Correlates of Spreading Depolarization, Spreading Depression, and Negative Ultraslow Potential in Epidural Versus Subdural Electrocorticography

Spreading depolarizations (SDs) are characterized by near-complete breakdown of the transmembrane ion gradients, neuronal oedema and activity loss (=depression). The SD extreme in ischemic tissue, termed 'terminal SD,' shows prolonged depolarization, in addition to a slow baseline variation called 'negative ultraslow potential' (NUP). The NUP is the largest bioelectrical signal ever recorded from the human brain and is thought to reflect the progressive recruitment of neurons into death in the wake of SD. However, it is unclear whether the NUP is a field potential or results from contaminating sensitivities of platinum electrodes. In contrast to Ag/AgCl-based electrodes in animals, platinum/iridium electrodes are the gold standard for intracranial direct current (DC) recordings in humans. Here, we investigated the full continuum including short-lasting SDs under normoxia, long-lasting SDs under systemic hypoxia, and terminal SD under severe global ischemia using platinum/iridium electrodes in rats to better understand their recording characteristics. Sensitivities for detecting SDs or NUPs were 100% for both electrode types. Nonetheless, the platinum/iridium-recorded NUP was 10 times smaller in rats than humans. The SD continuum was then further investigated by comparing subdural platinum/iridium and epidural titanium peg electrodes in patients. In seven patients with either aneurysmal subarachnoid hemorrhage or malignant hemispheric stroke, two epidural peg electrodes were placed 10 mm from a subdural strip. We found that 31/67 SDs (46%) on the subdural strip were also detected epidurally. SDs that had longer negative DC shifts and spread more widely across the subdural strip were more likely to be observed in epidural recordings. One patient displayed an SD-initiated NUP while undergoing brain death despite continued circulatory function. The NUP's amplitude was -150 mV subdurally and -67 mV epidurally. This suggests that the human NUP is a bioelectrical field potential rather than an artifact of electrode sensitivity to other factors, since the dura separates the epidural from the subdural compartment and the epidural microenvironment was unlikely changed, given that ventilation, arterial pressure and peripheral oxygen saturation remained constant during the NUP. Our data provide further evidence for the clinical value of invasive electrocorticographic monitoring, highlighting important possibilities as well as limitations of less invasive recording techniques

Correlates of Spreading Depolarization, Spreading Depression, and Negative Ultraslow Potential in Epidural Versus Subdural Electrocorticography

Spreading depolarizations (SDs) are characterized by near-complete breakdown of the transmembrane ion gradients, neuronal oedema and activity loss (=depression). The SD extreme in ischemic tissue, termed ‘terminal SD,’ shows prolonged depolarization, in addition to a slow baseline variation called ‘negative ultraslow potential’ (NUP). The NUP is the largest bioelectrical signal ever recorded from the human brain and is thought to reflect the progressive recruitment of neurons into death in the wake of SD. However, it is unclear whether the NUP is a field potential or results from contaminating sensitivities of platinum electrodes. In contrast to Ag/AgCl-based electrodes in animals, platinum/iridium electrodes are the gold standard for intracranial direct current (DC) recordings in humans. Here, we investigated the full continuum including short-lasting SDs under normoxia, long-lasting SDs under systemic hypoxia, and terminal SD under severe global ischemia using platinum/iridium electrodes in rats to better understand their recording characteristics. Sensitivities for detecting SDs or NUPs were 100% for both electrode types. Nonetheless, the platinum/iridium-recorded NUP was 10 times smaller in rats than humans. The SD continuum was then further investigated by comparing subdural platinum/iridium and epidural titanium peg electrodes in patients. In seven patients with either aneurysmal subarachnoid hemorrhage or malignant hemispheric stroke, two epidural peg electrodes were placed 10 mm from a subdural strip. We found that 31/67 SDs (46%) on the subdural strip were also detected epidurally. SDs that had longer negative DC shifts and spread more widely across the subdural strip were more likely to be observed in epidural recordings. One patient displayed an SD-initiated NUP while undergoing brain death despite continued circulatory function. The NUP’s amplitude was -150 mV subdurally and -67 mV epidurally. This suggests that the human NUP is a bioelectrical field potential rather than an artifact of electrode sensitivity to other factors, since the dura separates the epidural from the subdural compartment and the epidural microenvironment was unlikely changed, given that ventilation, arterial pressure and peripheral oxygen saturation remained constant during the NUP. Our data provide further evidence for the clinical value of invasive electrocorticographic monitoring, highlighting important possibilities as well as limitations of less invasive recording techniques

Correlation of velocity and susceptibility in patients with aneurysmal subarachnoid hemorrhage

In many cerebral grey matter structures including the neocortex, spreading depolarization (SD) is the principal mechanism of the near-complete breakdown of the transcellular ion gradients with abrupt water influx into neurons. Accordingly, SDs are abundantly recorded in patients with traumatic brain injury, spontaneous intracerebral hemorrhage, aneurysmal subarachnoid hemorrhage (aSAH) and malignant hemispheric stroke using subdural electrode strips. SD is observed as a large slow potential change, spreading in the cortex at velocities between 2 and 9 mm/min. Velocity and SD susceptibility typically correlate positively in various animal models. In patients monitored in neurocritical care, the Co-Operative Studies on Brain Injury Depolarizations (COSBID) recommends several variables to quantify SD occurrence and susceptibility, although accurate measures of SD velocity have not been possible. Therefore, we developed an algorithm to estimate SD velocities based on reconstructing SD trajectories of the wave-front's curvature center from magnetic resonance imaging scans and time-of-SD-arrival- differences between subdural electrode pairs. We then correlated variables indicating SD susceptibility with algorithm-estimated SD velocities in twelve aSAH patients. Highly significant correlations supported the algorithm's validity. The trajectory search failed significantly more often for SDs recorded directly over emerging focal brain lesions suggesting in humans similar to animals that the complexity of SD propagation paths increase in tissue undergoing injury

Stimulation of the Sphenopalatine Ganglion Induces Reperfusion and Blood-Brain Barrier Protection in the Photothrombotic Stroke Model

The treatment of stroke remains a challenge. Animal studies showing that electrical stimulation of the sphenopalatine ganglion (SPG) exerts beneficial effects in the treatment of stroke have led to the initiation of clinical studies. However, the detailed effects of SPG stimulation on the injured brain are not known.The effect of acute SPG stimulation was studied by direct vascular imaging, fluorescent angiography and laser Doppler flowmetry in the sensory motor cortex of the anaesthetized rat. Focal cerebral ischemia was induced by the rose bengal (RB) photothrombosis method. In chronic experiments, SPG stimulation, starting 15 min or 24 h after photothrombosis, was given for 3 h per day on four consecutive days. Structural damage was assessed using histological and immunohistochemical methods. Cortical functions were assessed by quantitative analysis of epidural electro-corticographic (ECoG) activity continuously recorded in behaving animals.Stimulation induced intensity- and duration-dependent vasodilation and increased cerebral blood flow in both healthy and photothrombotic brains. In SPG-stimulated rats both blood brain-barrier (BBB) opening, pathological brain activity and lesion volume were attenuated compared to untreated stroke animals, with no apparent difference in the glial response surrounding the necrotic lesion.SPG-stimulation in rats induces vasodilation of cortical arterioles, partial reperfusion of the ischemic lesion, and normalization of brain functions with reduced BBB dysfunction and stroke volume. These findings support the potential therapeutic effect of SPG stimulation in focal cerebral ischemia even when applied 24 h after stroke onset and thus may extend the therapeutic window of currently administered stroke medications

Abstracts from the 20th International Symposium on Signal Transduction at the Blood-Brain Barriers

https://deepblue.lib.umich.edu/bitstream/2027.42/138963/1/12987_2017_Article_71.pd

Dynamiken und Mechanismen der Blut-Hirnschranken-Störung, des Zellschadens und der Veränderungen der zerebralen Perfusion nach kortikaler Photothrombose

Focal cerebral ischemia is one of the main causes of death and disability worldwide. An ischemic core and a surrounding dysfunctional region that is susceptible to cell injury often characterize the lesion. Assessing the propensity of the peri-ischemic brain to undergo secondary damage, understanding the underlying mechanisms, and adjusting treatment accordingly remain clinically unmet challenges. A significant hallmark of the peri- ischemic brain is a dysfunctional blood–brain barrier (BBB), yet the role of disturbed vascular permeability in stroke progression and for functional outcome is largely unclear. Here I describe a longitudinal in vivo fluorescence imaging approach for the evaluation of cortical perfusion, BBB dysfunction, free radical formation and cellular injury in the cortical photothrombosis model in male Sprague Dawley rats. In this model light- activated (532 nm) Rose Bengal initiates clot formation thereby preventing sufficient perfusion to the site of illumination. Cerebral perfusion and BBB permeability were quantified from intra- and extravascular distribution kinetics of fluorescein sodium salt following its intravenous bolus application. In parallel, propidium iodide − a membrane integrity marker − served as a cell damage marker and was compared to staining with annexin V, which binds to phosphatidylserin following its exposure on the outer leaflet of the membrane during cellular injury and death. Reactive oxygen and nitrogen species were visualized using ROSstar 650 and 4-amino-5-methylamino-2’,7’-difluorofluorescein diacetate and reduced using phenyl-N-t-butylnitrone, L-NGnitroarginine methyl ester and carboxy-PTIO. Imaging the peri-ischemic area demonstrated propagation and progression of hypoperfusion, BBB dysfunction and cell damage during the 3 hour monitoring period following photothrombosis. While hypoperfusion was identified within a belt of ~400 μm surrounding the ischemic core, blood-brain barrier dysfunction and cell damage also occurred at greater distances. Nitric oxide formation most prominently occurred in arterioles, whereas superoxide and hydroxyl radicals gave a diffuse peri-ischemic parenchymal signal. Inhibiting free radical signaling significantly reduced progressive cellular damage after photothrombosis, with no significant effect on blood flow changes and increases in BBB permeability. Of note, cellular injury was only prevented in normally perfused more distant peri-ischemic brain regions (> 400 μm away from the ischemic core). Hence our data is in agreement with previous studies stressing perfusion based imaging as insufficient to detect tissue at risk. Measurements of BBB permeability could serve as a novel approach to predict lesion progression. In addition, our approach allows a dynamic follow-up of cellular events and their response to therapeutics in the acutely injured cerebral cortex.Schlaganfall ist weltweit eine der Hauptursachen von Morbidität und Mortalität. Ein ischämischer Kern und eine weiter gefasste funktionsgestörte Region, in der es zu sekundärem Zellschaden kommen kann, kennzeichnen dieses Krankheitsbild. Die Anfälligkeit für sekundären Zellschaden zu erfassen, zugrunde liegende Mechanismen herauszufinden und Therapien entsprechend anzupassen, bleiben jedoch Herausforderungen der gegenwärtigen Forschung. Eine eingeschränkte Blut-Hirn-Schranken-Funktion ist im peri-ischämischen Gewebe üblich. Es bleibt jedoch weitgehend unklar, inwieweit diese zur Vergrößerung des Infarktkerns beiträgt und dadurch die klinische Genesung beeinflusst. In dieser Arbeit beschreibe ich eine longitudinale Methode der in vivo Fluoreszenzbildgebung zur Evaluierung des zerebralen Blutflusses, der Blut- Hirn-Schranken- Permeabilität, der Bildung freier Radikale und des Zelltodes im kortikalen Photothrombosemodell an männlichen Sprague Dawley Ratten. In diesem Modell führt lichtaktiviertes Bengalrosa zur Thrombusformation und verhindert so die ausreichende Blutzufuhr in zuvor illuminiertes Gewebe. Die zerebrale Perfusion und die Blut-Hirn-Schranken-Permeabilität wurden anhand der intra- und extravaskulären Verteilungskinetik von zuvor intravenös injiziertem Fluoresceinsalz quantifiziert. Parallel diente Propidiumjodid, ein Membranintegritätsmarker, als Indikator für Zellschaden. Dieser wurde mit einem zweiten Marker, Annexin V, verglichen. Annexin V bindet an Phosphatidylserin, welches bei Zellschaden und -tod vom Zellinneren an die äußere Lage der Lipiddoppelmembran transloziert wird. Sauerstoff- und Stickstoffradikale wurden durch die Farbstoffe ROSstar 650 und 4-Amino-5-Methylamino-2’,7’-Difluorofluorescein-Diacetat sichtbar gemacht, und deren Vorkommen im Gewebe bzw. deren Bildung wurde durch Phenyl-N-t-butylnitron,L-NG-Nitroargininmethylester und Carboxy-PTIO reduziert. Das peri-ischämische Gewebe wurde für drei Stunden nach Induktion der Photothrombose bildgebend überwacht. Innerhalb dieses Zeitraums kam es zu Propagation und Progression von Hypoperfusion, Blut-Hirn-Schrankenstörung und Zellschaden. Während hypoperfundiertes Gewebe nur in einem Abstand von ca. 400 μm um den Infarktkern detektiert wurde, war auch weiter entferntes Gewebe von einer Blut-Hirn-Schranken-Störung und Zellschaden betroffen. Stickstoffmonoxid stieg am prominentesten in Arteriolen an, während Superoxid- und Hydroxylradikale ein diffuses parenchymales peri-ischämisches Signal gaben. Die Blockade freier Radikale reduzierte die Progression von Zellschaden, hatte jedoch weder Einfluss auf die Perfusion noch auf die Blut-Hirn-Schranken- Permeabilität. Interessanterweise wurde die Ausbreitung des Zellschadens nur in normal perfundiertem peri-ischämischem Gewebe (> 400 μm vom Infarktkern entfernt) reduziert. Damit stimmen die Daten mit solchen Studien überein, die betonen, dass perfusionsbasierte Bildgebung nur unzureichend erfasst, ob Gewebe von Sekundärschaden bedroht ist. Messungen der Blut-Hirn- Schrankenpermeabilität könnten als zusätzlicher neuer Ansatz zur Abschätzung der Schadensprogression dienen. Die longitudinale Bildgebung, wie hier im Tiermodell beschrieben, macht es möglich, zelluläre Ereignisse im akut geschädigten Hirngewebe in ihrer Dynamik zu überwachen und den Effekt möglicher Therapeutika zu überprüfen

Event-Associated Oxygen Consumption Rate Increases ca. Five-Fold When Interictal Activity Transforms into Seizure-Like Events In Vitro

Neuronal injury due to seizures may result from a mismatch of energy demand and adenosine triphosphate (ATP) synthesis. However, ATP demand and oxygen consumption rates have not been accurately determined, yet, for different patterns of epileptic activity, such as interictal and ictal events. We studied interictal-like and seizure-like epileptiform activity induced by the GABAA antagonist bicuculline alone, and with co-application of the M-current blocker XE-991, in rat hippocampal slices. Metabolic changes were investigated based on recording partial oxygen pressure, extracellular potassium concentration, and intracellular flavine adenine dinucleotide (FAD) redox potential. Recorded data were used to calculate oxygen consumption and relative ATP consumption rates, cellular ATP depletion, and changes in FAD/FADH2 ratio by applying a reactive-diffusion and a two compartment metabolic model. Oxygen-consumption rates were ca. five times higher during seizure activity than interictal activity. Additionally, ATP consumption was higher during seizure activity (~94% above control) than interictal activity (~15% above control). Modeling of FAD transients based on partial pressure of oxygen recordings confirmed increased energy demand during both seizure and interictal activity and predicted actual FAD autofluorescence recordings, thereby validating the model. Quantifying metabolic alterations during epileptiform activity has translational relevance as it may help to understand the contribution of energy supply and demand mismatches to seizure-induced injury

Imaging blood\u2013brain barrier dysfunction in animal disease models

The blood\u2013brain barrier (BBB) is a highly complex structure, which separates the extracellular fluid of the central nervous system (CNS) from the blood of CNS vessels. A wide range of neurologic conditions, including stroke, epilepsy, Alzheimer\u2019s disease, and brain tumors, are associated with perturbations of the BBB that contribute to their pathology. The common consequence of a BBB dysfunction is increased permeability, leading to extravasation of plasma constituents and vasogenic brain edema. The BBB impairment can persist for long periods, being involved in secondary inflammation and neuronal dysfunction, thus contributing to disease pathogenesis. Therefore, reliable imaging of the BBB impairment is of major importance in both clinical management of brain diseases and in experimental research. From landmark studies by Ehrlich and Goldman, the use of dyes (probes) has played a critical role in understanding BBB functions. In recent years methodologic advances in morphologic and functional brain imaging have provided insight into cellular and molecular interactions underlying BBB dysfunction in animal disease models. These imaging techniques, which range from in situ staining to noninvasive in vivo imaging, have different spatial resolution, sensitivity, and capacity for quantitative and kinetic measures of the BBB impairment. Despite significant advances, the translation of these techniques into clinical applications remains slow. This review outlines key recent advances in imaging techniques that have contributed to the understanding of BBB dysfunction in disease and discusses major obstacles and opportunities to advance these techniques into the clinical realm.Peer reviewed: YesNRC publication: Ye

Event-Associated Oxygen Consumption Rate Increases ca. Five-Fold When Interictal Activity Transforms into Seizure-Like Events In Vitro

Neuronal injury due to seizures may result from a mismatch of energy demand and adenosine triphosphate (ATP) synthesis. However, ATP demand and oxygen consumption rates have not been accurately determined, yet, for different patterns of epileptic activity, such as interictal and ictal events. We studied interictal-like and seizure-like epileptiform activity induced by the GABAA antagonist bicuculline alone, and with co-application of the M-current blocker XE-991, in rat hippocampal slices. Metabolic changes were investigated based on recording partial oxygen pressure, extracellular potassium concentration, and intracellular flavine adenine dinucleotide (FAD) redox potential. Recorded data were used to calculate oxygen consumption and relative ATP consumption rates, cellular ATP depletion, and changes in FAD/FADH2 ratio by applying a reactive-diffusion and a two compartment metabolic model. Oxygen-consumption rates were ca. five times higher during seizure activity than interictal activity. Additionally, ATP consumption was higher during seizure activity (~94% above control) than interictal activity (~15% above control). Modeling of FAD transients based on partial pressure of oxygen recordings confirmed increased energy demand during both seizure and interictal activity and predicted actual FAD autofluorescence recordings, thereby validating the model. Quantifying metabolic alterations during epileptiform activity has translational relevance as it may help to understand the contribution of energy supply and demand mismatches to seizure-induced injury

Low neuronal metabolism during isoflurane-induced burst suppression is related to synaptic inhibition while neurovascular coupling and mitochondrial function remain intact

Deep anaesthesia may impair neuronal, vascular and mitochondrial function facilitating neurological complications, such as delirium and stroke. On the other hand, deep anaesthesia is performed for neuroprotection in critical brain diseases such as status epilepticus or traumatic brain injury. Since the commonly used anaesthetic propofol causes mitochondrial dysfunction, we investigated the impact of the alternative anaesthetic isoflurane on neuro-metabolism. In deeply anaesthetised Wistar rats (burst suppression pattern), we measured increased cortical tissue oxygen pressure (ptiO2), a 35% drop in regional cerebral blood flow (rCBF) and burst-associated neurovascular responses. In vitro, 3% isoflurane blocked synaptic transmission and impaired network oscillations, thereby decreasing the cerebral metabolic rate of oxygen (CMRO2). Concerning mitochondrial function, isoflurane induced a reductive shift in flavin adenine dinucleotide (FAD) and decreased stimulus-induced FAD transients as Ca2þ influx was reduced by 50%. Computer simulations based on experimental results predicted no direct effects of isoflurane on mitochondrial complexes or ATP-synthesis. We found that isoflurane-induced burst suppression is related to decreased ATP consumption due to inhibition of synaptic activity while neurovascular coupling and mitochondrial function remain intact. The neurometabolic profile of isoflurane thus appears to be superior to that of propofol which has been shown to impair the mitochondrial respiratory chai