Ansgar M. Brambrink

Neurodegeneration in Excitotoxicity, Global Cerebral Ischemia, and Target Deprivation: A Perspective on the Contributions of Apoptosis and Necrosis

In the human brain and spinal cord, neurons degenerate after acute insults (e.g., stroke, cardiac arrest, trauma) and during progressive, adult-onset diseases [e.g., amyotrophic lateral sclerosis, Alzheimers disease]. Glutamate receptor-mediated excitotoxicity has been implicated in all of these neurological conditions. Nevertheless, effective approaches to prevent or limit neuronal damage in these disorders remain elusive, primarily because of an incomplete understanding of the mechanisms of neuronal death in in vivo settings. Therefore, animal models of neurodegeneration are crucial for improving our understanding of the mechanisms of neuronal death. In this review, we evaluate experimental data on the general characteristics of cell death and, in particular, neuronal death in the central nervous system (CNS) following injury. We focus on the ongoing controversy of the contributions of apoptosis and necrosis in neurodegeneration and summarize new data from this laboratory on the classification of neuronal death using a variety of animal models of neurodegeneration in the immature or adult brain following excitotoxic injury, global cerebral ischemia, and axotomy/target deprivation. In these different models of brain injury, we determined whether the process of neuronal death has uniformly similar morphological characteristics or whether the features of neurodegeneration induced by different insults are distinct. We classified neurodegeneration in each of these models with respect to whether it resembles apoptosis, necrosis, or an intermediate form of cell death falling along an apoptosis-necrosis continuum. We found that N-methyl-D-aspartate (NMDA) receptor- and non-NMDA receptor-mediated excitotoxic injury results in neurodegeneration along an apoptosis-necrosis continuum, in which neuronal death (appearing as apoptotic, necrotic, or intermediate between the two extremes) is influenced by the degree of brain maturity and the subtype of glutamate receptor that is stimulated. Global cerebral ischemia produces neuronal death that has commonalities with excitotoxicity and target deprivation. Degeneration of selectively vulnerable populations of neurons after ischemia is morphologically nonapoptotic and is indistinguishable from NMDA receptor-mediated excitotoxic death of mature neurons. However, prominent apoptotic cell death occurs following global ischemia in neuronal groups that are interconnected with selectively vulnerable populations of neurons and also in nonneuronal cells. This apoptotic neuronal death is similar to some forms of retrograde neuronal apoptosis that occur following target deprivation. We conclude that cell death in the CNS following injury can coexist as apoptosis, necrosis, and hybrid forms along an apoptosis-necrosis continuum. These different forms of cell death have varying contributions to the neuropathology resulting from excitotoxicity, cerebral ischemia, and target deprivation/axotomy. Degeneration of different populations of cells (neurons and nonneuronal cells) may be mediated by distinct or common causal mechanisms that can temporally overlap and perhaps differ mechanistically in the rate of progression of cell death.

Isoflurane-induced Neuroapoptosis in the Neonatal Rhesus Macaque Brain

Background:Brief isoflurane anesthesia induces neuroapoptosis in the developing rodent brain, but susceptibility of non-human primates to the apoptogenic action of isoflurane has not been studied. Therefore, we exposed postnatal day 6 (P6) rhesus macaques to a surgical plane of isoflurane anesthesia for 5 h, and studied the brains 3 h later for histopathologic changes. Method:With the same intensity of physiologic monitoring typical for human neonatal anesthesia, five P6 rhesus macaques were exposed for 5 h to isoflurane maintained between 0.7 and 1.5 end-tidal Vol% (endotracheally intubated and mechanically ventilated) and five controls were exposed for 5 h to room air without further intervention. Three hours later, the brains were harvested and serially sectioned across the entire forebrain and midbrain, and stained immunohistochemically with antibodies to activated caspase-3 for detection and quantification of apoptotic neurons. Results:Quantitative evaluation of brain sections revealed a median of 32.5 (range, 18.0-48.2) apoptotic cells/mm3 of brain tissue in the isoflurane group and only 2.5 (range, 1.1-5.2) in the control group (difference significant at P = 0.008). Apoptotic neuronal profiles were largely confined to the cerebral cortex. In the control brains, they were sparse and randomly distributed, whereas in the isoflurane brains they were abundant and preferentially concentrated in specific cortical layers and regions. Conclusion:The developing non-human primate brain is sensitive to the apoptogenic action of isoflurane and displays a 13-fold increase in neuroapoptosis after 5 h exposure to a surgical plane of isoflurane anesthesia.

Routine Clinical Practice Effectiveness of the Glidescope in Difficult Airway Management An Analysis of 2,004 Glidescope Intubations, Complications, and Failures from Two Institutions

Introduction: The Glidescope video laryngoscope has been shown to be a useful tool to improve laryngeal view. However, its role in the daily routine of airway management remains poorly characterized. Methods: This investigation evaluated the use of the Glidescope at two academic medical centers. Electronic records from 71,570 intubations were reviewed, and 2,004 cases were indentified where the Glidescope was used for airway management. We analyzed the success rate of Glidescope intubation in various intubation scenarios. In addition, the incidence and character of complications associated with Glidescope use were recorded. Predictors of Glidescope intubation failure were determined using a logistic regression analysis. Results: Overall success for Glidescope intubation was 97% (1,944 of 2,004). As a primary technique, success was 98% (1,712 of 1,755), whereas success in patients with predictors of difficult direct laryngoscopy was 96% (1,377 of 1,428). Success for Glidescope intubation after failed direct laryngoscopy was 94% (224 of 239). Complications were noticed in 1% (21 of 2,004) of patients and mostly involved minor soft tissue injuries, but major complications, such as dental, pharyngeal, tracheal, or laryngeal injury, occurred in 0.3% (6 of 2,004) of patients. The strongest predictor of Glidescope failure was altered neck anatomy with presence of a surgical scar, radiation changes, or mass. Conclusion: These data demonstrate a high success rate of Glidescope intubation in both primary airway management and rescue-failed direct laryngoscopy. However, Glidescope intubation is not always successful and certain predictors of failure can be identified. Providers should maintain their competency with alternate methods of intubation, especially for patients with neck pathology.

Comparative Effectiveness of the C-MAC Video Laryngoscope versus Direct Laryngoscopy in the Setting of the Predicted Difficult Airway

Background: Video laryngoscopy may be useful in the setting of the difficult airway, but it remains unclear if intubation success is improved in routine difficult airway management. This study compared success rates for tracheal intubation with the C-MAC® video laryngoscope (Karl Storz, Tuttlingen, Germany) with conventional direct laryngoscopy in patients with predicted difficult airway. Methods: We conducted a two arm, single-blinded randomized controlled trial that involved 300 patients. Inclusion required at least one of four predictors of difficult intubation. The primary outcome was successful tracheal intubation on first attempt. Results: The use of video laryngoscopy resulted in more successful intubations on first attempt (138/149; 93%) as compared with direct laryngoscopy (124/147; 84%), P = 0.026. Cormack-Lehane laryngeal view was graded I or II in 139/149 of C-MAC attempts versus 119/147 in direct laryngoscopy attempts (P < 0.01). Laryngoscopy time averaged 46 s (95% CI, 40–51) for the C-MAC group and was shorter in the direct laryngoscopy group, 33 s (95% CI, 29–36), P < 0.001. The use of a gum-elastic bougie and/or external laryngeal manipulation were required less often in the C-MAC intubations (24%, 33/138) compared with direct laryngoscopy (37%, 46/124, P = 0.020). The incidence of complications was not significantly different between the C-MAC (20%, 27/138) versus direct laryngoscopy (13%, 16/124, P = 0.146). Conclusion: A diverse group of anesthesia providers achieved a higher intubation success rate on first attempt with the C-MAC in a broad range of patients with predictors of difficult intubation. C-MAC laryngoscopy seems to be a useful technique for the initial approach to a potentially difficult airway.

Propofol-induced apoptosis of neurones and oligodendrocytes in fetal and neonatal rhesus macaque brain

BACKGROUND Exposure of the fetal or neonatal non-human primate (NHP) brain to isoflurane or ketamine for 5 h causes widespread apoptotic degeneration of neurones, and exposure to isoflurane also causes apoptotic degeneration of oligodendrocytes (OLs). The present study explored the apoptogenic potential of propofol in the fetal and neonatal NHP brain. METHOD Fetal rhesus macaques at gestational age 120 days were exposed in utero, or postnatal day 6 rhesus neonates were exposed directly for 5 h to propofol anaesthesia (n=4 fetuses; and n=4 neonates) or to no anaesthesia (n=4 fetuses; n=5 neonates), and the brains were systematically evaluated 3 h later for evidence of apoptotic degeneration of neurones or glia. RESULTS Exposure of fetal or neonatal NHP brain to propofol caused a significant increase in apoptosis of neurones, and of OLs at a stage when OLs were just beginning to myelinate axons. Apoptotic degeneration affected similar brain regions but to a lesser extent than we previously described after isoflurane. The number of OLs affected by propofol was approximately equal to the number of neurones affected at both developmental ages. In the fetus, neuroapoptosis affected particularly subcortical and caudal regions, while in the neonate injury involved neocortical regions in a distinct laminar pattern and caudal brain regions were less affected. CONCLUSIONS Propofol anaesthesia for 5 h caused death of neurones and OLs in both the fetal and neonatal NHP brain. OLs become vulnerable to the apoptogenic action of propofol when they are beginning to achieve myelination competence.

Isoflurane-induced apoptosis of oligodendrocytes in the neonatal primate brain

Previously we reported that exposure of 6‐day‐old (P6) rhesus macaques to isoflurane for 5 hours triggers a robust neuroapoptosis response in developing brain. We have also observed (unpublished data) that isoflurane causes apoptosis of cellular profiles in the white matter that resemble glia. We analyzed the cellular identity of the apoptotic white matter profiles and determined the magnitude of this cell death response to isoflurane.

Primary sensory and forebrain motor systems in the newborn brain are preferentially damaged by hypoxia‐ischemia

Cerebral hypoxia‐ischemia causes encephalopathy and neurologic disabilities in newborns by unclear mechanisms. We tested the hypothesis that hypoxia‐ischemia causes brain damage in newborns that is system‐preferential and related to regional oxidative metabolism. One‐week‐old piglets were subjected to 30 minutes of hypoxia and then seven minutes of airway occlusion, producing asphyxic cardiac arrest, followed by cardiopulmonary resuscitation and four‐day recovery. Brain injury in hypoxic‐ischemic piglets (n = 6) compared to controls (n = 5) was analyzed by hematoxylin‐eosin, Nissl, and silver staining; relationships between regional vulnerability and oxidative metabolism were evaluated by cytochrome oxidase histochemistry. Profile counting‐based estimates showed that 13% and 27% of neurons in layers II/III and layers IV/V of somatosensory cortex had ischemic cytopathology, respectively; CA1 neuronal perikarya appeared undamaged, and <10% of CA3 and CA4 neurons were injured; and neuronal damage was 79% in putamen, 17% in caudate, but nucleus accumbens was undamaged. Injury was found preferentially in primary sensory neocortices (particularly somatosensory cortex), basal ganglia (predominantly putamen, subthalamic nucleus, and substantia nigra reticulata), ventral thalamus, geniculate nuclei, and tectal nuclei. In sham piglets, vulnerable regions generally had higher cytochrome oxidase levels than less vulnerable areas. Postischemic alterations in cytochrome oxidase were regional and laminar, with reductions (31–66%) occurring in vulnerable regions and increases (20%) in less vulnerable areas. We conclude that neonatal hypoxia‐ischemia causes highly organized, system‐preferential and topographic encephalopathy, targeting regions that function in sensorimotor integration and movement control. This distribution of neonatal encephalopathy is dictated possibly by regional function, mitochondrial activity, and connectivity. J. Comp. Neurol. 377:262–285, 1997.

Brain protection by anesthetic agents.

Purpose of review Patients at risk for perioperative stroke, or those who have suffered recent cerebral injury, may benefit from neuroprotective properties of anesthetic agents during surgery. This manuscript reviews recent clinical and experimental evidence for neuroprotective effects of common anesthetic agents, and presents potential mechanisms involved in anesthetic neuroprotection. Recent findings Although strong experimental data support a neuroprotective potential of several anesthetic agents, specifically isoflurane and xenon, consistent long-term protection by either agent has not been demonstrated. Unfortunately, there is a lack of clinical studies that would support the use of any one anesthetic agent over the others. Mechanisms of neuroprotection by anesthetic agents appear to involve suppression of excitatory neurotransmission, and potentiation of inhibitory activity, which may contribute to the reduction of excitotoxic injury. Activation of intracellular signaling cascades that lead to altered expression of protective genes may also be involved. Summary Solid experimental evidence supports neuroprotection by anesthetic agents. It is too early to recommend any specific agent for clinical use as a neuroprotectant, however. Further study is warranted to unravel relevant mechanisms and to appreciate the potential clinical relevance of experimental findings.

Isoflurane-induced Apoptosis of Neurons and Oligodendrocytes in the Fetal Rhesus Macaque Brain

Background:The authors have previously shown that exposure of the neonatal nonhuman primate (NHP) brain to isoflurane for 5 h causes widespread acute apoptotic degeneration of neurons and oligodendrocyte. The current study explored the potential apoptogenic action of isoflurane in the fetal NHP brain. Methods:Fetal rhesus macaques at gestational age of 120 days (G120) were exposed in utero for 5 h to isoflurane anesthesia (n = 5) or to no anesthesia (control condition; n = 4), and all regions of the brain were systematically evaluated 3 h later for evidence of apoptotic degeneration of neurons or glia. Results:Exposure of the G120 fetal NHP brain to isoflurane caused a significant increase in apoptosis of neurons and of oligodendrocytes at a stage when oligodendrocytes were just beginning to myelinate axons. The neuroapoptosis response was most prominent in the cerebellum, caudate, putamen, amygdala, and several cerebrocortical regions. Oligodendrocyte apoptosis was diffusely distributed over many white matter regions. The total number of apoptotic profiles (neurons + oligodendrocytes) in the isoflurane-exposed brains was increased 4.1-fold, compared with the brains from drug-naive controls. The total number of oligodendrocytes deleted by isoflurane was higher than the number of neurons deleted. Conclusions:Isoflurane anesthesia for 5 h causes death of neurons and oligodendrocytes in the G120 fetal NHP brain. In the fetal brain, as the authors previously found in the neonatal NHP brain, oligodendrocytes become vulnerable when they are just achieving myelination competence. The neurotoxic potential of isoflurane increases between the third trimester (G120) and the neonatal period in the NHP brain.

Effect of arrest time and cerebral perfusion pressure during cardiopulmonary resuscitation on cerebral blood flow, metabolism, adenosine triphosphate recovery, and pH in dogs

OBJECTIVES To test the hypothesis that greater cerebral perfusion pressure (CPP) is required to restore cerebral blood flow (CBF), oxygen metabolism, adenosine triphosphate (ATP), and intracellular pH (pHi) levels after variable periods of no-flow than to maintain them when cardiopulmonary resuscitation (CPR) is started immediately. DESIGN Prospective, randomized, comparison of three arrest times and two perfusion pressures during CPR in 24 anesthetized dogs. SETTING University cerebral resuscitation laboratory. INTERVENTIONS We used radiolabeled microspheres to determine CBF and magnetic resonance spectroscopy to derive ATP and pHi levels before and during CPR. Ventricular fibrillation was induced, epinephrine administered, and thoracic vest CPR adjusted to provide a CPP of 25 or 35 mm Hg after arrest times of O, 6, or 12 mins. MEASUREMENTS AND MAIN RESULTS When CPR was started immediately after arrest with a CPP of 25 mm Hg, CBF and ATP were 57 +/- 10% and 64 +/- 14% of prearrest (at 10 mins of CPR). In contrast, CBF and ATP were minimally restored with a CPP at 25 mm Hg after a 6-min arrest time (23 +/- 5%, 16 +/- 5%, respectively). With a CPP of 35 mm Hg, extending the no-flow arrest time from 6 to 12 mins reduced reflow from 71 +/- 11% to 37 +/- 7% of pre-arrest and reduced ATP recovery from 60 +/- 11% to 2 +/- 1% of pre-arrest. After 6- or 12-min arrest times, brainstem blood flow was restored more than supratentorial blood flow, but cerebral pHi was never restored. CONCLUSIONS A CPP of 25 mm Hg maintains supratentorial blood flow and ATP at 60% to 70% when CPR starts immediately on arrest, but not after a 6-min delay. A higher CPP of 35 mm Hg is required to restore CBF and ATP when CPR is delayed for 6 mins. After a 12-min delay, even the CPP of 35 mm Hg is unable to restore CBF and ATP. Therefore, increasing the arrest time at these perfusion pressures increases the resistance to reflow sufficient to impair restoration of cerebral ATP.