Focal Ischemia Research Articles

Cerebral ischemia-induced gene expression changes in diabetic mice from acute to subacute stage

Diabetes is a risk factor for stroke; however, its impact on stroke progression at the genomic level is not well understood. To address this gap, we used transcriptome sequencing to explore the relationship between lncRNA and mRNA expression patterns and the reperfusion duration in the cortex of diabetically induced stroke mice. First, focal ischemia was induced in adult male ob/ob mice, which were then subjected to reperfusion periods of one, three, or seven days. Total RNA was extracted from the ischemic cortical tissue for RNA sequencing, and the resulting reads were aligned to the GRCm38 murine reference genome. A total of 672 mRNAs and 301 lncRNAs were identified as differentially expressed one day post-stroke, 1195 mRNAs and 66 lncRNAs at three days post-stroke, and 1069 mRNAs and 75 lncRNAs at seven days post-stroke. Stage-specific differentially expressed mRNAs were bioinformatically analyzed and found significantly enriched in processes such as apoptosis, angiogenesis, and lipid metabolism at one, three, and seven days post-stroke, respectively. Stage-specific DElncRNA-mRNA cis-regulatory relationships were constructed using these biological processes as examples, revealing the potential roles of four pairs of lncRNA-mRNAs (Gm39787-Lcn2, Gm46111-Drd2, D3300500i16Rik-Fosl1, and Gm41689-Egr1) in apoptosis. Additionally, Gm40237-Tie1 and Gm52352-Pdgfrb are associated with angiogenesis and lipid metabolism, respectively. In conclusion, our study demonstrated that lncRNA and mRNA expression in the cortex of transient focal ischemia-induced diabetic mice undergo extensive alterations, providing insights into complex molecular interactions underlying diabetic stroke.

Dynamics of functional impairments during focal transient ischemia in three-dimensional cortical space

Ischemic injury in the cerebral cortex results in decreased electrical activity across all frequency bands and the emergence of abnormal electrophysiological patterns, including spreading depolarization (SD) and negative ultraslow potential (NUP) [1, 2]. Despite this, the specific dynamics of these changes in electrical activity within the three-dimensional cortical space during ischemia remain incompletely described and poorly understood. To simultaneously investigate changes in electrical activity across the layers of the cerebral cortex and in the horizontal cortical space, we used two linear 16-channel silicone probes (Neuronexus, USA) in combination with a flexible transparent 60-channel matrix of subdural electrodes (MIPT, Russia) and intrinsic optical signal imaging (IOS, 665 nm, transillumination mode) during focal ischemia induced by intracortical injection of the potent vasoconstrictor endothelin-1 (ET-1, 1 μL, 1 μM). These experiments were carried out in head-restrained rats under urethane anesthesia (1.5 g/kg). Formation of the ischemic lesion following ET-1 administration was correlated with clusters of SDs that demonstrated considerable variability in their propagation patterns within both vertical and horizontal planes. The initial SDs originated from the injection site of ET-1 and diffused across all cortical layers. Subsequently, the initiation point of the ensuing SDs gradually shifted towards the deeper layers while the electrical activity showed inadequate recovery between SDs within the injection site. SDs originating in the surrounding cortex did not invade the area near the injection point. Instead, they tended to spread around, often compartmentalizing in the superficial layers of the cortex. Some SDs were observed deep in the cortex, while others were detected on the surface via superficial electrodes and IOS, without leaving typical intracortical electrode traces. Electrographic activity was significantly depressed, especially in the superficial layers around the injection site, three hours after ET-1 administration. However, it returned to pre-ET-1 levels at the remote site, with a spatial gradient observed in subdural electrodes. Functional impairments corresponded to the histological lesion observed in coronal brain sections. Recently discovered NUPs were initiated by SD and were most prominent in the electrodes closest to the ET-1 injection site. These NUPs reached their maximal amplitude at one hour and subsided three hours after ET-1 injection. Our research indicates intricate dynamics in the creation of an ischemic focal point. The data gathered suggest that the emergence of cerebral harm during focal ischemia is associated with the growth of a focus extending both horizontally and vertically across cortical dimensions. This growth is fueled by the generation of SDs within the ischemic penumbra.

Electrical resistance of brain tissue during terminal ischemia

Terminal ischemia is characterized by several electrophysiological processes, including hyperpolarization, spreading depolarization (tSD), and negative ultraslow potentials (NUP) [1]. tSD is knowin to induce depolarization of neuron membranes to roughly 0 mV, accompanied by the influx of positive ions into cells, which results in the displacement of extracellular fluids into neurons, leading to cell swelling. In addition to the movement of extracellular fluid, cerebrospinal fluid flows into the perivascular region of the penetrating vessels. These processes result in a reduction of extracellular space and brain edema [2], which can elevate the likelihood of patient mortality up to 80% [3]. The decrease in extracellular space may lead to an upsurge in the electrical resistance of the tissue, enabling it to function as an important indicator of the anatomical and functional state of the brain during traumatic injuries and hemorrhages [4]. Our goal was to assess the resistance of the extracellular space in the rat’s barrel cortex by measuring voltage step amplitudes caused by current injection between the V1 cortex and the tail vein of an animal. We used 16-channel probes with iridium electrode sites and glass microelectrodes containing Ag/AgCl conductors filled with NaCl solution. We used a metal Ag/AgCl electrode as a reference, implanted in the cerebellum. The experimental animals were euthanized via inhalation of isoflurane at a lethal concentration. Isoflurane-induced respiratory arrest led to the subsequent development of a sequence of electrophysiological processes that included hyperpolarization, terminal spreading depression, and negative ultraslow potential. The NUP persisted throughout the entirety of the recording period (30–90 minutes), with a significant increase in extracellular space resistance occurring during this time. The rise in resistance began concurrently with the end of respiration. The resistance increased by 40 [23–57]% (median [25th–75th percentile], p=0.002, n=11) after 30 minutes of breath cessation, and by 46 [(–15)–64]% (p=0.109, n=8) after 60 minutes. The resistance demonstrated equivalent changes in all cerebral cortex depths. Additionally, the NUP amplitude following respiratory arrest had a correlation with the increase of voltage step amplitudes at 30 minutes (R=–0.713, p=0.014). The resistances measured using signals from both Ag/AgCl- and Ir electrodes did not vary in any of the experiments conducted (p=1, n=5). Thus, the rise in electrical resistance in brain tissue was shown to initiate at the point of respiratory arrest in the animal and progresses alongside the terminal processes in the cerebral cortex. This correlation is specifically linked to the fluctuations in the negative ultraslow potential. In tDS and NUP, a significant increase in tissue edema occurs due to the movement of water from the extracellular spaces into the cells, which leads to a decrease in extracellular space volume and an increase in resistance. The process can last for several minutes and is characterized by a continuous increase in voltage step amplitudes. These results suggest that tissue resistance increases during focal ischemia, as SD and NUP are also present during the formation of the ischemic focus [5].

Sex-Specific Acute Cerebrovascular Responses to Photothrombotic Stroke in Mice.

Mechanisms underlying cerebrovascular stroke outcomes are poorly understood, and the effects of biological sex on cerebrovascular regulation post-stroke have yet to be fully comprehended. Here, we explore the overlapping roles of gonadal sex hormones and rho-kinase (ROCK), two important modulators of cerebrovascular tone, on the acute cerebrovascular response to photothrombotic (PT) focal ischemia in mice. Male mice were gonadectomized and female mice were ovariectomized to remove gonadal hormones, whereas control ("intact") animals received a sham surgery prior to stroke induction. Intact wild-type (WT) males showed a delayed drop in cerebral blood flow (CBF) compared with intact WT females, whereby maximal CBF drop was observed 48 h following stroke. Gonadectomy in males did not alter this response. However, ovariectomy in WT females produced a "male-like" phenotype. Intact Rock2+/- males also showed the same phenotypic response, which was not altered by gonadectomy. Alternatively, intact Rock2+/- females showed a significant difference in CBF values compared with intact WT females, displaying higher CBF values immediately post-stroke and showing a maximal CBF drop 48 h post-stroke. This pattern was not altered by ovariectomy. Altogether, these data illustrate sex differences in acute CBF responses to PT stroke, which seem to involve gonadal female sex hormones and ROCK2. Overall, this study provides a framework for exploring sex differences in acute CBF responses to focal ischemic stroke in mice.

In situ direct reprogramming of astrocytes to neurons via polypyrimidine tract-binding protein 1 knockdown in a mouse model of ischemic stroke.

JOURNAL/nrgr/04.03/01300535-202410000-00025/figure1/v/2024-02-06T055622Z/r/image-tiff In situ direct reprogramming technology can directly convert endogenous glial cells into functional neurons in vivo for central nervous system repair. Polypyrimidine tract-binding protein 1 (PTB) knockdown has been shown to reprogram astrocytes to functional neurons in situ. In this study, we used AAV-PHP.eB-GFAP-shPTB to knockdown PTB in a mouse model of ischemic stroke induced by endothelin-1, and investigated the effects of GFAP-shPTB-mediated direct reprogramming to neurons. Our results showed that in the mouse model of ischemic stroke, PTB knockdown effectively reprogrammed GFAP-positive cells to neurons in ischemic foci, restored neural tissue structure, reduced inflammatory response, and improved behavioral function. These findings validate the effectiveness of in situ transdifferentiation of astrocytes, and suggest that the approach may be a promising strategy for stroke treatment.

High frequency of endoluminal thrombus in patients with ischaemic stroke following AARS-CoV-2 infection

BackgroundIschaemic stroke may be a major complication of SARS-CoV-2 infection. Studying and characterising the different aetiological subtypes, clinical characteristics, and functional outcomes may be valuable in guiding patient selection for optimal management and treatment. MethodsData were collected retrospectively on consecutive patients with COVID-19 who developed acute focal brain ischaemia (between 1 March and 19 April 2020) at a tertiary university hospital in Madrid (Spain). ResultsDuring the study period, 1594 patients were diagnosed with COVID-19. We found 22 patients with ischaemic stroke (1.38%), 6 of whom did not meet the inclusion criteria. The remaining 16 patients were included in the study (15 cases of ischaemic stroke and one case of transient ischaemic attack).Median baseline National Institutes of Health Stroke Scale score was 9 (interquartile range: 16), and mean (standard deviation) age was 73 years (12.8). Twelve patients (75%) were men. Mean time from COVID-19 symptom onset to stroke onset was 13 days. Large vessel occlusion was identified in 12 patients (75%).We detected elevated levels of D-dimer in 87.5% of patients and C-reactive protein in 81.2%. The main aetiology was atherothrombotic stroke (9 patients, 56.3%), with the predominant subtype being endoluminal thrombus (5 patients, 31.2%), involving the internal carotid artery in 4 cases and the aortic arch in one. The mortality rate in our series was 44% (7 of 16 patients). ConclusionsIn patients with COVID-19, the most frequent stroke aetiology was atherothrombosis, with a high proportion of endoluminal thrombus (31.2% of patients). Our clinical and laboratory data support COVID-19–associated coagulopathy as a relevant pathophysiological mechanism for ischaemic stroke in these patients.

An exploratory pilot study on diffusion MRI measurement of cortical microstructural neurodegeneration for the assessment of dementia in a Japanese hospital setting

AbstractBackgroundThe use of diffusion MRI (dMRI) to assess cortical microstructure has demonstrated promise for measuring neurodegeneration in Alzheimer’s disease (PMID:33174658, PMID:36281682) and frontotemporal dementia (PMID:32641807, PMID:34686217) using retrospective research datasets. The ultimate translation of research findings to clinical practice requires prospective studies in hospital settings. A pilot study at the National Hospital Organization, Niigata, was designed and successfully delivered.MethodThe study was registered on Oct 5th 2021 (jRCT1032210367). Scanning took place between Oct 13th 2021 and June 30th 2022, on a 1.5T Philips Ingenia. MRI data was transferred using a secure cloud gateway (Cimar). A total of 10 healthy controls (HC), 10 Alzheimer’s disease (AD) patients, and 5 frontotemporal dementia (FTD) patients were recruited.Structural (3D T1‐weighted) and dMRI (2mm isotropic, 32 directions at b = 1000 s/mm2) scans were used to calculate cortical mean diffusivity and three neuropathologically‐inspired cortical diffusivity measures: the angle between the radial minicolumnar direction and the principal diffusion direction (AngleR); the principal diffusion component parallel with the minicolumns (ParlPD), and the diffusion components perpendicular to the minicolumns (PerpPD+). Metrics were also summarized over AD signature regions (Ridgway et al., CTAD 2022).ResultOf the n = 25 cases, 18 had clear clinical diagnoses after neurological and radiological assessment, while 7 were more complex (including focal bleeding, focal ischemia, meningioma, supra‐sellar tumour). Characteristics are given in Table 1.Figure 1 plots two whole‐brain measures: PerpPD+ and AngleR. A threshold of PerpPD+ > 1.325 has 100% sensitivity to the 8 clear AD cases, with 86% specificity for 7 controls vs AD. Results were less clear for FTD using whole‐brain measures alone.The Cohen’s d effect size for clear AD versus clear HC was 2.82 using whole‐brain PerpPD+. Using signature regions (Figure 2b) improved separation to d = 3.08 for PerpPD+, compared to d = 1.87 for the cortical thickness signature (Figure 2a).ConclusionThe pilot study demonstrated practical feasibility and good performance for distinguishing AD from HC using cortical diffusivity measures acquired at the 1.5T field‐strength common in hospitals in Japan. Future work will investigate FTD signature regions. A larger multicentre study assessing diagnostic accuracy using additional AD biomarkers is warranted.

An exploratory pilot study on diffusion MRI measurement of cortical microstructural neurodegeneration for the assessment of dementia in a Japanese hospital setting

AbstractBackgroundThe use of diffusion MRI (dMRI) to assess cortical microstructure has demonstrated promise for measuring neurodegeneration in Alzheimer’s disease (PMID:33174658, PMID:36281682) and frontotemporal dementia (PMID:32641807, PMID:34686217) using retrospective research datasets. The ultimate translation of research findings to clinical practice requires prospective studies in hospital settings. A pilot study at the National Hospital Organization, Niigata, was designed and successfully delivered.MethodThe study was registered on Oct 5th 2021 (jRCT1032210367). Scanning took place between Oct 13th 2021 and June 30th 2022, on a 1.5T Philips Ingenia. MRI data was transferred using a secure cloud gateway (Cimar). A total of 10 healthy controls (HC), 10 Alzheimer’s disease (AD) patients, and 5 frontotemporal dementia (FTD) patients were recruited. Structural (3D T1‐weighted) and dMRI (2mm isotropic, 32 directions at b = 1000 s/mm2) scans were used to calculate cortical mean diffusivity and three neuropathologically‐inspired cortical diffusivity measures: the angle between the radial minicolumnar direction and the principal diffusion direction (AngleR); the principal diffusion component parallel with the minicolumns (ParlPD), and the diffusion components perpendicular to the minicolumns (PerpPD+). Metrics were also summarized over AD signature regions (Ridgway et al., CTAD 2022).ResultOf the n = 25 cases, 18 had clear clinical diagnoses after neurological and radiological assessment, while 7 were more complex (including focal bleeding, focal ischemia, meningioma, supra‐sellar tumour). Characteristics are given in Table 1. Figure 1 plots two whole‐brain measures: PerpPD+ and AngleR. A threshold of PerpPD+ > 1.325 has 100% sensitivity to the 8 clear AD cases, with 86% specificity for 7 controls vs AD. Results were less clear for FTD using whole‐brain measures alone. The Cohen’s d effect size for clear AD versus clear HC was 2.82 using whole‐brain PerpPD+. Using signature regions (Figure 2b) improved separation to d = 3.08 for PerpPD+, compared to d = 1.87 for the cortical thickness signature (Figure 2a).ConclusionThe pilot study demonstrated practical feasibility and good performance for distinguishing AD from HC using cortical diffusivity measures acquired at the 1.5T field‐strength common in hospitals in Japan. Future work will investigate FTD signature regions. A larger multicentre study assessing diagnostic accuracy using additional AD biomarkers is warranted.

The Contribution of Hyperhomocysteinemia to Aβ‐Induced Endothelial Cell Apoptosis, Blood Brain Barrier Dysfunction, and Angiogenic Deficits

AbstractBackgroundCerebrovascular dysfunction has been implicated as a major contributor to Alzheimer’s Disease (AD) pathology, with particularly endothelial cell (EC) stress promoting the focal ischemia, cerebral blood flow impairments, and blood brain barrier (BBB) permeability that are pathologically characteristic in AD. Recent evidence has emerged suggesting a link between cardiovascular (CV) diseases and AD pathology, particularly showing that CV/cerebrovascular risk factors, including hyperhomocysteinemia (Hhcy), contribute to AD pathology and risk. Despite this, the underlying molecular mechanisms for this interaction remain unclear. Previously, our lab has demonstrated that amyloid beta (Aβ), particularly Aβ40‐Q22 (the vasculotropic Dutch mutant), promotes TRAIL death receptor (DR)‐mediated apoptosis, barrier permeability, and angiogenic impairment within human cerebral ECs. We tested the hypothesis that Hhcy exacerbates Aβ‐induced cerebral EC TRAIL DR‐mediated apoptosis, barrier dysfunction, and angiogenesis defects.MethodHuman cerebral microvascular ECs (HCMECs) were challenged with AβQ22 and/or homocysteine (Hcy). Apoptotic mediator expression, caspase activation, cytochrome c release, and DNA fragmentation were measured to assess apoptosis. Junction protein expression, trans‐endothelial‐electrical‐resistance (TEER), and inflammatory mediators were measured to assess EC barrier integrity. Angiogenesis inhibition and activation (cytokine expression, pVEGFR2Y1175 expression/localization, VEGF‐A ELISA) assays were utilized to measure EC angiogenic capability and wound healing and actin polymerization assays were utilized to measure EC wound healing ability.ResultCombined challenge of ECs with AβQ22 and Hcy potentiated increases in TRAIL DR‐related apoptotic mediators' expression, caspase activity, cytochrome c release, and DNA fragmentation and potentiated decreases in anti‐apoptotic mediators’ expression at differential timepoints. Treatment of ECs with AβQ22 and Hcy additively decreased TEER and, at certain timepoints, potentiated decreased junction protein expression, increased inflammatory mediator expression, and phosphorylated BBB protein expression. Combined treatment of ECs with AβQ22 and Hcy additively decreased angiogenesis progression, angiogenesis‐stimulating cytokine expression, wound healing capability, and actin polymerization levels. Additionally, Hcy decreased HCMEC soluble VEGF‐A levels.ConclusionThe presence of high Hcy exacerbates Aβ induced‐EC apoptosis, barrier dysfunction, and angiogenic impairment. This study revealed several of the specific molecular mechanism through which amyloidosis and CV risk factors may additively act to produce EC dysfunction and death in AD.

Positive Neuroprotective Effect of Argon Inhalation after Photochemically Induced Ischemic Stroke Model in Rats.

We studied the effect of 2-h inhalation of argon-oxygen mixture (Ar 70%/O2 30%) after photochemically induced stroke and on days 2 and 3 after stroke modeling on the severity of neurological deficit and brain damage (by MRI data) in Wistar rats. Neurological deficit was assessed within 14 days using the limb placement test. MRI and histological study of the brain with an assessment of the size of damage were performed on day 14 after ischemia. Significant differences were obtained in limb placement scores on days 3, 7, and 14, as well as in the volume of ischemic focus by MRI in comparison with the control (ischemia+N2 70%/O2 30%). Inhalation of argon-oxygen mixture for 2 h a day over 3 days after photoinduced stroke decreased the volume of brain damage by 2 times and reduced the severity of neurological deficit.

Cerebral Small Vessel Disease, Hypertension, and Vascular Contributions to Cognitive Impairment and Dementia.

Hypertension-associated cerebral small vessel disease is a common finding in older people. Strongly associated with age and hypertension, small vessel disease is found at autopsy in over 50% of people aged ≥65 years, with a spectrum of clinical manifestations. It is the main cause of lacunar stroke and a major source of vascular contributions to cognitive impairment and dementia. The brain areas affected are subcortical and periventricular white matter and deep gray nuclei. Neuropathological sequelae are diffuse white matter lesions (seen as white matter hyperintensities on T2-weighted magnetic resonance imaging), small ischemic foci (lacunes or microinfarcts), and less commonly, subcortical microhemorrhages. The most common form of cerebral small vessel disease is concentric, fibrotic thickening of small penetrating arteries (up to 300 microns outer diameter) termed arteriolosclerosis. Less common forms are small artery atheroma and lipohyalinosis (the lesions described by C. Miller Fisher adjacent to lacunes). Other microvascular lesions that are not reviewed here include cerebral amyloid angiopathy and venous collagenosis. Here, we review the epidemiology, neuropathology, clinical management, genetics, preclinical models, and pathogenesis of hypertensive small vessel disease. Knowledge gaps include initiating factors, molecular pathogenesis, relationships between arterial pathology and tissue damage, possible reversibility, pharmacological targets, and molecular biomarkers. Progress is anticipated from multicell transcriptomic and proteomic profiling, novel experimental models and further target-finding and interventional clinical studies.

Functional ultrasound imaging of stroke in awake rats

Anesthesia is a major confounding factor in preclinical stroke research as stroke rarely occurs in sedated patients. Moreover, anesthesia affects both brain functions and the stroke outcome acting as neurotoxic or protective agents. So far, no approaches were well suited to induce stroke while imaging hemodynamics along with simultaneous large-scale recording of brain functions in awake animals. For this reason, the first critical hours following the stroke insult and associated functional alteration remain poorly understood. Here, we present a strategy to investigate both stroke hemodynamics and stroke-induced functional alterations without the confounding effect of anesthesia, i.e., under awake condition. Functional ultrasound (fUS) imaging was used to continuously monitor variations in cerebral blood volume (CBV) in +65 brain regions/hemispheres for up to 3 hr after stroke onset. The focal cortical ischemia was induced using a chemo-thrombotic agent suited for permanent middle cerebral artery occlusion in awake rats and followed by ipsi- and contralesional whiskers stimulation to investigate on the dynamic of the thalamocortical functions. Early (0–3 hr) and delayed (day 5) fUS recording enabled to characterize the features of the ischemia (location, CBV loss), spreading depolarizations (occurrence, amplitude) and functional alteration of the somatosensory thalamocortical circuits. Post-stroke thalamocortical functions were affected at both early and later time points (0–3 hr and 5 days) after stroke. Overall, our procedure facilitates early, continuous, and chronic assessments of hemodynamics and cerebral functions. When integrated with stroke studies or other pathological analyses, this approach seeks to enhance our comprehension of physiopathologies towards the development of pertinent therapeutic interventions.

Functional ultrasound imaging of stroke in awake rats.

Anesthesia is a major confounding factor in preclinical stroke research as stroke rarely occurs in sedated patients. Moreover, anesthesia affects both brain functions and the stroke outcome acting as neurotoxic or protective agents. So far, no approaches were well suited to induce stroke while imaging hemodynamics along with simultaneous large-scale recording of brain functions in awake animals. For this reason, the first critical hours following the stroke insult and associated functional alteration remain poorly understood. Here, we present a strategy to investigate both stroke hemodynamics and stroke-induced functional alterations without the confounding effect of anesthesia, i.e., under awake condition. Functional ultrasound (fUS) imaging was used to continuously monitor variations in cerebral blood volume (CBV) in +65 brain regions/hemispheres for up to 3 hr after stroke onset. The focal cortical ischemia was induced using a chemo-thrombotic agent suited for permanent middle cerebral artery occlusion in awake rats and followed by ipsi- and contralesional whiskers stimulation to investigate on the dynamic of the thalamocortical functions. Early (0-3 hr) and delayed (day 5) fUS recording enabled to characterize the features of the ischemia (location, CBV loss), spreading depolarizations (occurrence, amplitude) and functional alteration of the somatosensory thalamocortical circuits. Post-stroke thalamocortical functions were affected at both early and later time points (0-3 hr and 5 days) after stroke. Overall, our procedure facilitates early, continuous, and chronic assessments of hemodynamics and cerebral functions. When integrated with stroke studies or other pathological analyses, this approach seeks to enhance our comprehension of physiopathologies towards the development of pertinent therapeutic interventions.

Micro-ultrasound based characterization of cerebrovasculature following focal ischemic stroke and upon short-term rehabilitation

Notwithstanding recanalization treatments in the acute stage of stroke, many survivors suffer long-term impairments. Physical rehabilitation is the only widely available strategy for chronic-stage recovery, but its optimization is hindered by limited understanding of its effects on brain structure and function. Using micro-ultrasound, behavioral testing, and electrophysiology, we investigated the impact of skilled reaching rehabilitation on cerebral hemodynamics, motor function, and neuronal activity in a rat model of focal ischemic stroke. A 50 MHz micro-ultrasound transducer and intracortical electrophysiology were utilized to characterize neurovascular changes three weeks following focal ischemia elicited by endothelin-1 injection into the sensorimotor cortex. Sprague-Dawley rats were rehabilitated through tray reaching, and their fine skilled reaching was assessed via the Montoya staircase. Focal ischemia led to a sustained deficit in forelimb reaching; and increased tortuosity of the penetrating vessels in the perilesional cortex; with no lateralization of spontaneous neuronal activity. Rehabilitation improved skilled reaching; decreased cortical vascularity; was associated with elevated peri- vs. contralesional hypercapnia-induced flow homogenization and increased perilesional spontaneous cortical neuronal activity. Our study demonstrated neurovascular plasticity accompanying rehabilitation-elicited functional recovery in the subacute stage following stroke, and multiple micro-ultrasound-based markers of cerebrovascular structure and function modified in recovery from ischemia and upon rehabilitation.

Carotid free-floating thrombus stemming from carotid web: co-occurrence of two rare causes of ischemic stroke

BackgroundCarotid web (CaW) and carotid free-floating thrombus (CFFT) are rare yet critical causes of ischemic stroke in young adults.Case presentationA 54-year-old woman presented with a fluctuating right sensory-motor faciobrachial syndrome. A brain MRI scan revealed multiple small recent asynchronous cortico-subcortical ischemic foci in the vascular territory of the left internal carotid artery. A CT angiography identified a CFFT in the left internal carotid artery arising from an underlying CaW. The patient was treated with excellent clinical outcomes with carotid artery stenting and dual antiplatelet therapy.ConclusionsWe provide a structured pathophysiological rationale connecting CaW and CFFT and highlight pivotal therapeutic implications. Further studies are needed to investigate this relationship and guide assessment and treatment.

Abstract 241: Neurologic Complications Associated With Extracorporeal Cardiopulmonary Resuscitation for In-Hospital Cardiac Arrest

Background: Neurologic complications are frequently encountered after ECPR and there is a limited understanding of the associated clinical trajectories following ECPR for in-hospital cardiac arrest (IHCA). Aim: To determine the prevalence, properties, and outcomes of radiographic neurologic complications following ECPR for IHCA. Methods: In this single-center retrospective chart review, we collected data on all adult IHCA patients who underwent ECPR from 1/1/2018 to 12/31/2019. Measured variables included demographics, clinical data, timing of diagnostic studies, and outcomes. Results: Data were analyzed from 36 adult ECPR patients, 16 (44%) of whom survived to hospital discharge. Twenty (56%) underwent head computed tomography (CT) and 9 (25%) had pathology identified, diagnosed a median of 1.8 days afterECPR. The most common complications were intracranial hemorrhage and focal ischemia (4/9 patients each). Neurologic complications were not associated with total ECMO duration (median 4 vs 5 days, p=0.57) and patients with complications trended toward poorer survival outcomes (22% vs 52% survival to discharge; p=0.25). Conclusions: Radiographic neurologic complications occurred in one quarter of IHCA ECPR patients and were associated with trends towards poorer clinical outcomes. Consistent with data in the non-ECPR literature, rates of global anoxic injury and hospital mortality were lower in this IHCA cohort than in previously published OHCA and mixed ECPR cohorts. This study is limited by its small sample size, single center design, and lack of a standardized neuroimaging protocol. Future prospective studies are needed to assess the efficacy of ECPR for IHCA.

Histopathological lesions induced by stroke in the encephalon.

Strokes are conditions with a high degree of morbidity and mortality worldwide. These conditions profoundly affect the quality of life of patients; in addition to physical disabilities, patients present various mental disorders, such as mood disorders, anxiety, depression, behavioral disorders, fatigue, etc. Microscopic lesions of the brain parenchyma explain the clinical symptoms and correlate with the severity of the stroke. Our study consisted of the histopathological (HP) and immunohistochemical analysis of brain fragments, collected from 23 patients, with a clinical and imagistic diagnosis of stroke, who died during hospital admission. The microscopic analysis showed that both neurons and glial cells are affected in the ischemic focus. Neuronal death in the ischemic focus was mostly caused by cell necrosis and only about 10% by apoptosis. Regarding vascular lesions, it was observed that the most frequent HP lesion of intracerebral arterioles was arteriosclerosis. The lumen of the arterioles was reduced, and the vascular endothelium had a discontinuous aspect, which indicates a change in the blood-brain barrier. Sometimes the arteriole lumen was completely obstructed, with fibrinoid necrosis in the internal and middle tunic, or with the proliferation of fibroblasts and the formation of young intraluminal connective tissue. Intraparenchymal blood capillaries in the ischemic area showed endothelium discontinuities, lumen collapse, and sometimes massive perivascular edema. As for neuroinflammation, the presence of numerous neutrophils, lymphocytes, plasma cells and macrophages was found in the ischemic focus, forming a complex and inhomogeneous cellular mixture. Of the inflammatory cells present in the ischemic focus and in the ischemic penumbra area, the most numerous were the macrophages. The HP analysis showed that neuroinflammation is very complex and different in intensity from one patient to another, most likely due to associated comorbidities, age, treatment administered until death, etc.

Transient global amnesia in patients with cerebral autosomal dominant arteriopathy with subcortical infarcts, and leukoencephalopathy (CADASIL)

Transient global amnesia (TGA) is characterized by a sudden onset of total memory dysfunction, and loss of the ability to memorize information, followed by anterograde, and sometimes, retrograde amnesia, and reproduction of current events with patient’s consciousness alert, accompanied by one’s repetitive questioning during this very episode that lasts less than 24 hours (usually 1-8 hours). The process involves mesial temporal lobes and hippocampi. Pathogenesis of TGA includes venous congestion with Valsalva-like activities, other vascular dysfunction, and migraine. The article contains literature review on the development of TGA in patients with Multiple Sclerosis and Cerebral Autosomal Dominant Arteriopathy and Subcortical Infarcts and Leukoencephalopathy (CADASIL), which is not common for such patients, and describes a rare clinical case of confirmed CADASIL, in whom first acute vascular ischemic attack presented by TGA developed. In this patient, CADASIL was diagnosed earlier, and was based on the presence of migraine in anamnesis, common for CADASIL patients MRI abnormalities, and the result of molecular-genetic analysis, which showed gene NOTCH3 heterozygous mutation: р.207 R>R/C. Neurological examination revealed no focal neurological signs. Brain MRI in diffusion option (DWI) revealed a small pinpoint ischemic focus in left hyppocampus, which is characteristic for TGA. Despite of frequent review of migraine and subcortical ischemic events in these patients, the history of TGA is solely reported in literature. Reports of TGA development in patients with multiple sclerosis are also sole.

(S)-roscovitine, a CDK inhibitor, decreases cerebral edema and modulates AQP4 and α1-syntrophin interaction on a pre-clinical model of acute ischemic stroke.

Cerebral edema is one of the deadliest complications of ischemic stroke for which there is currently no pharmaceutical treatment. Aquaporin-4 (AQP4), a water-channel polarized at the astrocyte endfoot, is known to be highly implicated in cerebral edema. We previously showed in randomized studies that (S)-roscovitine, a cyclin-dependent kinase inhibitor, reduced cerebral edema 48 h after induction of focal transient ischemia, but its mechanisms of action were unclear. In our recent blind randomized study, we confirmed that (S)-roscovitine was able to reduce cerebral edema by 65% at 24 h post-stroke (t test, p = .006). Immunofluorescence analysis of AQP4 distribution in astrocytes revealed that (S)-roscovitine decreased the non-perivascular pool of AQP4 by 53% and drastically increased AQP4 clusters in astrocyte perivascular end-feet (671%, t test p = .005) compared to vehicle. Non-perivascular and clustered AQP4 compartments were negatively correlated (R = -0.78; p < .0001), suggesting a communicating vessels effect between the two compartments. α1-syntrophin, AQP4 anchoring protein, was colocalized with AQP4 in astrocyte endfeet, and this colocalization was maintained in ischemic area as observed on confocal microscopy. Moreover, (S)-roscovitine increased AQP4/α1-syntrophin interaction (40%, MW p = .0083) as quantified by proximity ligation assay. The quantified interaction was negatively correlated with brain edema in both treated and placebo groups (R = -.57; p = .0074). We showed for the first time, that a kinase inhibitor modulated AQP4/α1-syntrophin interaction, and was implicated in the reduction of cerebral edema. These findings suggest that (S)-roscovitine may hold promise as a potential treatment for cerebral edema in ischemic stroke and as modulator of AQP4 function in other neurological diseases.

Longer Periods of Hypothermia Provide Greater Protection Against Focal Ischemia: A Systematic Review of Animal Studies Manipulating Treatment Duration.

Decades of animal research show therapeutic hypothermia (TH) to be potently neuroprotective after cerebral ischemic injuries. While there have been some translational successes, clinical efficacy after ischemic stroke is unclear. One potential reason for translational failures could be insufficient optimization of dosing parameters. In this study, we conducted a systematic review of the PubMed database to identify all preclinical controlled studies that compared multiple TH durations following focal ischemia, with treatment beginning at least 1 hour after ischemic onset. Six studies met our inclusion criteria. In these six studies, six of seven experiments demonstrated an increase in cerebroprotection at the longest duration tested. The average effect size (mean Cohen's d ± 95% confidence interval) at the shortest and longest durations was 0.4 ± 0.3 and 1.9 ± 1.1, respectively. At the longest durations, this corresponded to percent infarct volume reductions between 31.2% and 83.9%. Our analysis counters previous meta-analytic findings that there is no relationship, or an inverse relationship between TH duration and effect size. However, underreporting often led to high or unclear risks of bias for each study as gauged by the SYRCLE Risk of Bias tool. We also found a lack of investigations of the interactions between duration and other treatment considerations (e.g., method, delay, and ischemic severity). With consideration of methodological limitations, an understanding of the relationships between treatment parameters is necessary to determine proper "dosage" of TH, and should be further studied, considering clinical failures that contrast with strong cerebroprotective results in most animal studies.