Acute CNS Insults Research Articles

Abstract TMP50: Suppression of α-synuclein Mitigates Secondary Ischemic Brain Damage

α-synuclein (α-syn) is one of the most abundant proteins in mammalian brain that is known to be a major player in the neurodegeneration observed in chronic conditions like Parkinson’s disease (PD). Several mechanisms were proposed for α-syn-induced neuronal death in PD including inflammation, oxidative stress, mitochondrial fission, and autophagy. Interestingly, all these pathophysiologic mechanisms also mediate neuronal death after acute CNS insults like stroke. Therefore, we examined whether α-syn contributes to post-stroke neuronal death and neurological dysfunction. Rodents were subjected to transient middle cerebral artery occlusion (tMCAO) and α-syn induction was silenced with a cocktail of α-syn-specific siRNAs. The levels of α-syn were estimated with qPCR and Western Blots. Post-ischemic motor deficit was evaluated with rotarod, beam walk and adhesive removal test 1 to 7 days after ischemia, and infarct volume was measured on cresyl violet stained brain sections. Cellular changes after ischemia were examined using immunofluorescence staining. Following tMCAO, α-syn levels were significantly up-regulated and phosphorylated at serine-129 in ischemic penumbra. Interestingly, in the ischemic brain non-phosphorylated and phosphorylated α-syn species translocated into the neuronal nuclei. We also observed that silencing α-syn by α-syn siRNA cocktail before tMCAO significantly decreased the infarction and improved the motor function. The neuroprotective effects were still observed when α-syn siRNAs were administered 30 min after the tMCAO. We found that the rate of survival at 7 days after a 90 min tMCAO was significantly higher in α-syn KO mice compared to wild-type control mice. Furthermore, α-syn suppression significantly mitigated the post-ischemic oxidative stress (8-OHdG and 3-NT), apoptosis (cleaved Caspase-3) and mitochondrial dysfunction (phospho-Drp1). Thus, we show that α-syn plays a critical role in neuronal death following stroke. Furthermore, phospho-S129 α-syn in the neuronal nuclei might be a potential mediator of the induction of ischemic brain damage. Preventing α-syn expression is a potential therapeutic target to minimize post-stroke brain damage.

Cooperative effect of p150Glued and microtubule stabilization to suppress excitotoxicity-induced axon degeneration

Glutamate excitotoxicity is implicated in chronic neurological disorders and acute CNS insults and causes neuronal degeneration including axons. The molecular mechanism underlying excitotoxicity-induced axon degeneration is poorly understood. Recently, we found that components of the dynein–dynactin complex that governs microtubule-dependent retrograde transport play important roles in modulating the process of excitotoxicity-induced neurodegeneration. Here we used hippocampal cultures and searched for pathways that function in concert with the components of the dynein–dynactin complex and identified microtubule stabilization as a cooperative pathway to suppress axon degeneration. We find that overexpression of p150Glued, a major component of the dynactin complex, and microtubule stabilization cooperatively suppress axon degeneration. The protective effect of p150Glued is dependent on the C-terminal region as excitotoxicity-induced C-terminal truncated form of p150Glued was unable to interact with APP cargo and altered the localization of APP in neurites when overexpressed. C-terminal truncation of p150Glued is not rescued by microtubule stabilization suggesting that the downstream effects of p150Glued and microtubule stabilization to protect axon degeneration are mutually exclusive.

Dynein and dynactin components modulate neurodegeneration induced by excitotoxicity

Glutamate excitotoxicity causes neuronal dysfunction and degeneration. It is implicated in chronic disorders, including Alzheimer's disease, and in acute CNS insults such as ischemia. These disorders share prominent morphological features, including axon degeneration and cell body death. However, the molecular mechanism underlying excitotoxicity-induced neurodegeneration remains poorly understood. A key molecular feature of neurodegeneration is deficits in microtubule-based cargo transport that plays a pivotal role in maintaining the balance of survival and stress signaling in the axon. We developed an excitotoxicity-induced neurodegeneration system in primary neuronal cultures. We find that excitotoxicity generates a C-terminal truncated form of p150Glued, a major component of the dynactin complex, which exacerbates axon degeneration. This p150Glued truncated form was identified in brain tissues of patients with Alzheimer's disease. Overexpression of wild-type (WT) dynein intermediate chain (DIC), a dynein component that interacts with p150Glued and links dynein and dynactin complexes, DIC (S84D) mutant, and WT p150Glued suppressed axon degeneration. These modulating effects of p150Glued and DIC on excitotoxicity-induced axon degeneration are also observed in apoptosis and cell body death. Thus, our findings identify retrograde transport proteins, p150Glued and DIC, as novel modulators of neurodegeneration induced by glutamate excitotoxicity.

Albumin induces upregulation of matrix metalloproteinase-9 in astrocytes via MAPK and reactive oxygen species-dependent pathways

BackgroundAstrocytes are an integral component of the blood–brain barrier (BBB) which may be compromised by ischemic or traumatic brain injury. In response to trauma, astrocytes increase expression of the endopeptidase matrix metalloproteinase (MMP)-9. Compromise of the BBB leads to the infiltration of fluid and blood-derived proteins including albumin into the brain parenchyma. Albumin has been previously shown to activate astrocytes and induce the production of inflammatory mediators. The effect of albumin on MMP-9 activation in astrocytes is not known. We investigated the molecular mechanisms underlying the production of MMP-9 by albumin in astrocytes.MethodsPrimary enriched astrocyte cultures were used to investigate the effects of exposure to albumin on the release of MMP-9. MMP-9 expression was analyzed by zymography. The involvement of mitogen-activated protein kinase (MAPK), reactive oxygen species (ROS) and the TGF-β receptor-dependent pathways were investigated using pharmacological inhibitors. The production of ROS was observed by dichlorodihydrofluorescein diacetate fluorescence. The level of the MMP-9 inhibitor tissue inhibitor of metalloproteinase (TIMP)-1 produced by astrocytes was measured by ELISA.ResultsWe found that albumin induces a time-dependent release of MMP-9 via the activation of p38 MAPK and extracellular signal regulated kinase, but not Jun kinase. Albumin-induced MMP-9 production also involves ROS production upstream of the MAPK pathways. However, albumin-induced increase in MMP-9 is independent of the TGF-β receptor, previously described as a receptor for albumin. Albumin also induces an increase in TIMP-1 via an undetermined mechanism.ConclusionsThese results link albumin (acting through ROS and the p38 MAPK) to the activation of MMP-9 in astrocytes. Numerous studies identify a role for MMP-9 in the mechanisms of compromise of the BBB, epileptogenesis, or synaptic remodeling after ischemia or traumatic brain injury. The increase in MMP-9 produced by albumin further implicates both astrocytes and albumin in the acute and long-term complications of acute CNS insults, including cerebral edema and epilepsy.

Mass‐spectrometry based oxidative lipidomics and lipid imaging: applications in traumatic brain injury

Lipids, particularly phospholipids, are fundamental to CNS tissue architecture and function. Endogenous polyunsaturated fatty acid chains of phospholipids possess cis-double bonds each separated by one methylene group. These phospholipids are very susceptible to free-radical attack and oxidative modifications. A combination of analytical methods including different versions of chromatography and mass spectrometry allows detailed information to be obtained on the content and distribution of lipids and their oxidation products thus constituting the newly emerging field of oxidative lipidomics. It is becoming evident that specific oxidative modifications of lipids are critical to a number of cellular functions, disease states and responses to oxidative stresses. Oxidative lipidomics is beginning to provide new mechanistic insights into traumatic brain injury which may have significant translational potential for development of therapies in acute CNS insults. In particular, selective oxidation of a mitochondria-specific phospholipid, cardiolipin, has been associated with the initiation and progression of apoptosis in injured neurons thus indicating new drug discovery targets. Furthermore, imaging mass-spectrometry represents an exciting new opportunity for correlating maps of lipid profiles and their oxidation products with structure and neuropathology. This review is focused on these most recent advancements in the field of lipidomics and oxidative lipidomics based on the applications of mass spectrometry and imaging mass spectrometry as they relate to studies of phospholipids in traumatic brain injury.

Targeting individual calpain isoforms for neuroprotection

Targeting individual calpain isoforms for neuroprotection

Treating epilepsy across its different stages

Epilepsy is a chronic condition requiring long-term treatment with drugs that have intrinsic limitations. Antiepileptic drugs (AEDs) are effective in suppressing seizures but do not alter the disease process. They have a suboptimal tolerability profile and can be teratogenic. Second-generation compounds may be better tolerated but no more effective than traditional AEDs. In this light, as drug therapy is purely symptomatic, acute symptomatic seizures (i.e. seizures occurring in close temporal relationship with acute CNS insults) may require treatment only until recovery or stabilization of the injury. Treatment of the first unprovoked seizure may be considered in patients with abnormal EEG and imaging findings and in those in whom the relapse has severe social, emotional and personal implications. In these cases and in patients with epilepsy (i.e. repeated unprovoked seizures), drugs for partial seizures supported by class I regulatory trials or pragmatic trials are oxcarbazepine in children, carbamazepine or lamotrigine in adults, and lamotrigine or gabapentin in the elderly. Pragmatic trials support use of valproate for generalized seizures, except for women of childbearing age for whom the drug should be tailored to the individual patient. The lowest maintenance dose should be chosen, based on the efficacy and tolerability of the assigned drug. If the first monotherapy fails, the safety profile of a drug is important when opting for another monotherapy or for an add-on therapy. The epilepsy syndrome and the social, psychological and emotional profile of the patient all contribute to the individualization of treatment discontinuation after long-term seizure remission.

Attenuation of AD‐like neuropathology by harnessing peripheral immune cells: local elevation of IL‐10 and MMP‐9

Immunization with an altered myelin-derived peptide (MOG45D) improves recovery from acute CNS insults, partially via recruitment of monocyte-derived macrophages that locally display a regulatory activity. Here, we investigated the local alterations in the cellular and molecular immunological milieu associated with attenuation of Alzheimer's disease-like pathology following immunotherapy. We found that immunization of amyloid precursor protein/presenilin 1 double-transgenic mice with MOG45D peptide, loaded on dendritic cells, led to a substantial reduction of parenchymal and perivascular amyloid beta (Abeta)-plaque burden and soluble Abeta((1-42)) peptide levels as well as reduced astrogliosis and levels of a key glial scar protein (chondroitin sulphate proteoglycan). These changes were associated with a shift in the local innate immune response, manifested by increased Iba1+/CD45(high) macrophages that engulfed Abeta, reduced pro-inflammatory (tumor necrosis factor-alpha) and increased anti-inflammatory (interleukin-10) cytokines, as well as a significant increase in growth factors (IGF-1 and TGFbeta) in the brain. Furthermore, the levels of matrix metalloproteinase-9, an enzyme shown to degrade Abeta and is associated with glial scar formation, were significantly elevated in the brain following immunization. Altogether, these results indicate that boosting systemic immune cells leads to a local immunomodulation manifested by elevated levels of anti-inflammatory cytokines and metalloproteinases that contribute to ameliorating Alzheimer's disease pathology.

Mitochondrial uncoupling as a potential therapeutic target in acute central nervous system injury.

Mitochondrial dysfunction, resulting from the disruption of calcium homeostasis and the generation of toxic reactive oxygen species, is a central process leading to neuronal injury and death following acute CNS insults. Interventions aimed at preventing disturbances in mitochondrial function have therefore become targets of intense investigation. Mitochondrial uncoupling is a condition in which electron transport is disconnected from the production of ATP. As a consequence, there is a decrease in the mitochondrial membrane potential, which can temporarily decrease calcium influx and attenuate free radical formation. The potential use of pharmacological agents with uncoupling properties may provide a novel therapeutic approach for the treatment of acute neuronal injury.

Excitotoxicity and the putative involvement of excitatory amino acids in neurodegenerative diseases

Excitatory amino acids (EAA) represent major brain neurotransmitters. They are present in numerous neuronal systems and thus are involved in almost all aspects of normal and pathological brain activity. Changes in EAA transmission have been associated with the functional impairments characterizing major neurological disorders, including epilepsy and schizophrenia. There is also a suspicion that EAA systems underlie the neuronal death associated not only with acute CNS insults, such as in ischemia or post-traumatic lesions, but also with neurodegenerative diseases such as ALS, Huntington's disease and Parkinson's disease. The neurotoxicity of EAA, referred to as excitotoxicity, is presumably mediated primarily through an excess of EAA synaptic receptor stimulation. Indeed, overstimulation of the ionotropic NMDA or AMPA/kainate receptor subtypes has been shown to produce an intense membrane depolarisation and further a massive increase in intracellular calcium leading to cell damage. The extreme diversity and specific pattern of expression of EAA receptor subunits could account for the differential vulnerability of certain brain areas to such excitotoxic processes. In addition, it is now believed that besides abnormalities in receptor functioning or in release processes, alterations in EAA transmission may result from dysfunction of the EAA uptake system, which represents the mechanism for EAA removal from the synapse. From the five transporter proteins cloned, termed EAAT1-5, the primarily glial transporters EAAT1 and EAAT2 have been shown to mediate the bulk of EAA uptake in the brain and it has then been suggested that they play a major role in the prevention of excitotoxic processes. In this respect, the degeneration of motor neurons in ALS has been associated with altered expression or inactivation of EAAT2. Moreover, recent evidence has been provided that pharmacological alteration of glutamate transport can also induce astrocyte degeneration, as observed in neurodegenerative insults, but through a mechanism independent of stimulation of EAA receptors. Thus, one can obviously consider that these EAATs can represent a key target for further development of new neuroprotective agents.

Activation of group I metabotropic glutamate receptors reduces neuronal apoptosis but increases necrotic cell death in vitro.

Glutamate released during acute CNS insults acts at metabotropic glutamate receptors (mGluR), including group I mGluR. Blockade of group I mGluR during in vitro neuronal trauma provides neuroprotection, whereas activation exacerbates such injury. However, the effects of group I mGluR agonists or antagonists have been primarily studied in in vitro models characterized by necrotic cell death. We examined the role of group I mGluR in the modulation of neuronal injury induced during oxygen-glucose deprivation (OGD), a well-studied model of necrosis, and by application of two well established pro-apoptotic agents: staurosporine and etoposide. Inhibition of group I mGluR attenuated necrosis induced by OGD, whereas selective activation of group I mGluR exacerbated such injury. In contrast, activation of group I mGluR, including selective activation of mGluR5, significantly attenuated apoptotic cell death induced by both staurosporine and etoposide. This effect was completely reversed by co-application of a group I mGluR antagonist. Thus, group I mGluR appear to exhibit opposite effects on necrotic and apoptotic neuronal cell death. Our findings suggest that activation of mGluR1 exacerbates neuronal necrosis whereas both mGluR1 and mGluR5 play a role in attenuation of neuronal apoptosis.

Estrogens attenuate neuronal injury due to hemoglobin, chemical hypoxia, and excitatory amino acids in murine cortical cultures

A growing body of evidence supports the hypothesis that estrogens may be beneficial in Alzheimer's disease and other neurodegenerative processes. Less is known of their therapeutic potential in acute CNS insults. In this study, we assessed the effect of estrogens in three injury paradigms that may be relevant to CNS hemorrhage, trauma, and ischemia. Supraphysiologic concentrations of 17β-estradiol, estrone, or equilin attenuated neuronal loss due to prolonged exposure to the pro-oxidant hemoglobin, with complete protection at 10 μM. Most of this effect persisted despite concomitant treatment with the antiestrogen ICI 182,780 or the protein synthesis inhibitor cycloheximide. In contrast, the non-estrogenic steroid methylprednisolone, which is currently in clinical use in spinal cord injury, reduced neuronal loss by only about 30%. High concentrations of equilin or estrone also attenuated the submaximal neuronal injury induced by 3.5–4.5 h exposure to the cytochrome oxidase inhibitor sodium azide, with near complete protection at 30 μM. Estrogens had a weaker and somewhat variable effect on pure excitotoxic injury, reducing neuronal loss due to 24 h kainate exposure by about half, and due to 24 h NMDA exposure by 15–65%; similar neuroprotection was provided by the antioxidant 21-aminosteroid U74500A. These results suggest that estrogens may be beneficial in acute CNS injuries associated with oxidative and excitotoxic stress. Investigation of high dose estrogen therapy in in vivo models of CNS hemorrhage, trauma, and ischemia is warranted.

Incidence of Acute Symptomatic Seizures in Rochester, Minnesota, 1935‐1984

We determined the incidence of seizures due to acute CNS insults for residents of Rochester, Minnesota, U.S.A., from 1935 through 1984. The age-adjusted incidence rates for 1955-1984, the period of most complete case ascertainment, was 39.0/100,000 person-years (United States 1970 population as standard). The age-adjusted incidence was considerably higher in men: 52.0 as compared with 29.5 in women. The 3.6% risk of experiencing an acute symptomatic seizure in an 80-year lifespan approaches that of developing epilepsy. The major causes of acute symptomatic seizures were traumatic brain injury, cerebrovascular disease, drug withdrawal, and CNS infections. Each type of acute symptomatic seizure has age, gender, and time period patterns that reflect the occurrence of the underlying cause.

Excitotoxicity and motor neurone disease: A review of the evidence

Excitotoxic mechanisms have a well established role in the pathogenesis of neuronal injury following acute CNS insults such as ischaemia and trauma. Their role in the selective cell death which occurs in chronic neurodegenerative disorders such as motor neurone disease (MND) is more speculative. The traditional classification of glutamate receptor subtypes which mediate excitotoxicity requires modification in the light of new molecular data. There is much greater structural and functional diversity in this receptor family than previously envisaged and it is quite possible that specific populations of neurones will be characterised by a unique profile of glutamate receptor subtypes which may be a factor determining their selective vulnerability. The molecular mechanisms underlying excitotoxic neuronal injury are still being elucidated but it is clear that the cascade of events resulting from elevation of intracellular free calcium is likely to play a major role. As well as being a primary mechanism of neuronal injury, excitotoxicity can secondarily damage neurones whose energy metabolism is impaired from some primary pathological process. The 8 lines of evidence that primary or secondary excitotoxic mechanisms may be involved in the selective neuronal injury of MND are discussed. The evidence, while still circumstantial, is sufficient to warrant further research effort in this field, not least because the emergence of pharmacological agents which modify specific aspects of excitatory amino acid neurotransmission offer the possibility of therapeutic intervention in MND.

Bacterial meningitis in children: pathophysiology and treatment.

Recent studies of the pathophysiology of bacterial meningitis have suggested that the development of neuronal injury is related to the release of vasoactive substances or alteration of blood-brain barrier permeability. Cerebral edema, increased intracranial pressure (ICP), systemic hypotension, decreased cerebral perfusion pressure, vascular inflammation, thrombosis, and a variety of other vascular changes may result in global or regional reductions in cerebral blood flow (CBF), which contribute to this insult. Approximately one-third of infants and children with bacterial meningitis will have markedly reduced CBF, and even in those children with normal total flow, regional hypoperfusion is common. Reduced CBF is associated with cerebral edema and a poor prognosis. A poor prognosis also is associated with reduced cerebral perfusion pressure. This occurs early in the course of meningitis and is primarily due to increased ICP rather than systemic hypotension. Autoregulation is preserved, suggesting that local ischemic tissue injury is more related to factors such as regional edema formation, focal vascular pathology, or specific intrinsic flow/metabolic abnormalities than to a reduction in systemic blood pressure. In contrast with other acute CNS insults, CBF/PCO2 reactivity is well preserved in many patients with meningitis; this raises the possibility that hyperventilation may cause further ischemic injury in those patients with marginal CBF. Although it is still unclear that treatment of increased ICP will affect outcome, we propose a treatment paradigm based on the results of neuroimaging studies and ICP measurements.

Abnormal Shoulder Girdle Muscle Tone in Premature Infants During Their First 18 Months of Life

To document the incidence of and neonatal factors associated with abnormal shoulder girdle muscle tone in premature infants at follow-up, we studied 125 consecutively admitted infants weighing less than 1,750 g treated in The Children's Hospital of Philadelphia intensive care nursery and subsequently seen in the Neonatal Follow-up Program up to 18 months of age. Fifty-seven infants (46%) displayed abnormal shoulder girdle muscle tone which presented clinically as scapular retractions. These infants had significantly lower birth weights (P less than .001) and gestational age (P less than .001) as well as a higher incidence of acute and chronic pulmonary disease (P less than 0.01) and CNS insults (P less than .05) when compared with infants without scapular retractions. The 57 infants with scapular retractions were further divided into two groups: 42 infants (74%) in whom scapular retractions were associated with generalized mild hypertonicity and 15 infants (26%) in whom scapular retractions compensated for trunk and neck hypotonicity. The infants with scapular retractions and hypotonicity had a significantly higher incidence of neonatal neurologic morbidity including seizures, major resuscitations, and birth asphyxia (P less than .01) when compared with the infants with scapular retractions and hypertonicity. Shoulder girdle tone abnormalities in the first year of life inhibit crawling, sitting, and object manipulation and, therefore, may manifest as delays in motor development. Identification of infants with significant neonatal risk factors for tone abnormalities is important to allow for earlier therapeutic intervention.