Exogenous sex hormones have been extensively studied for their influence on stroke risk and outcome. This meta-analysis served to update the pre-clinical acute ischemic stroke (AIS) literature and provide the first synthesis of the intracerebral hemorrhage (ICH) and subarachnoid hemorrhage (SAH) literature on how estrogen, progesterone, and testosterone affect post-stroke outcomes. This study was pre-registered with PROSPERO (CRD42024544794). Medline, EMBASE, Scopus, and Web of Science were searched; studies using animal models of stroke investigating exogenous estrogen, progesterone, or testosterone, alone or in combination, compared to non-treated controls were included. Assessments of injury volume, edema, and behaviour (neurological deficits, sensorimotor and cognitive outcomes) were analyzed via hierarchical meta-analyses. Risk of bias was assessed via SYRCLE and CAMARADES, and evidence certainty via an adaptation of the GRADE tool. In total, 211 studies were included. Estrogen and progesterone improved all post-AIS outcomes (SMDs = 0.32–1.30, 95

Preclinical studies show that therapeutic hypothermia (TH) effectively reduces cerebral ischemic injury. In contrast, TH has not been consistently beneficial in clinical trials of stroke and cardiac arrest, perhaps from suboptimal dosing (e.g., delay, depth, and duration), among other factors. This systematic review aimed to find an optimal depth of TH from in vivo adult preclinical studies of global and focal ischemia. To study depth, without other confounds, we examined studies that compared ≥2 depths of TH versus normothermic controls. Our primary outcomes were infarct size (focal ischemia) and hippocampal cell death (global ischemia), while secondary outcomes were behavior, edema, and striatal cell death. Studies were assessed with the SYRCLE Risk of Bias tool (e.g., use of blinding) and additional indices of translational rigor (e.g., use of aged animals). Thirty studies were included from a search of the PubMed database in 2025. Many studies were rated as exhibiting a high risk of bias with low translational rigor. Overall, TH provided considerable protection on all endpoints, sometimes up to 100%, but no consistent dose-response patterns emerged, nor was an optimal depth of cooling readily evident. To explore the latter finding, specifically sampling variability, we conducted Monte Carlo simulations using the pooled standard deviation of the preclinical studies to generate three populations based upon a theoretical 5% protection per 1°C relationship (37°C vs. 32°C vs. 27°C groups run 75 times). Dose-dependent effects were statistically detectable in only 36% of comparisons, which showed comparably noisy patterns of protection. Thus, the variable dose-dependent effects in the reviewed animal studies likely arise, at least partially, from sampling error owing to using small samples from variable populations (average n = 8/group in focal ischemia). Overall, these findings highlight weaknesses in the extant dose-response literature that limit our ability to precisely guide clinical trials.

Primary intracranial tumors make up ~ 2% of all cancer cases but contribute disproportionately to cancer-related morbidity and mortality. While spontaneous preclinical intracranial tumor models offer valuable insights into the mechanisms underlying primary tumor formation and progression, few have been identified to date. As such, tracking and characterizing spontaneous tumor development in aging laboratory animals represents an opportunity to advance preclinical research, while informing best practices for animal welfare. Here, we investigated intracranial tumor incidence and origin in a cohort of spontaneously hypertensive rats aged 20–24 months (SHRs; N = 60), initially intended for preclinical stroke experiments. Upon gross inspection, we identified large intracranial tumors in 30% of aged SHRs. Using histological, immunohistological, and ultrastructural methods, we found that ~ 89% of these tumors were pituitary neuroendocrine tumors (NETs), with an average tumor diameter of 6.7 ± 1.22 mm (mean ± 95% confidence interval). These pituitary NETs displayed rapid growth, invasive behavior, and aggressive proliferation, as indicated by Ki67 staining. Our results reveal a predisposition in aged SHRs to develop aggressive pituitary NETs, providing a diagnostic and pathological foundation for future mechanistic and translational studies, while emphasizing the need for enhanced surveillance and care in aging laboratory animal populations.

Objective: Conditions associated with mass effect, such as intracranial tumors and hemorrhagic stroke (intracerebral hemorrhage; ICH), disrupt intracranial pressure (ICP) regulation by exhausting the brain's compliance reserves. Displacement of cerebrospinal fluid, blood, and brain tissue ("tissue compliance") can buffer ICP elevations, but these reserves are limited. Brain tumor patients face elevated ICH risk, yet how pre-existing mass effect influences acute intracranial compliance remains unclear. This study examined how chronic intracranial mass effect alters tissue compliance following acute ICH. Methods: In an aged spontaneously hypertensive rat (SHR) cohort, a high incidence of spontaneous intracranial tumors were discovered. This prompted an exploratory analysis in 18 SHRs (n = 6/group: ICH-24 h, ICH-72 h, sham) to evaluate the impact of chronic mass effect on post-ICH tissue compliance. Following collagenase striatal hemorrhage, brains were collected for macroscopic and microscopic volumetric morphological analysis. Results: Tissue compliance persisted in tumor-bearing animals, reflected by reduced contralateral hemisphere volume (similar to 6% at 24; similar to 14 % at 72 h) and neuronal soma shrinkage in hippocampal and cortical regions. Despite smaller hematoma volumes, rats with tumors exhibited nearly twofold greater tissue compliance responses than non-tumor animals. Conclusion: Pre-existing mass effect may modify the recruitment of intracranial compliance reserves following stroke in aged animals. (c) 2025 International Hemorrhagic Stroke Association. Publishing services by Elsevier B.V. on behalf of KeAi Communications Co. Ltd. This is an open access article under the CC BY license (http://creativecom-mons.org/licenses/by/4.0/).

Intracerebral hemorrhage (ICH) along with aggravating factors, such as edema, can raise intracranial pressure (ICP) to pathological levels. Diversion of some cerebrospinal fluid (CSF) and venous blood out of the cranium can limit ICP rises while maintaining cerebral perfusion pressure. Brain tissue itself is widely considered immutable in volume but prone to distortion (e.g., midline shift). However, distal brain regions shrink acutely following ICH in rodents. Tissue contraction arises from cell shrinkage and increased packing density. This “tissue compliance” is hypothesized to be an additional mechanism to limit ICP rises. Here, we examined whether and by how much parenchyma volume reduction occurs in ICH patients. We conducted a retrospective analysis on computed tomography (CT) scans of 96 ICH patients (average age 63.8 years old, 55

Malnutrition after stroke may lessen the beneficial effects of rehabilitation on motor recovery through influences on both brain and skeletal muscle. Enriched rehabilitation (ER), a combination of environmental enrichment and forelimb reaching practice, is used preclinically to study recovery of skilled reaching after stroke. However, the chronic food restriction typically used to motivate engagement in reaching practice is a barrier to using ER to investigate interactions between nutritional status and rehabilitation. Thus, our objectives were to determine if a modified ER program comprised of environmental enrichment and skilled reaching practice motivated by a short fast would enhance post-stroke forelimb motor recovery and preserve forelimb muscle size and metabolic fiber type, relative to a group exposed to stroke without ER. At one week after photothrombotic cortical stroke, male, Sprague-Dawley rats were assigned to modified ER or standard care for 2 weeks. Forelimb recovery was assessed in the Montoya staircase and cylinder task before stroke and on days 5-6, 22-23, and 33-34 after stroke. ER failed to improve forelimb function in either task (p > 0.05). Atrophy of extensor digitorum communis (EDC) and triceps brachii long head (TBL) muscles was not evident in the stroke-targeted forelimb on day 35, but the area occupied by hybrid fibers was increased in the EDC muscle (p = 0.038). ER bilaterally increased EDC (p = 0.046), but not TBL, muscle size; EDC muscle fiber type was unchanged by ER. While the modified ER did not promote forelimb motor recovery, it does appear to have utility for studying the role of skeletal muscle plasticity in post-stroke recovery.

Intracerebral hemorrhage (ICH) is a stroke subtype with a high mortality rate ( 40

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.

Therapeutic hypothermia (TH) mitigates damage in ischemic stroke models. However, safer and easier TH methods (e.g., pharmacological) are needed to circumvent physical cooling complications. This study evaluated systemic and pharmacologically induced TH using the adenosine A1 receptor agonist, N6-cyclohexyladenosine (CHA), with control groups in male Sprague-Dawley rats. CHA was administered intraperitoneally 10 minutes following a 2-hour intraluminal middle cerebral artery occlusion. We used a 1.5 mg/kg induction dose, followed by three 1.0 mg/kg doses every 6 hours for a total of 4 doses, causing 20-24 hours of hypothermia. Animals assigned to physical hypothermia and CHA-hypothermia had similar induction rates and nadir temperatures, but forced cooling lasted ∼6 hours longer compared with CHA-treated animals. The divergence is likely attributable to individual differences in CHA metabolism, which led to varied durations at nadir, whereas physical hypothermia was better regulated. Physical hypothermia significantly reduced infarction (primary endpoint) on day 7 (mean reduction of 36.8 mm3 or 39% reduction; p = 0.021 vs. normothermic animals; Cohen's d = 0.75), whereas CHA-induced hypothermia did not (p = 0.33). Similarly, physical cooling improved neurological function (physical hypothermia median = 0, physical normothermia median = 2; p = 0.008) and CHA-induced cooling did not (p > 0.99). Our findings demonstrate that forced cooling was neuroprotective compared with controls, but prolonged CHA-induced cooling was not neuroprotective.

Therapeutic hypothermia (TH) lessens ischemic brain injury. Cytoprotective agents can augment protection, although it is unclear which combinations are most effective. The objective of this study is to identify which cytoprotective drug works best with delayed TH. Following PRISMA guidelines, a systematic review (PubMed, Web of Science, MEDLINE, Scopus) identified controlled experiments that used an in vivo focal ischemic stroke model and evaluated the efficacy of TH (delay of >= 1 hour) coupled with cytoprotective agents. This combination was our main intervention compared with single treatments with TH, drug, or no treatment. Endpoints were brain injury and neurological impairment. The CAMARADES checklist for study quality and the SYRCLE's risk of bias tool gauged study quality. Twenty-five studies were included. Most used young, healthy male rats, with only one using spontaneously hypertensive rats. Two studies used mice models, and six used adult animals. Study quality was moderate (median score = 6), and risk of bias was high. Pharmacological agents provided an additive effect on TH for all outcomes measured. Magnesium coupled with TH had the greatest impact compared with other agent-TH combinations on all outcomes. Longer TH durations improved both behavioral and histological outcomes and had greater cytoprotective efficacy than shorter durations. Anti-inflammatories were the most effective in reducing infarction (standardized mean difference [SMD]: -1.64, confidence interval [CI]: [-2.13, -1.15]), sulfonylureas reduced edema the most (SMD: -2.32, CI: [-3.09, -1.54]), and antiapoptotic agents improved behavioral outcomes the most (normalized mean difference: 52.38, CI: [45.29, 59.46]). Statistically significant heterogeneity was observed (I-2 = 82 - 98%, all p < 0.001), indicating that studies wildly differ in their effect size estimates. Our results support the superiority of adding cytoprotective therapies with TH (vs. individual or no therapy). Additional exploratory and confirmatory studies are required to identify and thoroughly assess combination therapies owing to limited work and inconsistent translational quality.

Intracerebral hemorrhage (ICH) is a stroke subtype with a high mortality rate (similar to 40%). After ICH, the mass effect of the hematoma and edema contribute to raised intracranial pressure (ICP) and poor outcome. Endogenous compensatory mechanisms that blunt ICP elevations include redirection of venous blood and cerebrospinal fluid, along with brain tissue compliance (e.g., decreased cell volume, increased cell density); however, these limited reserves can be exhausted after severe stroke, resulting in decompensated ICP that requires careful clinical management. Management strategies can include administration of hypertonic saline (HTS), an osmotic agent that putatively attenuates edema, and thereby ICP elevations. Evidence regarding the efficacy of HTS treatment following ICH remains limited. In this study, adult male rats were given a collagenase-induced striatal ICH and a bolus of either 3% HTS or 0.9% saline vehicle at 2- and 14-hours post-stroke onset. Neurological deficits, edema, ipsilateral cell volume and density (in areas S1 and CA1), and contralateral CA1 ultrastructural morphology were assessed 24 h post-ICH. Animals had large bleeds (median 108.2 mu L), extensive edema (median 83.9% brain water content in ipsilateral striatum), and evident behavioural deficits (median 5.4 neurological deficit scale score). However, HTS did not affect edema (p >= 0.4797), behaviour (p = 0.6479), cell volume (p >= 0.1079), or cell density (p >= 0.0983). Qualitative ultrastructural assessment of contralateral area CA1 suggested that HTS administration was associated with paradoxical cellular swelling in ICH animals. Overall, there was no benefit with administering 3% HTS after ICH.

Ketone bodies, or ketones, are an alternative energy source and have several nonmetabolic signaling actions, such as inhibiting inflammation. Because of this, exogenous ketone supplementation has been used to help treat various diseases. β-hydroxybutyrate (βHB) is the major ketone body that has reduced neurological injury and brain edema in animal models of ischemic stroke and traumatic brain injury. However, the therapeutic potential of βHB in intracerebral hemorrhage (ICH) has not yet been determined. Here we investigated the effects of exogenous βHB treatment following ICH on inflammation, edema, injury size, and functional outcomes. To do this, we administered 250 mg/kg of βHB (subcutaneously every 12 hours) starting 2 hours after collagenase-induced ICH in rats over 3 experiments. First, we observed that βHB-treated rats had significant reductions in transcript expression of pro-inflammatory markers Il1b (p = 0.0210), Tnfa (p = 0.0108), and Mcp1 (p = 0.0473) 3 days post-ICH. Second, βHB significantly improved neurological deficits measured by the neurological deficit scale on day 3 (p = 0.0416) in another cohort of rats, despite no treatment effect on edema (p = 0.2110). To test whether the effects of acute βHB treatment (for 7 days post-ICH) were chronically sustained, the third experiment used serial behavioural testing which confirmed that βHB significantly improved neurological deficit scores (p = 0.0459) 3 days post-ICH. These effects were not sustained at 7, 14, and 28 days post-ICH (all p≥0.1546). Similarly, βHB treatment did not yield differences in forelimb use asymmetry (all p>0.45) or brain lesion volume (p = 0.3381), the primary endpoint of this study. Thus, our studies show that an acute βHB treatment post-ICH can provide some early signs of functional benefit without evidence of lasting effects or neuroprotection. However, it remains to be tested whether other βHB dosing regimens may favorably affect these and other neurological, behavioral, and biochemical parameters, particularly given the early signals of reduced striatal inflammation.

Intracerebral hemorrhage (ICH) is a devastating stroke with many mechanisms of injury. Edema worsens outcome and can lead to mortality after ICH. Glibenclamide (GLC), a sulfonylurea 1- transient receptor potential melastatin 4 (Sur1-Trpm4) channel blocker, has been shown to attenuate edema in ischemic stroke models, raising the possibility of benefit in ICH. This meta-analysis synthesizes current pre-clinical (rodent) literature regarding the efficacy of post-ICH GLC administration (vs. vehicle controls) on behaviour (i.e., neurological deficit, motor, and memory outcomes), edema, hematoma volume, and injury volume. Six studies (5 in rats and 1 in mice) were included in our meta-analysis (PROSPERO registration = CRD42021283614). GLC significantly improved behaviour (standardized mean difference (SMD) = -0.63, [-1.16, -0.09], n = 70-74) and reduced edema (SMD = -0.91, [-1.64, -0.18], n = 70), but did not affect hematoma volume (SMD = 0.0788, [-0.5631, 0.7207], n = 18-20), or injury volume (SMD = 0.2892, [-0.4950, 1.0734], n = 24). However, these results should be interpreted cautiously. Findings were conflicted with 2 negative and 4 positive reports, and Egger regressions indicated missing negative edema data (p = 0.0001), and possible missing negative behavioural data (p = 0.0766). Experimental quality assessed via the SYRCLE and CAMARADES checklists was concerning, as most studies demonstrated high risks of bias. Studies were generally low-powered (e.g., average n = 14.4 for behaviour), and future studies should employ sample sizes of 41 to detect our observed effect size in behaviour and 33 to detect our observed effect in edema. Overall, missing negative studies, low study quality, high risk of bias, and incomplete attention to key recommendations (e.g., investigating female, aged, and co-morbid animals) suggest that further high-powered confirmatory studies are needed before conclusive statements about GLC's efficacy in ICH can be made, and before further clinical trials are performed.