Galanin diminishes cortical spreading depolarization across rodents - A candidate for treatment?

Galanin (Gal) is a neuropeptide with the potential to ameliorate cortical spreading depolarization (CSD), an electrophysiological phenomenon occurring after brain injury or in migraine aura. Gal is expressed in all cortical neurons both in rat and in mouse cortices. Here we investigated whether the effect of Gal on CSD previously described in the rat is conserved in the mouse cortex. In rats, the topical application of Gal to the cortex for 1 h did not induce any change in CSD amplitudes, propagation velocity, or threshold of elicitation. Rather, topical application of Gal for 3 h was necessary to obtain a significant decrease in these CSD parameters and to develop a remarkable increase in the KCl threshold to elicit a CSD in rat cortex. In contrast, the topical application of Gal on cortical surface for 1 h in mice was sufficient to significantly attenuate CSD amplitudes and increase threshold. A thinner cortex, a faster diffusion or different affinity/expression of receptors for Gal are possible reasons to explain this difference in the time course between rats and mice. Our data are relevant to postulate Gal as a potential target for inhibition of CSD under pathological situations such as stroke or ischemia. SIGNIFICANCE STATEMENT: The neuropeptide Galanin (Gal) is expressed in all neurons throughout the cerebral cortex, both in rats and mice, and is able to reduce or even inhibit Cortical Spreading Depolarization, thus, Gal has the potential to control neuronal excitability that may identify Gal as a target in drug development against CSD.

Transient Receptor Potential Ankyrin 1-dependent Activation of Extracellular Signal-regulated Kinase 2 in the Cerebral Cortices Contributes to Cortical Spreading Depolarization.

Extracellular signal-regulated kinase (ERK) are serine/threonine-selective proteins and ERK1/2 can be phosphorylated in peripheral and central brain regions after cortical spreading depolarization (CSD) and calcitonin gene-related peptide; However, it remains unclear about whether and how ERK activity modulates CSD that correlates to migraine aura. Here, we determined the role of ERK in regulating CSD and explored the underlying mechanism involving transient receptor potential ankyrin 1 (TRPA1), a stress-sensing cation channel. CSD was recorded using intrinsic optical imaging in mouse brain slices, and electrophysiology in rats. Phosphorylated ERK (pERK1/2) and interleukin-1ß (IL-1ß) protein levels were detected using Western blot or enzyme-linked immunosorbent assay, respectively. IL-1ß mRNA level was detected using qPCR. The results showed that an ERK inhibitor, SCH77298, markedly prolonged CSD latency and reduced propagation rate in mouse brain slices. Corresponding to this, CSD induction increased levels of cytosolic pERK1/2 in ipsilateral cerebral cortices of rats, the elevation of which correlated to the level of IL-1ß mRNA. Mechanistic analysis showed that pre-treatment of an anti-TRPA1 antibody reduced the cytosolic pERK2 level but not pERK1 following CSD in cerebral cortices of rats and this level of pERK2 correlated with that of cerebral cortical IL-1ß protein. Furthermore, an ERK activator, AES16-2M, but not its scrambled control, reversed the prolonged CSD latency by a TRPA1 inhibitor, HC-030031, in mouse brain slices. These data revealed a crucial role of ERK activity in regulating CSD, and elevation of pERK and IL-1ß production induced by CSD is predominantly TRPA1 channel-dependent, thereby contributing to migraine pathogenesis.

Different vulnerability of fast and slow cortical oscillations to suppressive effect of spreading depolarization: state-dependent features potentially relevant to pathogenesis of migraine aura.

BACKGROUND: Spreading depolarization (SD), underlying mechanism of migraine aura and potential activator of pain pathways, is known to elicit transient local silencing cortical activity. Sweeping across the cortex, the electrocorticographic depression is supposed to underlie spreading negative symptoms of migraine aura. Main information about the suppressive effect of SD on cortical oscillations was obtained in anesthetized animals while ictal recordings in conscious patients failed to detect EEG depression during migraine aura. Here, we investigate the suppressive effect of SD on spontaneous cortical activity in awake animals and examine whether the anesthesia modifies the SD effect. METHODS: Spectral and spatiotemporal characteristics of spontaneous cortical activity following a single unilateral SD elicited by amygdala pinprick were analyzed in awake freely behaving rats and after induction of urethane anesthesia. RESULTS: In wakefulness, SD transiently suppressed cortical oscillations in all frequency bands except delta. Slow delta activity did not decline its power during SD and even increased it afterwards; high-frequency gamma oscillations showed the strongest and longest depression under awake conditions. Unexpectedly, gamma power reduced not only during SD invasion the recording cortical sites but also when SD occupied distant subcortical/cortical areas. Contralateral cortex not invaded by SD also showed transient depression of gamma activity in awake animals. Introduction of general anesthesia modified the pattern of SD-induced depression: SD evoked the strongest cessation of slow delta activity, milder suppression of fast oscillations and no distant changes in gamma activity. CONCLUSION: Slow and fast cortical oscillations differ in their vulnerability to SD influence, especially in wakefulness. In the conscious brain, SD produces stronger and spatially broader depression of fast cortical oscillations than slow ones. The frequency-specific effects of SD on cortical activity of awake brain may underlie some previously unexplained clinical features of migraine aura.

Spontaneous and optogenetically induced cortical spreading depolarization in familial hemiplegic migraine type 1 mutant mice.

Mechanisms underlying the migraine aura are incompletely understood, which to large extent is related to a lack of models in which cortical spreading depolarization (CSD), the correlate of the aura, occurs spontaneously. Here, we investigated electrophysiological and behavioural CSD features in freely behaving mice expressing mutant CaV2.1 Ca2+ channels, either with the milder R192Q or the severer S218L missense mutation in the α1 subunit, known to cause familial hemiplegic migraine type 1 (FHM1) in patients. Very rarely, spontaneous CSDs were observed in mutant but never in wildtype mice. In homozygous Cacna1aR192Q mice exclusively single-wave CSDs were observed whereas heterozygous Cacna1aS218L mice displayed multiple-wave events, seemingly in line with the more severe clinical phenotype associated with the S218L mutation. Spontaneous CSDs were associated with body stretching, one-directional slow head turning, and rotating movement of the body. Spontaneous CSD events were compared with those induced in a controlled manner using minimally invasive optogenetics. Also in the optogenetic experiments single-wave CSDs were observed in Cacna1aR192Q and Cacna1aS218L mice (whereas the latter also showed multiple-wave events) with movements similar to those observed with spontaneous events. Compared to wildtype mice, FHM1 mutant mice exhibited a reduced threshold and an increased propagation speed for optogenetically induced CSD with a more profound CSD-associated dysfunction, as indicated by a prolonged suppression of transcallosal evoked potentials and a reduction of unilateral forepaw grip performance. When induced during sleep, the optogenetic CSD threshold was particularly lowered, which may explain why spontaneous CSD events predominantly occurred during sleep. In conclusion, our data show that key neurophysiological and behavioural features of optogenetically induced CSDs mimic those of rare spontaneous events in FHM1 R192Q and S218L mutant mice with differences in severity in line with FHM1 clinical phenotypes seen with these mutations.

Increased presynaptic excitability in a migraine with aura mutation.

Migraine is a common and disabling neurological disorder. The headache and sensory amplifications of migraine are attributed to hyperexcitable sensory circuits, but a detailed understanding remains elusive. A mutation in casein kinase 1 delta (CK1δ) was identified in non-hemiplegic familial migraine with aura and advanced sleep phase syndrome. Mice carrying the CK1δT44A mutation were more susceptible to spreading depolarization (the phenomenon that underlies migraine aura), but mechanisms underlying this migraine-relevant phenotype were not known. We used a combination of whole-cell electrophysiology and multiphoton imaging, in vivo and in brain slices, to compare CK1δT44A mice (adult males) to their wild-type littermates. We found that despite comparable synaptic activity at rest, CK1δT44A neurons were more excitable upon repetitive stimulation than wild-type, with a reduction in presynaptic adaptation at excitatory but not inhibitory synapses. The mechanism of this adaptation deficit was a calcium-dependent enhancement of the size of the readily releasable pool of synaptic vesicles, and a resultant increase in glutamate release, in CK1δT44A compared to wild-type synapses. Consistent with this mechanism, CK1δT44A neurons showed an increase in the cumulative amplitude of excitatory post-synaptic currents, and a higher excitation-to-inhibition ratio during sustained activity compared to wild-type. At a local circuit level, action potential bursts elicited in CK1δT44A neurons triggered an increase in recurrent excitation compared to wild-type, and at a network level, CK1δT44A mice showed a longer duration of 'up state' activity, which is dependent on recurrent excitation. Finally, we demonstrated that the spreading depolarization susceptibility of CK1δT44A mice could be returned to wild-type levels with the same intervention (reduced extracellular calcium) that normalized presynaptic adaptation. Taken together, these findings show a stimulus-dependent presynaptic gain of function at glutamatergic synapses in a genetic model of migraine, that accounts for the increased spreading depolarization susceptibility and may also explain the sensory amplifications that are associated with the disease.

Isoflurane lowers the cerebral metabolic rate of oxygen and prevents hypoxia during cortical spreading depolarization in vitro: An integrative experimental and modeling study.

Cortical spreading depolarization (SD) imposes a massive increase in energy demand and therefore evolves as a target for treatment following acute brain injuries. Anesthetics are empirically used to reduce energy metabolism in critical brain conditions, yet their effect on metabolism during SD remains largely unknown. We investigated oxidative metabolism during SD in brain slices from Wistar rats. Extracellular potassium ([K+]o), local field potential and partial tissue oxygen pressure (ptiO2) were measured simultaneously. The cerebral metabolic rate of oxygen (CMRO2) was calculated using a reaction-diffusion model. By that, we tested the effect of clinically relevant concentrations of isoflurane on CMRO2 during SD and modeled tissue oxygenation for different capillary pO2 values. During SD, CMRO2 increased 2.7-fold, resulting in transient hypoxia in the slice core. Isoflurane decreased CMRO2, reduced peak [K+]o, and prolonged [K+]o clearance, which indicates reduced synaptic transmission and sodium-potassium ATPase inhibition. Modeling tissue oxygenation during SD illustrates the need for increased capillary pO2 levels to prevent hypoxia. In the absence thereof, isoflurane could improve tissue oxygenation by lowering CMRO2. Therefore, isoflurane is a promising candidate for pre-clinical studies on neuronal survival in conditions involving SD.

Commentary to Brain-wide continuous functional ultrasound imaging for real-time monitoring of hemodynamics during ischemic stroke.

Functional ultrasound (FUS) has emerged as a novel imaging method to reliably assess relative cerebral blood volume (rCBV) and infer perfusion, with good spatiotemporal resolution. Brunner and colleagues provide what appears to be its first application to characterize peri-infarct spreading depolarizations (SDs) in experimental stroke through recording of transient hyperemic events. They also report incomplete overlap between acute perfusion deficits and subsequent infarct distribution, specifically noting a rostral expansion to involve penumbral territory from which propagating depolarizations had preferentially originated. This observation would not be straightforward using other methodologies. Other strengths and limitations of the study are considered.

Traveling waves in a model for cortical spreading depolarization with slow-fast dynamics.

Cortical spreading depression and spreading depolarization (CSD) are waves of neuronal depolarization that spread across the cortex, leading to a temporary saturation of brain activity. They are associated with various brain disorders such as migraine and ischemia. We consider a reduced version of a biophysical model of a neuron-astrocyte network for the initiation and propagation of CSD waves [Huguet et al., Biophys. J. 111(2), 452-462, 2016], consisting of reaction-diffusion equations. The reduced model considers only the dynamics of the neuronal and astrocytic membrane potentials and the extracellular potassium concentration, capturing the instigation process implicated in such waves. We present a computational and mathematical framework based on the parameterization method and singular perturbation theory to provide semi-analytical results on the existence of a wave solution and to compute it jointly with its velocity of propagation. The traveling wave solution can be seen as a heteroclinic connection of an associated system of ordinary differential equations with a slow-fast dynamics. The presence of distinct time scales within the system introduces numerical instabilities, which we successfully address through the identification of significant invariant manifolds and the implementation of the parameterization method. Our results provide a methodology that allows to identify efficiently and accurately the mechanisms responsible for the initiation of these waves and the wave propagation velocity.

History of migraine.

Migraine symptoms were described in ancient Babylonia, and supernatural forces were felt to play a role in etiology and treatment. This changed in the Greco-Roman period, when the (dis)balance of humors was considered in (patho)physiology and treatment based on this. Aretaeus distinguished between cephalalgia, cephalea, and heterocrania. The latter term was changed to hemicrania by Galen. Physicians in the 17th century attributed headache to the meninges, extracranial periost, and cranial blood vessels. As for the pathophysiology, Willis suggested intracranial vasoconstriction with subsequent dilatation. Tissot and Fothergill gave comprehensive descriptions of migraine, including visual symptoms. Symptomatic and idiopathic hemicrania were distinguished in the early 19th century. Vasomotor pathophysiology was scientifically studied in the 1860s, leading to sympathicotonic and angioparalytic theories. Latham combined them, stating the latter follows the first. Ergot was introduced in 1868; ergotamine was isolated in 1918. This led to the vasodilatation theory of migraine (Wolff), the discovery of 5-HT, and later the specific agonists. Aura and cortical spreading depression were studied in the early 1940s and related to spreading oligemia in the 1980s. Subsequently, hyperemia followed by oligemia after CSD was found. After the discovery of CGRP, a new a class of drugs became the subject of clinical studies.

Diversity of cortical activity changes beyond depression during Spreading Depolarizations.

Spreading depolarizations (SDs) are classically thought to be associated with spreading depression of cortical activity. Here, we found that SDs in patients with subarachnoid hemorrhage produce variable, ranging from depression to booming, changes in electrocorticographic activity, especially in the delta frequency band. In rats, depression of activity was characteristic of high-potassium-induced full SDs, whereas partial superficial SDs caused either little change or a boom of activity at the cortical vertex, supported by volume conduction of signals from spared delta generators in the deep cortical layers. Partial SDs also caused moderate neuronal depolarization and sustained excitation, organized in gamma oscillations in a narrow sub-SD zone. Thus, our study challenges the concept of homology between spreading depolarization and spreading depression by showing that SDs produce variable, from depression to booming, changes in activity at the cortical surface and in different cortical layers depending on the depth of SD penetration.

Spreading Depolarizations Suppress Hematoma Growth in Hyperacute Intracerebral Hemorrhage in Mice.

BACKGROUND: Spreading depolarizations (SDs) occur in all types of brain injury and may be associated with detrimental effects in ischemic stroke and subarachnoid hemorrhage. While rapid hematoma growth during intracerebral hemorrhage triggers SDs, their role in intracerebral hemorrhage is unknown. METHODS: We used intrinsic optical signal and laser speckle imaging, combined with electrocorticography, to investigate the effects of SD on hematoma growth during the hyperacute phase (0-4 hours) after intracortical collagenase injection in mice. Hematoma expansion, SDs, and cerebral blood flow were simultaneously monitored under normotensive and hypertensive conditions. RESULTS: Spontaneous SDs erupted from the vicinity of the hematoma during rapid hematoma growth. We found that hematoma growth slowed down by >60% immediately after an SD. This effect was even stronger in hypertensive animals with faster hematoma growth. To establish causation, we exogenously induced SDs (every 30 minutes) at a remote site by topical potassium chloride application and found reduced hematoma growth rate and final hemorrhage volume (18.2±5.8 versus 10.7±4.1 mm3). Analysis of cerebral blood flow using laser speckle flowmetry revealed that suppression of hematoma growth by spontaneous or induced SDs coincided and correlated with the characteristic oligemia in the wake of SD, implicating the vasoconstrictive effect of SD as one potential mechanism of action. CONCLUSIONS: Our findings reveal that SDs limit hematoma growth during the early hours of intracerebral hemorrhage and decrease final hematoma volume.

Mechanisms of initiation of cortical spreading depression.

BACKGROUND: There is increasing evidence from human and animal studies that cortical spreading depression (CSD) is the neurophysiological correlate of migraine aura and a trigger of migraine pain mechanisms. The mechanisms of initiation of CSD in the brain of migraineurs remain unknown, and the mechanisms of initiation of experimentally induced CSD in normally metabolizing brain tissue remain incompletely understood and controversial. Here, we investigated the mechanisms of CSD initiation by focal application of KCl in mouse cerebral cortex slices. METHODS: High KCl puffs of increasing duration up to the threshold duration eliciting a CSD were applied on layer 2/3 whilst the membrane potential of a pyramidal neuron located very close to the site of KCl application and the intrinsic optic signal were simultaneously recorded. This was done before and after the application of a specific blocker of either NMDA or AMPA glutamate receptors (NMDARs, AMPARs) or voltage-gated Ca2+ (CaV) channels. If the drug blocked CSD, stimuli up to 12-15 times the threshold were applied. RESULTS: Blocking either NMDARs with MK-801 or CaV channels with Ni2+ completely inhibited CSD initiation by both CSD threshold and largely suprathreshold KCl stimuli. Inhibiting AMPARs with NBQX was without effect on the CSD threshold and velocity. Analysis of the CSD subthreshold and threshold neuronal depolarizations in control conditions and in the presence of MK-801 or Ni2+ revealed that the mechanism underlying ignition of CSD by a threshold stimulus (and not by a just subthreshold stimulus) is the CaV-dependent activation of a threshold level of NMDARs (and/or of channels whose opening depends on the latter). The delay of several seconds with which this occurs underlies the delay of CSD initiation relative to the rapid neuronal depolarization produced by KCl. CONCLUSIONS: Both NMDARs and CaV channels are necessary for CSD initiation, which is not determined by the extracellular K+ or neuronal depolarization levels per se, but requires the CaV-dependent activation of a threshold level of NMDARs. This occurs with a delay of several seconds relative to the rapid depolarization produced by the KCl stimulus. Our data give insights into potential mechanisms of CSD initiation in migraine.

Cortical spreading depolarization-induced constriction of penetrating arteries can cause watershed ischemia: A potential mechanism for white matter lesions.

Periventricular white matter lesions (WMLs) are common MRI findings in migraine with aura (MA). Although hemodynamic disadvantages of vascular supply to this region create vulnerability, the pathophysiological mechanisms causing WMLs are unclear. We hypothesize that prolonged oligemia, a consequence of cortical spreading depolarization (CSD) underlying migraine aura, may lead to ischemia/hypoxia at hemodynamically vulnerable watershed zones fed by long penetrating arteries (PAs). For this, we subjected mice to KCl-triggered single or multiple CSDs. We found that post-CSD oligemia was significantly deeper at medial compared to lateral cortical areas, which induced ischemic/hypoxic changes at watershed areas between the MCA/ACA, PCA/anterior choroidal and at the tip of superficial and deep PAs, as detected by histological and MRI examination of brains 2-4 weeks after CSD. BALB-C mice, in which MCA occlusion causes large infarcts due to deficient collaterals, exhibited more profound CSD-induced oligemia and were more vulnerable compared to Swiss mice such that a single CSD was sufficient to induce ischemic lesions at the tip of PAs. In conclusion, CSD-induced prolonged oligemia has potential to cause ischemic/hypoxic injury at hemodynamically vulnerable brain areas, which may be one of the mechanisms underlying WMLs located at the tip of medullary arteries seen in MA patients.

Aura phenomena do not initiate migraine attacks-Findings from neuroimaging.

OBJECTIVES/BACKGROUND: As cortical spreading depolarization (CSD) has been suggested to be the cause of migraine aura and as CSD can activate trigeminal nociceptive neurons in animals, it has been suggested that CSD may be the cause of migraine attacks. This raises the question of how migraine pain is generated in migraine attacks without aura and has led to the hypothesis that CSD may also occur in subcortical regions in the form of "silent" CSDs, and accordingly "silent auras". METHODS: In this case study, we provide evidence for common neuronal alterations preceding headache attacks with and without aura in a male patient with migraine, who underwent daily event-correlated functional magnetic resonance imaging of trigeminal nociception for a period of 30 days. During these days the man experienced migraine attacks with and without aura. RESULTS: Comparing the preictal phases between both attack types revealed a common hyperactivation of the hypothalamus (p < 0.01), which was already present 2 days before the actual attack. CONCLUSION: The time frame of the central pathophysiological orchestration of migraine attacks, irrelevant of the presence of later aura, strongly suggests that the aura is an epiphenomenon that is unrelated and does not initiate headache attacks.

A model of neurovascular coupling and its application to cortical spreading depolarization.

Cortical spreading depolarization (CSD) is a neuropathological condition involving propagating waves of neuronal silence, and is related to multiple diseases, such as migraine aura, traumatic brain injury (TBI), stroke, and cardiac arrest, as well as poor outcome of patients. While CSDs of different severity share similar roots on the ion exchange level, they can lead to different vascular responses (namely spreading hyperemia and spreading ischemia). In this paper, we propose a mathematical model relating neuronal activities to predict vascular changes as measured with near-infrared spectroscopy (NIRS) and fMRI recordings, and apply it to the extreme case of CSD, where sustained near-complete neuronal depolarization is seen. We utilize three serially connected models (namely, ion exchange, neurovascular coupling, and hemodynamic model) which are described by differential equations. Propagating waves of ion concentrations, as well as the associated vasodynamics and hemodynamics, are simulated by solving these equations. Our proposed model predicts vasodynamics and hemodynamics that agree both qualitatively and quantitatively with experimental literature. Mathematical modeling and simulation offer a powerful tool to help understand the underlying mechanisms of CSD and help interpret the data. In addition, it helps develop novel monitoring techniques prior to data collection. Our simulated results strongly suggest that fMRI is unable to reliably distinguish between spreading hyperemia and spreading ischemia, while NIRS signals are substantially distinct in the two cases.

Optogenetic cortical spreading depolarization induces headache-related behaviour and neuroinflammatory responses some prolonged in familial hemiplegic migraine type 1 mice.

BACKGROUND: Cortical spreading depolarization (CSD), the neurophysiological correlate of the migraine aura, can activate trigeminal pain pathways, but the neurobiological mechanisms and behavioural consequences remain unclear. Here we investigated effects of optogenetically-induced CSDs on headache-related behaviour and neuroinflammatory responses in transgenic mice carrying a familial hemiplegic migraine type 1 (FHM1) mutation. METHODS: CSD events (3 in total) were evoked in a minimally invasive manner by optogenetic stimulation through the intact skull in freely behaving wildtype (WT) and FHM1 mutant mice. Related behaviours were analysed using mouse grimace scale (MGS) scoring, head grooming, and nest building behaviour. Neuroinflammatory changes were investigated by assessing HMGB1 release with immunohistochemistry and by pre-treating mice with a selective Pannexin-1 channel inhibitor. RESULTS: In both WT and FHM1 mutant mice, CSDs induced headache-related behaviour, as evidenced by increased MGS scores and the occurrence of oculotemporal strokes, at 30 min. Mice of both genotypes also showed decreased nest building behaviour after CSD. Whereas in WT mice MGS scores had normalized at 24 h after CSD, in FHM1 mutant mice scores were normalized only at 48 h. Of note, oculotemporal stroke behaviour already normalized 5 h after CSD, whereas nest building behaviour remained impaired at 72 h; no genotype differences were observed for either readout. Nuclear HMGB1 release in the cortex of FHM1 mutant mice, at 30 min after CSD, was increased bilaterally in both WT and FHM1 mutant mice, albeit that contralateral release was more pronounced in the mutant mice. Only in FHM1 mutant mice, contralateral release remained higher at 24 h after CSD, but at 48 h had returned to abnormal, elevated, baseline values, when compared to WT mice. Blocking Panx1 channels by TAT-Panx308 inhibited CSD-induced headache related behaviour and HMGB1 release. CONCLUSIONS: CSDs, induced in a minimally invasive manner by optogenetics, investigated in freely behaving mice, cause various migraine relevant behavioural and neuroinflammatory phenotypes that are more pronounced and longer-lasting in FHM1 mutant compared to WT mice. Prevention of CSD-related neuroinflammatory changes may have therapeutic potential in the treatment of migraine.

Nonpharmacological modulation of cortical spreading depolarization.

AIMS: Cortical spreading depolarization (CSD) is a wave of pathologic neuronal dysfunction that spreads through cerebral gray matter, causing neurologic disturbance in migraine and promoting lesion development in acute brain injury. Pharmacologic interventions have been found to be effective in migraine with aura, but their efficacy in acutely injured brains may be limited. This necessitates the assessment of possible adjunctive treatments, such as nonpharmacologic methods. This review aims to summarize currently available nonpharmacological techniques for modulating CSDs, present their mechanisms of action, and provide insight and future directions for CSD treatment. MAIN METHODS: A systematic literature review was performed, generating 22 articles across 3 decades. Relevant data is broken down according to method of treatment. KEY FINDINGS: Both pharmacologic and nonpharmacologic interventions can mitigate the pathological impact of CSDs via shared molecular mechanisms, including modulating K+/Ca2+/Na+/Cl- ion channels and NMDA, GABAA, serotonin, and CGRP ligand-based receptors and decreasing microglial activation. Preclinical evidence suggests that nonpharmacologic interventions, including neuromodulation, physical exercise, therapeutic hypothermia, and lifestyle changes can also target unique mechanisms, such as increasing adrenergic tone and myelination and modulating membrane fluidity, which may lend broader modulatory effects. Collectively, these mechanisms increase the electrical initiation threshold, increase CSD latency, slow CSD velocity, and decrease CSD amplitude and duration. SIGNIFICANCE: Given the harmful consequences of CSDs, limitations of current pharmacological interventions to inhibit CSDs in acutely injured brains, and translational potentials of nonpharmacologic interventions to modulate CSDs, further assessment of nonpharmacologic modalities and their mechanisms to mitigate CSD-related neurologic dysfunction is warranted.

Estrogen modulation of cortical spreading depression.

BACKGROUND AND AIMS: Cortical spreading depression (CSD), a transient neuronal and glial depolarization that propagates slowly across the cerebral cortex, is the putative electrophysiological event underlying migraine aura and a headache trigger. Migraine is three times more prevalent in women than men, linked to circulating female hormones. High estrogen levels or estrogen withdrawal may be a migraine trigger for many women. We, therefore, aimed to examine whether sex, gonadectomy, and female hormone supplementation and withdrawal affect the susceptibility to CSD. METHODS: To determine CSD susceptibility, we recorded the frequency of CSDs triggered during 2-h topical KCl application in intact or gonadectomized female and male rats, without or with estradiol or progesterone supplementation via daily intraperitoneal injections. Estrogen or progesterone treatment followed by withdrawal was studied in a separate cohort. To take the first step towards identifying potential mechanisms, we studied glutamate and GABAA receptor binding using autoradiography. RESULTS: The CSD frequency in intact female rats was higher than intact male and ovariectomized rats. We did not detect a change in CSD frequency during different stages of the estrous cycle in intact females. Daily estrogen injections for three weeks did not change CSD frequency. However, one-week estrogen withdrawal after two weeks of treatment significantly increased CSD frequency compared with the vehicle group in gonadectomized females. The same protocol of estrogen treatment and withdrawal was ineffective in gonadectomized males. In contrast to estrogen, daily progesterone injections for three weeks elevated CSD susceptibility, and one-week withdrawal after two weeks of treatment partially normalized this effect. Autoradiography did not reveal significant changes in glutamate or GABAA receptor binding density after estrogen treatment and withdrawal. CONCLUSIONS: These data suggest that females are more susceptible to CSD, and sexual dimorphism is abrogated by gonadectomy. Moreover, estrogen withdrawal after prolonged daily treatment enhances CSD susceptibility. These findings may have implications for estrogen-withdrawal migraine, although the latter tends to be without aura.

Improving Neurotrauma by Depolarization Inhibition With Combination Therapy: A Phase 2 Randomized Feasibility Trial.

BACKGROUND AND OBJECTIVES: Spreading depolarizations (SDs) are a pathological mechanism that mediates lesion development in cerebral gray matter. They occur in ∼60% of patients with severe traumatic brain injury (TBI), often in recurring and progressive patterns from days 0 to 10 after injury, and are associated with worse outcomes. However, there are no protocols or trials suggesting how SD monitoring might be incorporated into clinical management. The objective of this protocol is to determine the feasibility and efficacy of implementing a treatment protocol for intensive care of patients with severe TBI that is guided by electrocorticographic monitoring of SDs. METHODS: Patients who undergo surgery for severe TBI with placement of a subdural electrode strip will be eligible for enrollment. Those who exhibit SDs on electrocorticography during intensive care will be randomized 1:1 to either (1) standard care that is blinded to the further course of SDs or (2) a tiered intervention protocol based on efficacy to suppress further SDs. Interventions aim to block the triggering and propagation of SDs and include adjusted targets for management of blood pressure, CO 2 , temperature, and glucose, as well as ketamine pharmacotherapy up to 4 mg/kg/ hour. Interventions will be escalated and de-escalated depending on the course of SD pathology. EXPECTED OUTCOMES: We expect to demonstrate that electrocorticographic monitoring of SDs can be used as a real- time diagnostic in intensive care that leads to meaningful changes in patient management and a reduction in secondary injury, as compared with standard care, without increasing medical complications or adverse events. DISCUSSION: This trial holds potential for personalization of intensive care management by tailoring therapies based on monitoring and confirmation of the targeted neuronal mechanism of SD. Results are expected to validate the concept of this approach, inform refinement of the treatment protocol, and lead to larger-scale trials.

Optogenetic Spreading Depolarizations Do Not Worsen Acute Ischemic Stroke Outcome.

BACKGROUND: Spreading depolarizations (SDs) are believed to contribute to injury progression and worsen outcomes in focal cerebral ischemia because exogenously induced SDs have been associated with enlarged infarct volumes. However, previous studies used highly invasive methods to trigger SDs that can directly cause tissue injury (eg, topical KCl) and confound the interpretation. Here, we tested whether SDs indeed enlarge infarcts when induced via a novel, noninjurious method using optogenetics. METHODS: Using transgenic mice expressing channelrhodopsin-2 in neurons (Thy1-ChR2-YFP), we induced 8 optogenetic SDs to trigger SDs noninvasively at a remote cortical location in a noninjurious manner during 1-hour distal microvascular clip or proximal an endovascular filament occlusion of the middle cerebral artery. Laser speckle imaging was used to monitor cerebral blood flow. Infarct volumes were then quantified at 24 or 48 hours. RESULTS: Infarct volumes in the optogenetic SD arm did not differ from the control arm in either distal or proximal middle cerebral artery occlusion, despite a 6-fold and 4-fold higher number of SDs, respectively. Identical optogenetic illumination in wild-type mice did not affect the infarct volume. Full-field laser speckle imaging showed that optogenetic stimulation did not affect the perfusion in the peri-infarct cortex. CONCLUSIONS: Altogether, these data show that SDs induced noninvasively using optogenetics do not worsen tissue outcomes. Our findings compel a careful reexamination of the notion that SDs are causally linked to infarct expansion.