callosum, clinically described as periventricular leukomalacia (PVL) of the severely preterm infant. Mouse pups and their nursing dams were placed in 80% oxygen from P4-P8, then removed to room air until P11. Corpus callosal sections were probed myelin immunofluorescence, tested for myelin basic protein concentration by Western blot, and both glial fibrillary acidic protein levels and apoptosis quantified. Density of corpus callosal capillaries were measured after lectin staining and hypoxia measured by Hypoxyprobe. Numbers of oligodendrocytes were quantified by immunohistochemistry. We next used hypoxiamimesis as a surrogate to hypoxia by comparing cerebral hypoxia inducible factor (HIF) stabilization to hepatic HIF stabilization. Hyperoxia induced hypomyelination and a reduction of corpus callosal capillaries. Hyperoxia decreased numbers of oligodendrocytes with an increase in corpus callosal fibrosis and apoptosis. Cerebral hypoxiamimesis induced hypomyelination whereas hepatic hypoxiamimesis alone increased myelination, oligodendrocyte numbers, and corpus callosal capillary density. Hepatic HIF-1 dependence on myelination was confirmed using the cre/lox hepatic HIF-1 knockout. These findings suggest that hyperoxia can induce hypomyelination through vasoobliteration and subsequent ischemia, adding a potential oxygen induced mechanism to the diverse causes of periventricular leukomalacia of the severely preterm infant. Targeting hepatic HIF-1 alone led to increased myelination.

Visual imaging as a predictor of neurodegeneration in experimental autoimmune demyelination and multiple sclerosis.

Thalamic volume is associated with clinical disability in multiple sclerosis (MS) and is vulnerable to secondary neurodegeneration due to its extensive connectivity throughout the central nervous system (CNS). Using a model of autoimmune demyelination that exhibits CNS-infiltrating immune cells in both spinal cord white matter and optic nerve, we sought to evaluate neurodegenerative changes due to lesions affecting the spino- and retino-thalamic pathways. We found comparable axonal loss in spinal cord white matter and optic nerve during the acute phase of disease consistent with synaptic loss, but not neuronal cell body loss in the thalamic nuclei that receive input from these discrete pathways. Loss of spinal cord neurons or retinal ganglion cells retrograde to their respective axons was not observed until the chronic phase of disease, where optical coherence tomography (OCT) documented reduced inner retinal thickness. In patients with relapsing-remitting MS without a history of optic neuritis, OCT measures of inner retinal volume correlated with retino-thalamic (lateral geniculate nucleus) and spino-thalamic (ventral posterior nucleus) volume as well as neuroperformance measures. These data suggest retinal imaging may serve as an important noninvasive predictor of neurodegeneration in MS.

Quantification of microglial contact and engulfment of oligodendrocyte progenitor cells in the rodent brain.

Microglia have emerged as essential regulators of neurodevelopment by phagocytosing synapses. Recently, we showed that microglia engulf viable oligodendrocyte progenitor cells (OPCs) during development to facilitate myelination. Here, we describe a protocol to quantify microglial engulfment of whole cells using 3D confocal microscopy to differentiate microglial contact. This protocol can be applied to assess whole-cell engulfment in a variety of contexts and cell types. For complete details on the use and execution of this protocol, please refer to Nemes-Baran et al. (2020).

Regulation of glutamate transport in developing rat oligodendrocytes.

Glutamate released from synaptic vesicles mediates excitatory neurotransmission by stimulating glutamate receptors. Glutamate transporters maintain low synaptic glutamate levels critical for this process, a role primarily attributed to astrocytes. Recently, vesicular release of glutamate from unmyelinated axons in the rat corpus callosum has been shown to elicit AMPA receptor-mediated currents in glial progenitor cells. Glutamate transporters are the only mechanism of glutamate clearance, yet very little is known about the role of glutamate transporters in normal development of oligodendrocytes (OLs) or in excitotoxic injury to OLs. We found that OLs in culture are capable of sodium-dependent glutamate uptake with a K(m) of 10 +/- 2 microm and a V(max) of 2.6, 5.0, and 3.8 nmol x min(-1) x mg(-1) for preoligodendrocytes, immature, and mature OLs, respectively. Surprisingly, EAAC1, thought to be exclusively a neuronal transporter, contributes more to [(3)H]l-glutamate uptake in OLs than GLT1 or GLAST. These data suggest that glutamate transporters on oligodendrocytes may serve a critical role in maintaining glutamate homeostasis at a time when unmyelinated callosal axons are engaging in glutamatergic signaling with glial progenitors. Furthermore, GLT1 was significantly increased in cultured mature OLs contrary to in vivo data in which we have shown that, although GLT1 is present on developing OLs when unmyelinated axons are prevalent in the developing rat corpus callosum, after myelination, GLT1 is not expressed on mature OLs. The absence of GLT1 in mature OLs in the rat corpus callosum and its presence in mature rat cultured OLs may indicate that a signaling process in vivo is not activated in vitro.

Fractalkine-Dependent Microglial Pruning of Viable Oligodendrocyte Progenitor Cells Regulates Myelination.

Oligodendrogenesis occurs during early postnatal development, coincident with neurogenesis and synaptogenesis, raising the possibility that microglia-dependent pruning mechanisms that modulate neurons regulate myelin sheath formation. Here we show a population of ameboid microglia migrating from the ventricular zone into the corpus callosum during early postnatal development, termed "the fountain of microglia," phagocytosing viable oligodendrocyte progenitor cells (OPCs) before onset of myelination. Fractalkine receptor-deficient mice exhibit a reduction in microglial engulfment of viable OPCs, increased numbers of oligodendrocytes, and reduced myelin thickness but no change in axon number. These data provide evidence that microglia phagocytose OPCs as a homeostatic mechanism for proper myelination. A hallmark of hypomyelinating developmental disorders such as periventricular leukomalacia and of adult demyelinating diseases such as multiple sclerosis is increased numbers of oligodendrocytes but failure to myelinate, suggesting that microglial pruning of OPCs may be impaired in pathological states and hinder myelination.

Reduction of AMPA receptor activity on mature oligodendrocytes attenuates loss of myelinated axons in autoimmune neuroinflammation.

Glutamate dysregulation occurs in multiple sclerosis (MS), but whether excitotoxic mechanisms in mature oligodendrocytes contribute to demyelination and axonal injury is unexplored. Although current treatments modulate the immune system, long-term disability ensues, highlighting the need for neuroprotection. Glutamate is elevated before T2-visible white matter lesions appear in MS. We previously reported that myelin-reactive T cells provoke microglia to release glutamate from the system xc - transporter promoting myelin degradation in experimental autoimmune encephalomyelitis (EAE). Here, we explore the target for glutamate in mature oligodendrocytes. Most glutamate-stimulated calcium influx into oligodendrocyte somas is AMPA receptor (AMPAR)-mediated, and genetic deletion of AMPAR subunit GluA4 decreased intracellular calcium responses. Inducible deletion of GluA4 on mature oligodendrocytes attenuated EAE and loss of myelinated axons was selectively reduced compared to unmyelinated axons. These data link AMPAR signaling in mature oligodendrocytes to the pathophysiology of myelinated axons, demonstrating glutamate regulation as a potential neuroprotective strategy in MS.

Hyperoxia causes maturation-dependent cell death in the developing white matter.

Periventricular leukomalacia is the predominant injury in the preterm infant leading to cerebral palsy. Oxygen exposure may be an additional cause of brain injury in these infants. In this study, we investigated pathways of maturation-dependent oligodendrocyte (OL) death induced by hyperoxia in vitro and in vivo. Developing and mature OLs were subjected to 80% oxygen (0-24 h). Lactate dehydrogenase (LDH) assay was used to assess cell viability. Furthermore, 3-, 6-, and 10-d-old rat pups were subjected to 80% oxygen (24 h), and their brains were processed for myelin basic protein staining. Significant cell death was detected after 6-24 h incubation in 80% oxygen in pre-OLs (O4+,O1-), but not in mature OLs (MBP+). Cell death was executed by a caspase-dependent apoptotic pathway and could be blocked by the pan-caspase inhibitor zVAD-fmk. Overexpression of BCL2 (Homo sapiens B-cell chronic lymphocytic leukemia/lymphoma 2) significantly reduced apoptosis. Accumulation of superoxide and generation of reactive oxygen species (ROS) were detected after 2 h of oxygen exposure. Lipoxygenase inhibitors 2,3,5-trimethyl-6-(12-hydroxy-5-10-dodecadiynyl-1,4-benzoquinone and N-benzyl-N-hydroxy-5-phenylpentamide fully protected the cells from oxidative injury. Overexpression of superoxide dismutase (SOD1) dramatically increased injury to pre-OLs but not to mature OLs. We extended these studies by testing the effects of hyperoxia on neonatal white matter. Postnatal day 3 (P3) and P6 rats, but not P10 pups, showed bilateral reduction in MBP (myelin basic protein) expression with 24 h exposure to 80% oxygen. Hyperoxia causes oxidative stress and triggers maturation-dependent apoptosis in pre-OLs, which involves the generation of ROS and caspase activation, and leads to white matter injury in the neonatal rat brain. These observations may be relevant to white matter injury observed in premature infants.

17beta-estradiol protects against hypoxic/ischemic white matter damage in the neonatal rat brain.

Developing oligodendrocytes (pre-OLs) are highly vulnerable to hypoxic-ischemic injury and associated excitotoxicity and oxidative stress. 17beta-Estradiol plays an important role in the development and function of the CNS and is neuroprotective. The sudden drop in circulating estrogen after birth may enhance the susceptibility of developing OLs to injury. Estrogen receptor (ER)-alpha and ER-beta are both expressed in OLs. We examined the effect of 17beta-estradiol on oxygen-glucose deprivation and oxidative stress-induced cell death in rat pre-OLs in vitro and on hypoxic-ischemic brain injury in vivo. Pre-OLs in culture were subjected to oxygen-glucose deprivation (OGD) or glutathione depletion in the presence or absence of 17beta-estradiol. LDH release, the Alamar blue assay, and phase-contrast microscopy were used to assess cell viability. Hypoxic-ischemic injury was generated in 6-day-old rats (P6) by unilateral carotid ligation and hypoxia (6% O(2) for 1 hr). Rat pups received one intraperitoneal injection of 300 or 600 microg/kg 17beta-estradiol or vehicle 12 hr prior to the surgical procedure. Injury was assessed by myelin basic protein (MBP) immunocytochemistry at P10. 17beta-Estradiol produced significant protection against OGD-induced cell death in primary OLs (EC(50) = 1.3 +/- 0.46 x 10(-9) M) and against oxidative stress. Moreover, 17beta-estradiol attenuated the loss of MBP labeling in P10 pups ipsilateral to the carotid ligation. These results suggest a potential role for estrogens in attenuation of hypoxic-ischemic and oxidative injury to developing OLs and in the prevention of periventricular leukomalacia.