Microglia are the brain's resident immune cells known for their dynamic responses to tissue and vascular injury. However, few studies have explored how microglial activity differs across the life stages of early development, adulthood, and aging. Using two-photon live imaging, we confirm that microglia in the adult cerebral cortex exhibit highly ramified processes and relatively immobile somata under basal conditions. Their responses to focal laser-induced injury occur over minutes and are highly coordinated among neighboring microglia. In contrast, neonatal microglia are denser and more mobile but less morphologically complex. Their responses to focal laser-induced injuries of capillaries or parenchymal tissue are uncoordinated, delayed, and persist over days. In the aged brain, microglia somata remain immobile under basal conditions, but their processes become less ramified. Their responses to focal injuries are coordinated but slower and less sensitive. These studies reveal the marked shifts in microglial morphology, distribution, dynamics, and injury response across the lifespan.

Pharmaceutical-grade medium-chain triglycerides (MCTs) are common excipients for in vivo pharmacological studies in laboratory animals and as an experimental therapeutic in certain metabolic and neurologic disorders. In this study, we examined the tolerability of repeated administration of a pharmaceutical-grade formulation of 3 MCTs-caprylic, capric, and lauric acid-in mice via the oral and intraperitoneal routes. We administered either 8 or 4 µL of 100% MCTs or saline/gram of body weight (∼7.56 or 3.78 g/kg, respectively) twice daily for 7 d. During administration, and for 7 d after, we monitored weight change and clinical presentation. On day 14, or upon meeting euthanasia criteria, animals were sacrificed for gross necropsy, histology, and CBC. We observed significant weight loss, clinical decline, and 100% mortality in animals receiving 8 µL/g MCTs via the intraperitoneal route of administration. Gross necropsy revealed serosanguinous fluid in the thoracic cavity, dark red mottled lungs, and adhesions in the abdominal cavity. Histology confirmed inflammation of the lungs, mediastinum, and peritoneum. Mild pathology and initial weight loss (through day 3) were also present in mice receiving 4 µL/g MCTs IP. However, these animals regained weight by day 7 and exhibited no clinical decline or mortality. These adverse effects were not seen in animals receiving either 8 µL/g MCTs PO or 8 µL/g saline IP. These findings suggest that repeated intraperitoneal administration of MCTs may cause dose-dependent toxicity and mortality at high doses, but it confers no adverse effects when administered via the oral route.

Vascular injury and pathologies underlie common diseases including ischemic stroke and cerebral small vessel disease (CSVD). Prior work has identified a key role for glial cells, including microglia, in the multifaceted and temporally evolving neuroimmune response to both stroke and CSVD. Transcriptional profiling has led to important advances including identification of distinct gene expression signatures in ischemia-exposed, flow cytometrically sorted microglia and more recently single cell RNA sequencing-identified microglial subpopulations or clusters. There is a reassuring degree of overlap in the results from these two distinct methodologies with both identifying a proliferative and a separate type I interferon responsive microglial element. Similar patterns were later seen using multimodal and spatial transcriptomal profiling in ischemia-exposed microglia and astrocytes. Methodological advances including enrichment of specific neuroanatomic/functional regions (such as the neurovascular unit) prior to single cell RNA sequencing has led to identification of novel cellular subtypes and generation of new credible hypotheses as to cellular function based on the enhanced cell sub-type specific gene expression patterns. A ribosomal tagging strategy focusing on the cellular translatome analyses carried out in the acute phases post stroke has revealed distinct inflammation-regulating roles for microglia and astrocytes in this setting. Early spatial transcriptomics experiments using cerebral ischemia models have identified regionally distinct microglial cell clusters in ischemic core versus penumbra. There is great potential for combination of these methods for multi-omics approaches to further elucidate glial responses in the context of both acute ischemic stroke and chronic CSVD.

Background: Acute ischemic stroke (AIS) is a leading cause of death and long-term disability. Both microglia (MG) and infiltrating macrophages (MP) are critical effector cell types in ischemic brain injury and recovery. K Ca 3.1 is a calcium-activated potassium channel that is upregulated in reactive MG and MP. Studies using either genetic deletion or pharmacological inhibition of K Ca 3.1 demonstrated that this channel is critical for pro-inflammatory activation of MG/MP as well as exacerbation of stroke pathophysiology. Senicapoc (ICA-17043) is a K Ca 3.1-specific inhibitor that has been used in human clinical trials for non-neurological indications (sickle cell anemia, asthma) and was proven safe. Here we evaluate the potential for repurposing senicapoc for AIS. Methods: Young adult male and female mice underwent 60 min middle cerebral artery occlusion (MCAO)/reperfusion. MCAO was monitored by laser doppler. Senicapoc’s pharmacokinetic (PK) profile was determined using high performance liquid chromatography - mass spectroscopy. Drug levels in plasma and brain were quantified at multiple time points post administration. Effects of senicapoc on post-stroke release of cytokines/chemokines was determined by multiplex ELISA. Inflammatory infiltrates were quantified with flow cytometry. Efficacy studies included: (i) infarct volume (MRI T2), white matter integrity (DTI) and longitudinal neurobehavioral outcomes (NBO). In-vitro chromogenic assay was used to assess senicapoc’s effect on proteolytic activity of tissue plasminogen activator (tPA). Results: Administration of senicapoc (40 mg/kg, i.p.) twice daily for seven days starting 12 h after MCAO resulted in ~55% reduction in infarct volume with corresponding improvements in NBO. Free senicapoc levels in brain ranged from 20 - 200 nM in stroked mice at 1, 4 & 12 hours following administration. All values exceeded senicapoc’s IC 50 (11 nM) for K Ca 3.1. Senicapoc, at up to 5 μM, had no effect on tPA’s proteolytic activity. Conclusions: We provide proof-of-concept data that senicapoc, administered in an extended temporal window, can markedly reduce infarct volume and improve NBO in a mouse model of AIS. Senicapoc demonstrated favorable PK and CNS penetration. It did not interfere with tPA.

Background: Ischemic preconditioning (IPC) is a robust protective phenomenon whereby brief ischemic exposure confers protection against a subsequent prolonged ischemic challenge. IPC has been studied primarily in gray matter predominant models, however, stroke significantly impacts white matter (WM). We have previously reported development of a WM IPC model in the mouse optic nerve (MON), a fully myelinated CNS WM tract. We identified innate immune signaling pathways as required for axonal protection, however the cell type(s) responsible for IPC in WM are unknown. Here we characterize the effects of microglial depletion on IPC-induced protection against OGD-mediated injury to: (i) axonal compound action potential (CAP) recovery, (ii) axonal structural integrity, (iii) oligodendrocyte viability and (iv) nodes of Ranvier (NoR) in MON. Methods: Following microglial depletion by pharmacologic treatment with colony stimulating factor 1 receptor (CSF1R) inhibitor PLX5622, MONs were exposed to transient ischemia in vivo, acutely isolated, and subjected to oxygen-glucose deprivation (OGD) ex vivo to simulate a severe ischemic injury. Functional and structural axonal recovery was assessed by electrophysiology with recording of CAP and immunofluorescent/confocal microscopy followed by quantitative stereology. Results: Microglial depletion eliminated IPC-mediated protection of axonal function but intriguingly had no effect on recovery after acute ischemic injury alone (i.e. in the absence of IPC). Microglial depletion abrogated IPC-mediated protective effects on both axonal integrity and the survival of mature (APC+) oligodendrocytes after exposure to OGD. IPC-mediated protection was determined to be independent of retinal injury. Effects on NoR remain under investigation. Conclusions: Based on these findings, we conclude that preconditioned, but not naïve, microglia are critical in the endogenous IPC-induced protective response against ischemic injury that occurs in WM. Thus, preconditioned microglia are a critical cellular target for future therapeutics designed to enhance WM recovery from acute ischemic injury (stroke). Furthermore, our data suggest that IPC-mediated protection in WM is anatomically intrinsic to WM.

Senicapoc, a small molecule inhibitor of the calcium-activated potassium channel KCa3.1, was safe and well-tolerated in clinical trials for sickle cell anemia. We previously reported proof-of-concept data suggesting that both pharmacological inhibition and genetic deletion of KCa3.1 reduces infarction and improves neurologic recovery in rodents by attenuating neuroinflammation. Here we evaluated the potential of repurposing senicapoc for ischemic stroke. In cultured microglia, senicapoc inhibited KCa3.1 currents with an IC 50 of 7 nM, reduced Ca 2+ signaling induced by the purinergic agonist ATP, suppressed expression of pro-inflammatory cytokines and enzymes (iNOS and COX-2), and prevented induction of the inflammasome component NLRP3 . When transient middle cerebral artery occlusion (tMCAO, 60 min) was induced in male C57BL/6 J mice, twice daily administration of senicapoc at 10 and 40 mg/kg starting 12 h after reperfusion dose-dependently reduced infarct area determined by T2-weighted magnetic resonance imaging (MRI) and improved neurological deficit on day 8. Ultra-high-performance liquid chromatography/mass spectrometry analysis of total and free brain concentrations demonstrated sufficient KCa3.1 target engagement. Senicapoc treatment significantly reduced microglia/macrophage and T cell infiltration and activation and attenuated neuronal death. A different treatment paradigm with senicapoc started at 3 h and MRI on day 3 and day 8 revealed that senicapoc reduces secondary infarct growth and suppresses expression of inflammation markers, including T cell cytokines in the brain. Lastly, we demonstrated that senicapoc does not impair the proteolytic activity of tissue plasminogen activator (tPA) in vitro. We suggest that senicapoc could be repurposed as an adjunctive immunocytoprotective agent for combination with reperfusion therapy for ischemic stroke.

GliaVolume 71, Issue 8 p. 1787-1789 ISSUE INFORMATION - TABLE OF CONTENTSFree Access Issue Information - Table of Contents First published: 11 June 2023 https://doi.org/10.1002/glia.24207AboutPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL Share a linkShare onEmailFacebookTwitterLinkedInRedditWechat Volume71, Issue8August 2023Pages 1787-1789 RelatedInformation

Ischemic preconditioning (IPC) is a phenomenon whereby a brief, non-injurious ischemic exposure enhances tolerance to a subsequent ischemic challenge. The mechanism of IPC has mainly been studied in rodent stroke models where gray matter (GM) constitutes about 85% of the cerebrum. In humans, white matter (WM) is 50% of cerebral volume and is a critical component of stroke damage. We developed a novel CNS WM IPC model using the mouse optic nerve (MON) and identified the involved immune signaling pathways. Here we tested the hypothesis that microglia are necessary for WM IPC. Microglia were depleted by treatment with the colony stimulating factor 1 receptor (CSF1R) inhibitor PLX5622. MONs were exposed to transient ischemia in vivo, acutely isolated 72 h later, and subjected to oxygen-glucose deprivation (OGD) to simulate a severe ischemic injury (i.e., stroke). Functional and structural axonal recovery was assessed by recording compound action potentials (CAPs) and by microscopy using quantitative stereology. Microglia depletion eliminated IPC-mediated protection. In control mice, CAP recovery was improved in preconditioned MONs compared with non-preconditioned MONs, however, in PLX5622-treated mice, we observed no difference in CAP recovery between preconditioned and non-preconditioned MONs. Microgliadepletion also abolished IPC protective effects on axonal integrity and survival of mature (APC+ ) oligodendrocytes after OGD. IPC-mediated protection was independent of retinal injury suggesting it results from mechanistic processes intrinsic to ischemia-exposed WM. We conclude that preconditioned microglia are critical for IPC in WM. The "preconditioned microglia" phenotype might protect against other CNS pathologies and is a neurotherapeutic horizon worth exploring.

Microglia maintain brain health and play important roles in disease and injury. Despite the known ability of microglia to proliferate, the precise nature of the population or populations capable of generating new microglia in the adult brain remains controversial. We identified Prominin-1 (Prom1; also known as CD133) as a putative cell surface marker of committed brain myeloid progenitor cells. We demonstrate that Prom1-expressing cells isolated from mixed cortical cultures will generate new microglia in vitro. To determine whether Prom1-expressing cells generate new microglia in vivo, we used tamoxifen inducible fate mapping in male and female mice. Induction of Cre recombinase activity at 10 weeks in Prom1-expressing cells leads to the expression of TdTomato in all Prom1-expressing progenitors and newly generated daughter cells. We observed a population of new TdTomato-expressing microglia at 6 months of age that increased in size at 9 months. When microglia proliferation was induced using a transient ischemia/reperfusion paradigm, little proliferation from the Prom1-expressing progenitors was observed with the majority of new microglia derived from Prom1-negative cells. Together, these findings reveal that Prom1-expressing myeloid progenitor cells contribute to the generation of new microglia both in vitro and in vivo. Furthermore, these findings demonstrate the existence of an undifferentiated myeloid progenitor population in the adult mouse brain that expresses Prom1. We conclude that Prom1-expressing myeloid progenitors contribute to new microglia genesis in the uninjured brain but not in response to ischemia/reperfusion. SIGNIFICANCE STATEMENT Microglia, the innate immune cells of the CNS, can divide to slowly generate new microglia throughout life. Newly generated microglia may influence inflammatory responses to injury or neurodegeneration. However, the origins of the new microglia in the brain have been controversial. Our research demonstrates that some newly born microglia in a healthy brain are derived from cells that express the stem cell marker Prominin-1. This is the first time Prominin-1 cells are shown to generate microglia.

We provide results concerning, in a given normal-form game, which sets of actions are best replies to some belief. Proposition 1 states that for any set S of actions, there is a belief under which all actions in S are simultaneously best replies if and only if no mixture of actions in S is strictly dominated. Similarly, Proposition 2 states that for any set S of actions, there is a full-support belief under which all actions in S are best replies if and only if no mixture of actions in S is weakly dominated. One important consequence is Corollary 1: a two-player game has a totally mixed Nash equilibrium if and only if neither player has a pair of mixed strategies such that one weakly dominates the other.

Predictions under common knowledge of payoffs may differ from those under arbitrarily, but finitely, many orders of mutual knowledge; Rubinstein's (1989) Email game is a seminal example. Weinstein and Yildiz (2007) showed that the discontinuity in the example generalizes: for all types with multiple rationalizable (ICR) actions, there exist similar types with unique rationalizable action. This paper studies how a wide class of departures from common belief in rationality impact Weinstein and Yildiz's discontinuity. We weaken ICR to ICR λ , where λ is a sequence whose term λ n is the probability players attach to ( n − 1)th‐order belief in rationality. We find that Weinstein and Yildiz's discontinuity remains when λ n is above an appropriate threshold for all n, but fails when λ n converges to 0. That is, if players' confidence in mutual rationality persists at high orders, the discontinuity persists, but if confidence vanishes at high orders, the discontinuity vanishes.

Ischemic preconditioning (IPC) is an experimental phenomenon in which a subthreshold ischemic insult applied to the brain reduces damage caused by a subsequent more severe ischemic episode. Identifying key molecular and cellular mediators of IPC will provide critical information needed to develop novel therapies for stroke. Here we report that the transcriptomic response of acutely isolated preconditioned cortical microglia is dominated by marked upregulation of genes involved in cell cycle activation and cellular proliferation. Notably, this transcriptional response occurs in the absence of cortical infarction. We employed ex vivo flow cytometry, immunofluorescent microscopy, and quantitative stereology methods on brain tissue to evaluate microglia proliferation following IPC. Using cellular colocalization of microglial (Iba1) and proliferation (Ki67 and BrdU) markers, we observed a localized increase in the number of microglia and proliferating microglia within the preconditioned hemicortex at 72, but not 24, hours post-IPC. Our quantification demonstrated that the IPC-induced increase in total microglia was due entirely to proliferation. Furthermore, microglia in the preconditioned hemisphere had altered morphology and increased soma volumes, indicative of an activated phenotype. Using transgenic mouse models with either fractalkine receptor (CX3CR1)-haploinsufficiency or systemic type I interferon signaling loss, we determined that microglial proliferation after IPC is dependent on fractalkine signaling but independent of type I interferon signaling. These findings suggest there are multiple distinct targetable signaling pathways in microglia, including CX3CR1-dependent proliferation that may be involved in IPC-mediated protection.

Ischemic preconditioning (IPC) is an experimental phenomenon in which a brief ischemic stimulus confers protection against a subsequent prolonged ischemic event. Initially thought to be due to mechanistic changes in neurons, our understanding of IPC has evolved to encompass a global reprogramming of the Central Nervous System (CNS) after transient ischemia/reperfusion that requires innate immune signaling pathways including Toll-like receptors (TLRs) and Type I interferons. Microglia are the CNS resident neuroimmune cells that express these key innate immune receptors. Studies suggest that microglia are required for IPC-mediated neuronal and axonal protection. Multiple paradigms targeting TLRs have converged on a distinctive Type I interferon response in microglia that is critical for preconditioning-mediated protection against ischemia. These pathways can be targeted through administration of TLR agonists, cytokines including interferon-β, and pharmaceutical agents that induce preconditioning through cross-tolerance mechanisms. Transcriptomic analyses and single cell RNA studies point to specific gene expression signatures in microglia that functionally shift these mutable cells to an immunomodulatory or protective phenotype. Although there are technological challenges and gaps in knowledge to overcome, the targeting of specific molecular signaling pathways in microglia is a promising direction for development of novel and effective pharmacotherapies for stroke. Studies on preconditioning in animal models, including nonhuman primates, show promise as prophylactic preconditioning treatments for selected at risk patient populations. In addition, our growing understanding of the mechanisms of IPC-mediated protection is identifying novel cellular and molecular targets for therapeutic interventions that could apply broadly to both acute stroke and chronic vascular cognitive impairment patients.

Ischemic preconditioning (IPC) is a robust neuroprotective phenomenon in which a brief period of cerebral ischemia confers transient tolerance to subsequent ischemic challenge. Research on IPC has implicated cellular, molecular, and systemic elements of the immune response in this phenomenon. Potent molecular mediators of IPC include innate immune signaling pathways such as Toll-like receptors and type 1 interferons. Brain ischemia results in release of pro- and anti-inflammatory cytokines and chemokines that orchestrate the neuroinflammtory response, resolution of inflammation, and transition to neurological recovery and regeneration. Cellular mediators of IPC include microglia, the resident central nervous system immune cells, astrocytes, and neurons. All of these cell types engage in cross-talk with each other using a multitude of signaling pathways that modulate activation/suppression of each of the other cell types in response to ischemia. As the postischemic neuroimmune response evolves over time there is a shift in function toward provision of trophic support and neuroprotection. Peripheral immune cells infiltrate the central nervous system en masse after stroke and are largely detrimental, with a few subtypes having beneficial, protective effects, though the role of these immune cells in IPC is largely unknown. The role of neural progenitor cells in IPC-mediated neuroprotection is another active area of investigation as is the role of microglial proliferation in this setting. A mechanistic understanding of these molecular and cellular mediators of IPC may not only facilitate more effective direct application of IPC to specific clinical scenarios, but also, more broadly, reveal novel targets for therapeutic intervention in stroke.

When a decision-maker is surprised by a pattern unaccounted for in his prior, he may wish to change his beliefs in a way which violates Bayes’ rule, but this practice (under suitable background assumptions) will cause dynamic inconsistency. He may wish to limit this inconsistency. We show that if these non-Bayesian updates, which we call “paradigm shifts,” are rare, in the sense that they occur only with a small probability α according to the decision-maker’s initial belief, the decision-maker will be “approximately” dynamically consistent. Our notion of “approximate” dynamic consistency is that the possible arbitrage against the decision-maker is small compared to the size of his transactions. The quantity α is equivalent to the level of a classical hypothesis test, so our results provide a decision-theoretic foundation for the classical criteria for rejecting a null hypothesis. Our results give the decision-maker some latitude to violate Bayes’ rule while bounding the resulting inconsistency, and justifies the classical criterion for hypothesis testing on decision-theoretic grounds. ∗Previous versions were circulated as “Provisional Beliefs and Paradigm Shifts” and “A Bayesian Foundation for Classical Hypothesis Testing” †I am grateful for conversations with Nabil Al-Najjar, Eddie Dekel, Tai-Wei Hu, Ehud Kalai, Eric Maskin, Allegra Petti, Phil Reny, Larry Samuelson, and Muhamet Yildiz, and seminar participants at Northwestern University, the Transatlantic Theory Conference, University of Montreal, the Stony Brook Game Theory Festival, and University of Pennsylvania. “...I equate the rational attitude and the critical attitude. The point is that, whenever we propose a solution to a problem, we ought to try as hard as we can to overthrow our solution, rather than defend it. Few of us, unfortunately, practice this precept...” Karl Popper, The Logic of Scientific Discovery

Tissue plasminogen activator (tPA) was first approved in the USA 25 years ago for those who had experienced a recent occlusion (<3 h) of a cerebral vessel. Now, advances in clot retrieval (stentriever), in concert with tPA, heralds new optimism for ischemic stroke victims, but adds more pressure to identify therapies that will minimize hypoxic damage, protect compromised cells, and promote rehabilitation. In the past preclinical investigations have been poor at predicting potential clinical therapy, but they have contributed enormously to understanding post-stroke pathology. Current clinical trials ( www.strokecenter.org/trials ) anticipate a broad range of approaches: from hypothermia, to cell therapy, to neuroprotection.

In a Bayesian game, assume that the type space is a complete, separable metric space, the action space is a compact metric space, and the payoff functions are continuous. We show that the iterative and fixed-point definitions of interim correlated rationalizability (ICR) coincide, and ICR is non-empty-valued and upper hemicontinuous. This extends the finite-game results of Dekel et al. (2007), who introduced ICR. Our result applies, for instance, to discounted infinite-horizon dynamic games. (C) 2017 Elsevier B.V. All rights reserved.

Microglia, the resident immune cells of the CNS, are primary regulators of the neuroimmune response to injury. Type I interferons (IFNs), including the IFNαs and IFNβ, are key cytokines in the innate immune system. Their activity is implicated in the regulation of microglial function both during development and in response to neuroinflammation, ischemia, and neurodegeneration. Data from numerous studies in multiple sclerosis (MS) and stroke suggest that type I IFNs can modulate the microglial phenotype, influence the overall neuroimmune milieu, regulate phagocytosis, and affect blood–brain barrier integrity. All of these IFN-induced effects result in numerous downstream consequences on white matter pathology and microglial reactivity. Dysregulation of IFN signaling in mouse models with genetic deficiency in ubiquitin specific protease 18 (USP18) leads to a severe neurological phenotype and neuropathological changes that include white matter microgliosis and pro-inflammatory gene expression in dystrophic microglia. A class of genetic disorders in humans, referred to as pseudo-TORCH syndrome (PTS) for the clinical resemblance to infection-induced TORCH syndrome, also show dysregulation of IFN signaling, which leads to severe neurological developmental disease. In these disorders, the excessive activation of IFN signaling during CNS development results in a destructive interferonopathy with similar induction of microglial dysfunction as seen in USP18 deficient mice. Other recent studies implicate “microgliopathies” more broadly in neurological disorders including Alzheimer’s disease (AD) and MS, suggesting that microglia are a potential therapeutic target for disease prevention and/or treatment, with interferon signaling playing a key role in regulating the microglial phenotype.

Innate immune signaling is important in the pathophysiology of ischemia/reperfusion (stroke)-induced injury and recovery. Several lines of evidence support a central role for microglia in these processes. Recent work has identified Toll-like receptors (TLRs) and type I interferon (IFN) signaling in both ischemia/reperfusion-induced brain injury and ischemic preconditioning-mediated neuroprotection. To determine the effects of “ischemia/reperfusion-like” conditions on microglia, we performed genomic analyses on wild-type (WT) and TLR4−/− cultured microglia after sequential exposure to hypoxia/hypoglycemia and normoxia/normoglycemia (H/H–N/N). We observed increased expression of type 1 IFN-stimulated genes (ISGs) as the predominant transcriptomal feature of H/H–N/N-exposed WT, but not TLR4−/−, microglia. Microarray analysis on ex vivo sorted microglia from ipsilateral male mouse cortex after a transient in vivo ischemic pulse also demonstrated robust expression of ISGs. Type 1 IFNs, including the IFN-αs and IFN-β, activate the interferon-α/β receptor (IFNAR) complex. We confirmed both in vitro H/H–N/N- and in vivo ischemia/reperfusion-induced microglial ISG responses by quantitative real-time PCR and demonstrated that both were dependent on IFNAR1. We characterized the effects of hypoxia/hypoglycemia on phosphorylation of signal transducer and activator of transcription 1 (STAT1), release of type 1 IFNs, and surface expression of IFNAR1 in microglia. We demonstrated that IFN-β induces dose-dependent secretion of ISG chemokines in cultured microglia and robust ISG expression in microglia both in vitro and in vivo. Finally, we demonstrated that the microglial ISG chemokine responses to TLR4 agonists were dependent on TLR4 and IFNAR1. Together, these data suggest novel ischemia/reperfusion-induced pathways for both TLR4-dependent and -independent, IFNAR1-dependent, type 1 IFN signaling in microglia. SIGNIFICANCE STATEMENT Stroke is the fifth leading cause of death in the United States and is a leading cause of serious long-term disability worldwide. Innate immune responses are critical in stroke pathophysiology, and microglia are key cellular effectors in the CNS response to ischemia/reperfusion. Using a transcriptional analysis approach, we identified a robust interferon (IFN)-stimulated gene response within microglia exposed to ischemia/reperfusion in both in vitro and in vivo experimental paradigms. Using a number of complementary techniques, we have demonstrated that these responses are dependent on innate immune signaling components including Toll-like receptor-4 and type I IFNs. We have also elucidated several novel ischemia/reperfusion-induced microglial signaling mechanisms.