Aberrant cortical activity, functional connectivity, and neural assembly architecture after photothrombotic stroke in mice.

Despite substantial progress in mapping the trajectory of network plasticity resulting from focal ischemic stroke, the extent and nature of changes in neuronal excitability and activity within the peri-infarct cortex of mice remains poorly defined. Most of the available data have been acquired from anesthetized animals, acute tissue slices, or infer changes in excitability from immunoassays on extracted tissue, and thus may not reflect cortical activity dynamics in the intact cortex of an awake animal. Here, in vivo two-photon calcium imaging in awake, behaving mice was used to longitudinally track cortical activity, network functional connectivity, and neural assembly architecture for 2 months following photothrombotic stroke targeting the forelimb somatosensory cortex. Sensorimotor recovery was tracked over the weeks following stroke, allowing us to relate network changes to behavior. Our data revealed spatially restricted but long-lasting alterations in somatosensory neural network function and connectivity. Specifically, we demonstrate significant and long-lasting disruptions in neural assembly architecture concurrent with a deficit in functional connectivity between individual neurons. Reductions in neuronal spiking in peri-infarct cortex were transient but predictive of impairment in skilled locomotion measured in the tapered beam task. Notably, altered neural networks were highly localized, with assembly architecture and neural connectivity relatively unaltered a short distance from the peri-infarct cortex, even in regions within 'remapped' forelimb functional representations identified using mesoscale imaging with anaesthetized preparations 8 weeks after stroke. Thus, using longitudinal two-photon microscopy in awake animals, these data show a complex spatiotemporal relationship between peri-infarct neuronal network function and behavioral recovery. Moreover, the data highlight an apparent disconnect between dramatic functional remapping identified using strong sensory stimulation in anaesthetized mice compared to more subtle and spatially restricted changes in individual neuron and local network function in awake mice during stroke recovery.

Ischemic Stroke.

This concise review of the epidemiology, pathophysiology, evaluation, acute management, and prevention of ischemic stroke targets internists, family practitioners, and emergency physicians who manage patient with stroke.

Adrenergic inhibition facilitates normalization of extracellular potassium after cortical spreading depolarization.

Cortical spreading depolarization (CSD) is a propagating wave of tissue depolarization characterized by a large increase of extracellular potassium concentration and prolonged subsequent electrical silencing of neurons. Waves of CSD arise spontaneously in various acute neurological settings, including migraine aura and ischemic stroke. Recently, we have reported that pan-inhibition of adrenergic receptors (AdRs) facilitates the normalization of extracellular potassium after acute photothrombotic stroke in mice. Here, we have extended that mechanistic study to ask whether AdR antagonists also modify the dynamics of KCl-induced CSD and post-CSD recovery in vivo. Spontaneous neural activity and KCl-induced CSD were visualized by cortex-wide transcranial Ca2+ imaging in G-CaMP7 transgenic mice. AdR antagonism decreased the recurrence of CSD waves and accelerated the post-CSD recovery of neural activity. Two-photon imaging revealed that astrocytes exhibited aberrant Ca2+ signaling after passage of the CSD wave. This astrocytic Ca2+ activity was diminished by the AdR antagonists. Furthermore, AdR pan-antagonism facilitated the normalization of the extracellular potassium level after CSD, which paralleled the recovery of neural activity. These observations add support to the proposal that neuroprotective effects of AdR pan-antagonism arise from accelerated normalization of extracellular K+ levels in the setting of acute brain injury.

Cholinergic upregulation by optogenetic stimulation of nucleus basalis after photothrombotic stroke in forelimb somatosensory cortex improves endpoint and motor but not sensory control of skilled reaching in mice.

A network of cholinergic neurons in the basal forebrain innerve the forebrain and are proposed to contribute to a variety of functions including cortical plasticity, attention, and sensorimotor behavior. This study examined the contribution of the nucleus basalis cholinergic projection to the sensorimotor cortex on recovery on a skilled reach-to-eat task following photothrombotic stroke in the forelimb region of the somatosensory cortex. Mice were trained to perform a single pellet skilled reaching task and their pre and poststroke performance, from Day 4 to Day 28 poststroke, was assessed frame-by-frame by video analysis with endpoint, movement and sensorimotor integration measures. Somatosensory forelimb lesions produced impairments in endpoint and movement component measures of reaching and increased the incidence of fictive eating, a sensory impairment in mistaking a missed reach for a successful reach. Upregulated acetylcholine (ACh) release, as measured by local field potential recording, elicited via optogenetic stimulation of the nucleus basalis improved recovery of reaching and improved movement scores but did not affect sensorimotor integration impairment poststroke. The results show that the mouse cortical forelimb somatosensory region contributes to forelimb motor behavior and suggest that ACh upregulation could serve as an adjunct to behavioral therapy for acute treatment of stroke.

Deep Brain Stimulation for Stroke: Continuous Stimulation of the Pedunculopontine Tegmental Nucleus has no Impact on Skilled Walking in Rats After Photothrombotic Stroke.

BACKGROUND: Gait impairment after stroke is considered as a loss of cerebral function but is also the result of dysfunctional cerebral signals travelling to the spinal motor centres. A therapeutic option to restore disturbed cerebral network activity is deep brain stimulation (DBS). METHODS: A promising target for neuromodulation might be the pedunculopontine tegmental nucleus (PPTg), which contributes to the initiation and control of gait. To test this hypothesis, we trained eighteen rats to cross a horizontal ladder and a wooden beam before inflicting a photothrombosis in the right sensorimotor cortex and implanting a stimulating electrode in the ipsilateral PPTg. RESULTS: Continuous high-frequency DBS (130 Hz; amplitude 55 ± 5 µA) of rats for 10 days yielded no significant improvement of skilled walking when examined with the ladder rung walking test and beam walking test compared to sham-stimulation. CONCLUSION: In contrast to DBS of the cuneiform nucleus, PPTg-stimulation improves neither control of gait nor balance after stroke.

Theta-burst transcranial magnetic stimulation promotes stroke recovery by vascular protection and neovascularization.

Rationale: The integrity and function of the blood-brain barrier (BBB) is compromised after stroke. The current study was performed to examine potential beneficial effects and underlying mechanisms of repetitive transcranial magnetic stimulation (rTMS) on angiogenesis and vascular protection, function, and repair following stroke, which are largely unknown. Methods: Using a rat photothrombotic (PT) stroke model, continuous theta-burst rTMS was administered once daily to the infarcted hemisphere for 5 min, beginning 3 h after PT stroke. This treatment was applied for 6 days. BBB integrity, blood flow, vascular associated proteins, angiogenesis, integrity of neuronal morphology and structure, and behavioral outcome were measured and analyzed at 6 and/or 22 days after PT stroke. Results: We report that rTMS significantly mitigated BBB permeabilization and preserved important BBB components ZO-1, claudin-5, occludin, and caveolin-1 from PT-induced degradation. Damage to vascular structure, morphology, and perfusion was ameliorated by rTMS, resulting in improved local tissue oxygenation. This was accompanied with robust protection of critical vascular components and upregulation of regulatory factors. A complex cytokine response was induced by PT, particularly at the late phase. Application of rTMS modulated this response, ameliorating levels of cytokines related to peripheral immune cell infiltration. Further investigation revealed that rTMS promoted and sustained post-ischemic angiogenesis long-term and reduced apoptosis of newborn and existing vascular endothelial cells. Application of rTMS also inhibited PT-induced excessive astrocyte-vasculature interactions and stimulated an A1 to A2 shift in vessel-associated astrocytes. Mechanistic studies revealed that rTMS dramatically increased levels of PDGFRß associated with A2 astrocytes and their adjacent vasculature. As well, A2 astrocytes displayed marked amplification of the angiogenesis-related factors VEGF and TGFß. PT induced a rise in vessel-associated expression of HIF-1α that was starkly intensified by rTMS treatment. Finally, rTMS preserved neuronal morphology, synaptic structure integrity and behavioral outcome. Conclusions: These results indicate that rTMS can exert powerful protective and restorative effects on the peri-infarct microvasculature after PT stroke by, in part, promoting HIF-1α signaling and shifting vessel-associated astrocytic polarization to the A2 phenotype. This study provides further support for the potent protective effects of rTMS in the context of ischemic stroke, and these findings implicate vascular repair and protection as an important underlying phenomenon.

Quantitative Morphology of Cerebral Thrombi Related to Intravital Contraction and Clinical Features of Ischemic Stroke.

BACKGROUND AND PURPOSE: The purpose was to assess quantitatively and qualitatively the composition and structure of cerebral thrombi and correlate them with the signs of intravital clot contraction (retraction), as well as with etiology, severity, duration, and outcomes of acute ischemic stroke. METHODS: We quantified high-resolution scanning electron micrographs of 41 cerebral thrombi for their detailed cellular and noncellular composition and analyzed histological images for the overall structure with the emphasis on red blood cell compression, fibrin age, and the signs of inflammation. RESULTS: Cerebral thrombi were quite compact and had extremely low porosity. The prevailing cell type was polyhedral compressed erythrocytes (polyhedrocytes) in the core, and fibrin-platelet aggregates were concentrated at the periphery; both findings are indicative of intravital contraction of the thrombi. The content of polyhedrocytes directly correlated with the stroke severity. The prevalence of fibrin bundles was typical for more severe cases, while the content of fibrin sponge prevailed in cases with a more favorable course. The overall platelet content in cerebral thrombi was surprisingly small, while the higher content of platelet aggregates was a marker of stroke severity. Fibrillar types of fibrin prevailed in atherothrombogenic thrombi. Older fibrin prevailed in thrombi from the patients who received thrombolytics, and younger fibrin dominated in cardioembolic thrombi. Alternating layers of erythrocytes and fibrin mixed with platelets were common for thrombi from the patients with more favorable outcomes. Thrombi with a higher number of leukocytes were associated with fatal cases. CONCLUSIONS: Most cerebral thrombi undergo intravital clot contraction (retraction) that may be of underestimated clinical importance. Despite the high variability of the composition and structure of cerebral thrombi, the content of certain types of blood cells and fibrin structures combined with the morphological signs of intravital contraction correlate with the clinical course and outcomes of acute ischemic stroke.

Treatment with AAV1-Rheb(S16H) provides neuroprotection in a mouse model of photothrombosis-induced ischemic stroke.

We recently reported that upregulation of the constitutively active ras homolog enriched in brain [Rheb(S16H)], which induces the activation of the mammalian target of rapamycin complex 1 (mTORC1) signaling pathway, can protect adult neurons, mediated by the induction of neurotrophic factors, such as brain-derived neurotrophic factor (BDNF), in animal models of neurodegenerative diseases. Here we show that neuronal transduction of Rheb(S16H) using adeno-associated virus serotype 1 provides neuroprotection in a mouse model of photothrombosis-induced ischemic stroke. Rheb(S16H)-expressing neurons exhibited neurotrophic effects, such as mTORC1 activation, increases in neuronal size, and BDNF production, in mouse cerebral cortex. Moreover, the upregulation of neuronal Rheb(S16H) significantly attenuated ischemic damage and behavioral impairments as compared to untreated mice, suggesting that Rheb(S16H) upregulation in cortical neurons may be a useful strategy to treat ischemic stroke.

Epicortical Brevetoxin Treatment Promotes Neural Repair and Functional Recovery after Ischemic Stroke.

Emerging literature suggests that after a stroke, the peri-infarct region exhibits dynamic changes in excitability. In rodent stroke models, treatments that enhance excitability in the peri-infarct cerebral cortex promote motor recovery. This increase in cortical excitability and plasticity is opposed by increases in tonic GABAergic inhibition in the peri-infarct zone beginning three days after a stroke in a mouse model. Maintenance of a favorable excitatory-inhibitory balance promoting cerebrocortical excitability could potentially improve recovery. Brevetoxin-2 (PbTx-2) is a voltage-gated sodium channel (VGSC) gating modifier that increases intracellular sodium ([Na+]i), upregulates N-methyl-D-aspartate receptor (NMDAR) channel activity and engages downstream calcium (Ca2+) signaling pathways. In immature cerebrocortical neurons, PbTx-2 promoted neuronal structural plasticity by increasing neurite outgrowth, dendritogenesis and synaptogenesis. We hypothesized that PbTx-2 may promote excitability and structural remodeling in the peri-infarct region, leading to improved functional outcomes following a stroke. We tested this hypothesis using epicortical application of PbTx-2 after a photothrombotic stroke in mice. We show that PbTx-2 enhanced the dendritic arborization and synapse density of cortical layer V pyramidal neurons in the peri-infarct cortex. PbTx-2 also produced a robust improvement of motor recovery. These results suggest a novel pharmacologic approach to mimic activity-dependent recovery from stroke.