Many central nervous system (CNS) diseases display sexual dimorphism. Exposure to circulating sex steroids is felt to be a chief contributor to this phenomenon; however, CNS diseases of childhood and the elderly also demonstrate gender predominance and/or a sexually dimorphic response to therapies. Here we show that XY and XX neurons cultured separately are differentially susceptible to various cytotoxic agents and treatments. XY neurons were more sensitive to nitrosative stress and excitotoxicity versus XX neurons. In contrast, XX neurons were more sensitive to etoposide- and staurosporine-induced apoptosis versus XY neurons. The responses to specific therapies were also sexually dimorphic. Moreover, gender proclivity in programmed cell death pathway was observed. After cytotoxic challenge, programmed cell death proceeded predominately via an apoptosis-inducing factor-dependent pathway in XY neurons versus a cytochrome c-dependent pathway in XX neurons. This gender-dependent susceptibility is related to the incapacity of XY neurons to maintain intracellular levels of reduced glutathione. In vivo studies further demonstrated an incapacity for male, but not female, 17-day-old rats to maintain reduced glutathione levels within cerebral cortex acutely after an 8-min asphyxial cardiac arrest. This gender difference in sensitivity to cytotoxic agents may be generalized to nonneuronal cells, as splenocytes from male and female 16–18-day-old rats show similar gender-dependent responses to nitrosative stress and staurosporine-induced apoptosis. These data support gender stratification in the evaluation of mechanisms and treatment of CNS disease, particularly those where glutathione may play a role in detoxification, such as Parkinson's disease, traumatic brain injury, and conditions producing cerebral ischemia, and may apply to non-CNS diseases as well.

Autophagy is a homeostatic process for recycling of proteins and organelles, induced by nutrient deprivation and regulated by oxygen radicals. Whether autophagy is induced after traumatic brain injury (TBI) is not established. We show that TBI in mice results in increased ultrastructural and biochemical evidence of autophagy. Specifically, autophagosomal vacuoles and secondary lysosomes were frequently observed in cell processes and axons in ipsilateral brain regions by electron microscopy, and lipidated microtubule-associated protein light chain 3, a biochemical footprint of autophagy referred to as LC3 II, was increased at 2 and 24 h after TBI versus controls. Since oxygen radicals are believed to be important in the pathogenesis of TBI and are essential for the process of starvation-induced autophagy in vitro, we also sought to determine if treatment with the antioxidant γ-glutamylcysteinyl ethyl ester (GCEE) reduced autophagy and influenced neurologic outcome after TBI in mice. Treatment with GCEE reduced oxidative stress and partially reduced LC3 II formation in injured brain at 24 h after TBI versus vehicle. Treatment with GCEE also led to partial improvement in behavioral and histologic outcome versus vehicle. Taken together, these data show that autophagy occurs after experimental TBI, and that oxidative stress contributes to overall neuropathology, in part by initiating or influencing autophagy.

Febrile‐infection related epilepsy syndrome (FIRES) is a devastating neurological condition characterized by a febrile illness preceding new onset refractory status epilepticus (NORSE). Increasing evidence suggests innate immune dysfunction as a potential pathological mechanism. We report an international retrospective cohort of 25 children treated with anakinra, a recombinant interleukin‐1 receptor antagonist, as an immunomodulator for FIRES. Anakinra was potentially safe with only one child discontinuing therapy due to infection. Earlier anakinra initiation was associated with shorter duration of mechanical ventilation, ICU and hospital length of stay. Our retrospective data lay the groundwork for prospective consensus‐driven cohort studies of anakinra in FIRES.

J. Neurochem. (2008) 104, 1700–1711. Poly‐ADP‐ribosylation is a post‐translational modification performed by poly(ADP‐ribose) polymerases (PARP), involved in many diverse cellular functions including DNA repair, transcription, and long‐term potentiation. Paradoxically, PARP over‐activation under pathologic conditions including traumatic brain injury (TBI) results in cell death. We previously demonstrated that intra‐mitochondrial poly‐ADP‐ribosylation occurs following excitotoxic and oxidative injury in vitro. Here we sought to identify mitochondrial proteins modified by poly‐ADP‐ribosylation after TBI in vivo. Poly‐ADP‐ribosylation within mitochondria from injured brain after experimental TBI in rats was first verified using western blot and immuno‐electron microscopy. Poly‐ADP‐ribosylated mitochondrial proteins identified using a targeted proteomic approach included voltage‐dependent anion channel‐1, mitofilin, mitochondrial stress proteins, and the electron transport chain components F₁F₀ ATPase, cytochrome c oxidase, and cytochrome c reductase. To examine the functional consequences of mitochondrial poly‐ADP‐ribosylation, isolated rat brain mitochondria were exposed to conditions of nitrosative stress known to activate PARP. PARP activation‐induced reductions in State 3 respiration were prevented by the PARP‐1 inhibitor 5‐iodo‐6‐amino‐1,2‐benzopyrone or exogenous poly(ADP‐ribose) glycohydrolase. As the effects of PARP activation on mitochondrial respiration appear regulated by poly(ADP‐ribose) glycohydrolase, a direct effect of poly‐ADP‐ribosylation on electron transport chain function is suggested. These findings may be of relevance to TBI and other diseases where mitochondrial dysfunction occurs.

Previous studies suggest that delayed neuronal death occurs in patients with inflicted traumatic brain injury (TBI) from child abuse. It is unknown whether the mode of this delayed neuronal death represents apoptosis or necrosis, a distinction that carries therapeutic ramifications. Cytochrome c, an electron transport chain component, can be released from mitochondria under conditions of cellular stress, whereupon it can initiate and serve as a biomarker of apoptosis. To resolve this issue, cytochrome c concentration was determined in 167 ventricular cerebrospinal fluid (CSF) samples from 67 infants and children with TBI (including 15 patients diagnosed with child abuse) by ELISA. Controls included lumbar CSF from 19 infants and children without trauma or meningitis. A multivariate model adjusted for multiple within-subject observations was used to identify clinical variables associated with CSF cytochrome c. Other apoptosis-related proteins were also examined in a subset of TBI patients. Increased CSF cytochrome c was independently associated with inflicted TBI (P=0.0001) and female gender (P=0.04), but not age, Glasgow coma scale score, or survival. Other apoptosis-related proteins including Fas and caspase-1 were increased in CSF after TBI, but did not independently discriminate between accidental and inflicted TBI. These data suggest that apoptosis, as detected by the presence of cytochrome c in CSF, is uniquely prominent among the subset of TBI patients diagnosed with child abuse. The degree of apoptosis after TBI also appears to be gender-dependent. Development of strategies targeting apoptosis after TBI, particularly in victims of child abuse and in girls, appears justified.

Objective Develop a clinically relevant model of pediatric asphyxial cardiopulmonary arrest in rats. Design Prospective interventional study. Setting University research laboratory. Subjects Postnatal day 16–18 rats (n = 9/group). Interventions Anesthetized rats were endotracheally intubated and mechanically ventilated, and vascular catheters were inserted. Vecuronium was administered, and the ventilator was disconnected from the rats for 8 mins, whereupon rats were resuscitated with epinephrine, sodium bicarbonate, and chest compressions until spontaneous circulation returned. Shams underwent all procedures except asphyxia. Measurements and Main Results Asphyxial arrest typically occurred by 1 min after the ventilator was disconnected. Return of spontaneous circulation typically occurred <30 secs after resuscitation. An isoelectric electroencephalograph was observed for 30 mins after asphyxia, and rats remained comatose for 12–24 hrs. Overall survival rate was 85%. Motor function measured using beam balance and inclined plane tests was impaired on days 1 and 2, but recovered by day 3, in rats after asphyxia vs. sham injury (p < .05). Spatial memory acquisition measured using the Morris-water maze on days 7–14 and 28–35 was also impaired in rats after asphyxia vs. sham injury (total latency 379 ± 28 vs. 501 ± 40 secs, respectively, p < .05). DNA fragmentation was detected in CA1 hippocampal neurons bilaterally 3–7 days after asphyxia. Neurodegeneration detected using Fluorojade B was seen in bilateral CA1 hippocampi and layer V cortical neurons 3–7 days after asphyxia, with persistent neurodegeneration in CA1 hippocampus detected up to 5 wks after asphyxia. CA1 hippocampal neuron survival after asphyxia was 39–43% (p < .001 vs. sham). Evidence of DNA or cellular injury was not detected in sham rats. Conclusions This model of asphyxial cardiopulmonary arrest in postnatal day 17 rats produces many of the clinical manifestations of pediatric hypoxic-ischemic encephalopathy. This model may be useful for the preclinical testing of novel and currently available interventions aimed at improving neurologic outcome in infants and children after cardiopulmonary arrest.