Fluids and Barriers of the CNS

Purpose In idiopathic normal pressure hydrocephalus (iNPH), longitudinal monitoring of ventricular size is important for diagnosis and treatment follow-up. This study aimed to validate a fully automated AI model for CT ventricular volumetry with subcompartments and to compare AI-derived volume changes with routine radiology assessments. Methods This retrospective, single-center study included 88 patients with iNPH and 456 non-contrast-enhanced head CT examinations. The model was trained on 38 manually labeled CT scans with 12 ventricular subcompartments. Outcomes included segmentation accuracy, correspondence between AI-derived longitudinal ventricular volume changes and radiology report categories (decreased, unchanged, increased), radiologist detection thresholds for ventricular change, and paired pre- and postoperative volume changes in 22 patients with ventriculoperitoneal shunt. Results Mean segmentation accuracy was high (Dice, 0.83). 91% of 100 segmentations were rated as excellent by an expert neuroradiologist. AI-derived ventricular volume changes corresponded well to radiology report categories (median total ventricular volume changes of -17% in cases reported as decreased, 0% in unchanged cases, and +22% in increased cases; all p < 0.001). Radiologists reported ventricular volume change in 50% of cases at an AI-measured relative volume change of +/-6%, and in 90% of cases at +21% for enlargement and -18% for decrease. After shunt placement, ventricular volume decreased by -8% (median), with the largest relative reductions observed in the right temporal and occipital horns. Conclusions Automated AI-based ventricular segmentation on CT enables accurate and reproducible assessment of ventricular volume changes in iNPH and complements routine radiological evaluation for longitudinal and postoperative monitoring.

A range of neurological pathologies can cause secondary hydrocephalus. For decades, treatment has been limited to surgical CSF diversion, as still no pharmacological options exist due to the elusive molecular nature of the CSF secretion apparatus and its regulatory properties. We have now identified phospholipid lysophosphatidic acid (LPA) as a biomarker for posthemorrhagic hydrocephalus (PHH) in patients with subarachnoid hemorrhage (SAH) and mimicked our results in an animal model of intraventricular hemorrhage (IVH). Intraventricular administration of LPA caused elevated brain water content and ventriculomegaly in experimental rats, via its action as an agonist of the choroidal transient receptor potential vanilloid 4 (TRPV4) channel. TRPV4 was revealed as a novel regulator of ICP in experimental rats via its ability to modulate the CSF secretion rate through its direct activation of the Na+, K+, 2Cl- cotransporter (NKCC1) implicated in CSF secretion. Together, our data reveal that a biomarker present in brain pathologies with hemorrhagic events promotes CSF hypersecretion and ensuing brain water accumulation via its direct action on TRPV4 and its downstream regulation of NKCC1. TRPV4 may therefore be a promising future pharmacological target for pathologies involving disturbed brain fluid dynamics.

The blood-tumor barrier (BTB) prevents effective central nervous system (CNS) drug delivery, especially in malignant gliomas. Brain endothelium predominates the BTB and connects through bicellular and tricellular tight junctions (TJ). Angulin-1/LSR, is a highly expressed endothelial tricellular TJ. Our studies explore the role of Angubindin-1, an Angulin-1/LSR binder, to disrupt tricellular TJ integrity, increase drug entry and hamper glioma progression. Using rat brain endothelial cells (RBMVEC) we tracked Angulin-1/LSR localization and expression to the membrane; binding tightest to Angubindin-1 2-8 hours post-treatment (p < 0.05). Angubindin-1 dose-dependently reduced bicellular and tricellular TJs 1-4 hours post treatment (p < 0.05), returning to baseline by 24 hours (p < 0.05). In human and rat-derived glioma cells, Angubindin-1 transiently reduced Angulin-1/LSR expression between 2-8 hours (p < 0.05), with return to baseline by 24 hours (p < 0.001). Silenced Angulin-1/LSR expression on endothelium resulted in decreased mRNA levels of bicellular (occludin, claudin-5, ZO-1) and tricellular (tricellulin/MARVELD2, angulin-1/LSR) TJs compared to control (p < 0.01). Angubindin-1 treatment also inhibited efflux transporter P-gp in both RBMVECs and glioma cells with high P-gp expression only. Orthotopic rat glioma models were treated with Doxil (3 mg/kg), Angubindin-1 (10 mg/kg), or combination to evaluate BTB permeability/drug accumulation, and overall survival. Combination therapy enhanced Doxil tumor accumulation by 20% (p < 0.001), reduced tumor volume by day 14 (77.5% vs. 81.6%, p < 0.05), and significantly extended survival compared to Doxil alone (24 days vs. 18 days, p < 0.0001). These findings demonstrate the effects of tricellular tight junction inhibition on disrupting the BTB, enhancing CNS drug delivery, and improving rodent glioma survival. SignificanceThis study demonstrates that Angubindin-1, a targeted modulator of tricellular tight junction protein Angulin-1/LSR, transiently disrupts BTB integrity to enhance chemotherapy delivery and prolong survival in glioma-bearing rats. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=145 SRC="FIGDIR/small/667901v1_ufig1.gif" ALT="Figure 1"> View larger version (76K): org.highwire.dtl.DTLVardef@b1b46corg.highwire.dtl.DTLVardef@bc552dorg.highwire.dtl.DTLVardef@7c1a74org.highwire.dtl.DTLVardef@1ace101_HPS_FORMAT_FIGEXP M_FIG C_FIG Angubindin-1 targets both bicellular tight junctions and the tricellular tight junction protein, Angulin-1/LSR, in brain endothelial and glioma cells leading to transient disruption of the blood-tumor barrier (BTB) and inhibition of P-glycoprotein towards enhanced Doxil penetration and reduced tumor burden.

Cerebrospinal fluid (CSF) is a Newtonian fluid that bathes the brain and spinal cord and oscillates in response to the physiological periodic changes in brain volume, of which the cardiac cycle is a major driver. Understanding this motion is essential for clarifying its contribution to solute transport, waste clearance, and drug delivery. In this work, we study oscillatory and steady streaming flow in the cranial subarachnoid space using a lubrication-based theoretical framework. The model represents the cranial CSF compartment as a thin fluid layer bounded internally by the brain surface and externally by the dura, driven by time-dependent brain surface displacements. We first derive simplified governing equations for flow over an arbitrary smooth sphere-like brain surface and obtain analytical solutions for an idealised spherical geometry with uniform displacements. We then incorporate realistic displacement fields reconstructed from MRI measurements in healthy subjects and solve the reduced equations numerically. The results show that oscillatory forcing produces a steady streaming component that may enhance solute transport compared with diffusion alone. This work provides a mechanistic description of the flow generated by physiological brain motion and highlights the potential presence of steady streaming in cranial subarachnoid fluid dynamics.

IntroductionIdiopathic normal pressure hydrocephalus is characterized by gait disturbance, cognitive decline, and urinary dysfunction which may improve with ventriculo-peritoneal shunting. This study evaluated the role of conventional MRI features and the CSF tap test (CSF-TT) in predicting shunt responsiveness in iNPH. MethodsThis is an Ambispective cohort study of 40 patients with probable iNPH evaluated between 2019 and 2024. Baseline MRI parameters, gait features, iNPH score, CSF opening pressure were analyzed. Functional outcome was assessed using the modified Rankin scale (mRS) at baseline, 24 hours after CSF-TT, and 24 weeks after VP shunt surgery. CSF-TT responders were defined as at least a 1-point improvement in mRS 24 hours after CSF-TT. The diagnostic performance of individual MRI parameters and composite diagnostic parameters were evaluated. ResultsForty patients underwent CSF-TT. There were no significant differences between CSF-TT responders and non-responders in the baseline clinical gait parameters, the iNPH scale, and MRI findings. Turning disturbance, wide-based stride, and reduced foot clearance showed significant improvement after CSF-TT. Individual MRI parameters and CSF-TT parameters showed limited value in predicting shunt responsiveness. Composite diagnostic criteria combining CSF-TT and CSF opening pressure >18cm H20 showed sensitivity of 62.5 % and specificity of 71.4% with the highest Youden index indicating modest diagnostic accuracy in predicting shunt responsiveness. ConclusionShunt surgery provided significant functional benefit in iNPH. Neither the CSF tap test nor conventional MRI markers alone reliably predicted shunt responsiveness. A multimodal assessment combining imaging, clinical evaluation, and CSF dynamics is required to optimize patient selection.

Traditionally, the meninges are described as 3 distinct layers, dura, arachnoid and pia. Yet, the classification of the connective meningeal membranes surrounding the brain is based on postmortem macroscopic examination. Ultrastructural and single cell transcriptome analyses have documented that the 3 meningeal layers can be subdivided into several distinct layers based on cellular characteristics. We here re-examined the existence of a 4th meningeal membrane, Subarachnoid Lymphatic-like Membrane or SLYM in Prox1-eGFP reporter mice. Imaging of freshly resected whole brains showed that SLYM covers the entire brain and brain stem and forms a roof shielding the subarachnoid cerebrospinal fluid (CSF)-filled cisterns and the pia-adjacent vasculature. Thus, SLYM is strategically positioned to facilitate periarterial influx of freshly produced CSF and thereby support unidirectional glymphatic CSF transport. Histological analysis showed that, in spinal cord and parts of dorsal cortex, SLYM fused with the arachnoid barrier layer, while in the basal brain stem typically formed a 1-3 cell layered membrane subdividing the subarachnoid space into two compartments. However, great care should be taken when interpreting the organization of the delicate leptomeningeal membranes in tissue sections. We show that hyperosmotic fixatives dehydrate the tissue with the risk of shrinkage and dislocation of these fragile membranes in postmortem preparations.

Cerebrospinal fluid (CSF) flows through the perivascular spaces (PVSs) surrounding cerebral arteries. Revealing the mechanisms driving that flow could bring improved understanding of brain waste transport and insights for disorders including Alzheimers disease and stroke. In vivo velocity measurements of CSF in surface PVSs in mice have been used to argue that flow is driven primarily by the pulsatile motion of artery walls -- perivascular pumping. However, fluid dynamics theory and simulation have predicted that perivascular pumping produces flows differing from in vivo observations starkly, particularly in the phase and relative amplitude of flow oscillation. Here we show that coupling theoretical and simulated flows to realistic end boundary conditions, using resistance and compliance values measured in mice, results in velocities that match observations closely in phase, relative amplitude of oscillation, and mean flow speed. This new, quantitative agreement among theory, simulation, and in vivo measurement further supports the idea that perivascular pumping is a primary CSF driver in physiological conditions.

BackgroundChoroid plexus (ChP) produces cerebrospinal fluid (CSF), and regulates brain development and adult subventricular zone (SVZ) neurogenesis, but its role in hippocampal subgranular zone (SGZ) neurogenesis in adulthood and early postnatal stages is not well understood. Current tools to directly manipulate neonatal ChP/CSF volume are very limited, representing an urgent need in the field. MethodsWe first discovered the specific "leaky" expression of DTR gene in the ChP of adult ROSA26-iDTR mice which can be used to specifically ablate ChP in adult brain that generated robust and long-lasting ablation of ChP and reduction of CSF volume. In this study, we the effectiveness of ROSA26-iDTR allele in ablating neonatal ChP. We also developed a novel AAV2/5-CMV-DTR vector with validated ChP tropism in both neonatal and adult mice, which induces substantial CSF loss in both neonates and adult mice. With both the ROSA26-iDTR genetic and AAV2/5-DTR viral-mediated ChP ablation in young adults and at defined postnatal ages, we quantified ventricular CSF volume by MRI and characterized postnatal neurogenesis. Doublecortin-positive (DCX+) neuroblasts, Ki67+ proliferating cells, and TUNEL+ apoptotic cells were quantified in SVZ and SGZ using confocal microscopy and machine learning-assisted cell counting. ResultsWe show that ROSA26-iDTR-mediated ChP ablation is inefficient before postnatal day 10, suggesting that this line may be of limited utility for CSF reduction in the early neonatal period before P10. P3-5 Dtx treatment of a previously used dosage of 20ng/g dosage did not lead to a reduction in CSF volume. Higher dosage of 40ng/gX3 Dtx dosage at p3-5 generated only moderate partial reduction of CSF in third ventricle and total CSF volume, with indication of toxicity associated with high Dtx dosage in general. In contrast, p10-12 injection of 20ng/gX3 Dtx led to robust CSF reduction. To target early neonatal days, AAV2/5 CMV-DTR virus shows high tropism for ChP epithelial cells and leads to near-complete ablation of CSF in neonatal brains. ChP/CSF loss in neonates or young adult mice leads to a substantial reduction of DCX+ cells at the SVZ but a moderate but significant reduction of SGZ DCX+ neuroblasts, without changes in Ki67+ or TUNEL+ cells. ConclusionsThis study reports a novel role of the ChP/CSF in maintaining the neuroblast pool in the neurogenic niches in both early postnatal and adult stages. Moreover, we expand the available tools to target the ChP and CSF production in the neonate, with potential uses in treating conditions such as neonatal hydrocephalus.

Intracerebral haemorrhage (ICH) is a severe form of stroke with high morbidity and mortality rates. For survivors, acute haematoma expansion strongly determines neurological outcome. Although blood pressure reduction is widely investigated as a strategy to limit haematoma growth, the haemodynamic mechanisms regulating haemorrhage development remain poorly understood. Zebrafish provide a tractable in vivo model to study cerebrovascular biology and spontaneous ICH, yet the contribution of vascular regulation to haemorrhage onset and expansion has not been explored in this species. Here, we investigated whether pharmacological modulation of vascular dilation influences ICH development in zebrafish larvae. We first characterised vascular changes during the developmental window in which spontaneous ICH occurs and observed increased heart rate and progressive reductions in arterial diameter between 2 and 3 days post-fertilisation, suggesting increased vascular resistance. We then tested whether vasoconstriction promotes haemorrhage using angiotensin II, which induced systemic and cerebrovascular vasoconstriction but did not increase ICH incidence or haematoma size in two independent ICH models. In contrast, pharmacological vasodilation using sodium nitroprusside or isoproterenol significantly reduced haematoma size in a high-incidence model of atorvastatin-induced ICH. Live imaging of cerebral blood flow revealed that vasodilation was associated with the confinement of red blood cells around affected vessels rather than dispersing into the brain ventricles. Together, these findings indicate that vascular dilation modulates haemorrhage progression in zebrafish ICH and establish this model as a platform to investigate haemodynamic mechanisms regulating haematoma expansion.

We have established and optimized a protocol for the high-yield isolation of primary epithelial cells from rat choroid plexus. The addition of cytosine arabinoside suppressed the growth of contaminating cells, and epithelial culture was grown into a confluent impermeable monolayer within 5-6 days after seeding. To form an in vitro blood-CSF barrier, epithelial cells were plated on inverted coated polycarbonate support of Transwell inserts. Morphologically, the polarized cells remained cuboidal in shape and expressed TJ proteins at a high rate. The filter-grown monolayers displayed transendothelial resistance (TEER) values in the range of 160 to 180 {Omega} x cm2 and remained at this level for 3 days, indicating the persistent formation of continuous TJs. The cells were able to secrete cerebrospinal fluid (CSF) actively. Epithelial cells showed expression of selective influx and efflux transporters. To conclude, our BCSFB model exhibits tight, functional barrier characteristics and shows the functional expression of the pharmaceutically important influx/efflux transporters. The recent model is suitable for in vitro investigations of BCSFB and routine pre-clinical drug discovery.

Understanding intracranial pressure (ICP) dynamics is essential for interpreting clinical infusion tests used in the diagnosis of cerebrospinal fluid circulation disorders. However, the complex coupling between vascular pulsations, cerebrospinal fluid flow, and intracranial compliance makes quantitative interpretation of these tests challenging. Here, I present a patient specific simulation framework based on an extended electrical analog model that reproduces intracranial pressure dynamics observed during clinical infusion tests. The model integrates physiological inputs including arterial blood pressure, heart rate, respiratory rhythm, and resistance to cerebrospinal fluid outflow derived from clinical data. Built upon the classical Ursino framework, the model incorporates several modifications enabling realistic representation of physiological pulsations and infusion test conditions. The resulting system functions as a hybrid electrical-numerical simulation model representing a simplified digital electrical twin of intracranial hydrodynamics. The model was validated using data from 21 clinical infusion tests performed in patients with suspected normal pressure hydrocephalus. Simulated intracranial pressure recordings were compared with clinical measurements using regression and residual analysis. The simulations demonstrated strong agreement with measured data, with a mean correlation coefficient of r = 0.95 (95% CI 0.94 - 0.96), mean residual values within -1.71 to +1.68 mmHg, and a mean root mean square error (RMSE) of 2.07 mmHg. These results demonstrate that the proposed model accurately reproduces the dynamic behavior of intracranial pressure observed during clinical infusion tests. The framework provides a physiologically grounded computational tool for studying patient specific intracranial dynamics and may support improved interpretation of infusion test results in clinical practice.

This study investigates the impact of rapamycin and propranolol on cerebral cavernous malformations (CCMs). Employing an unbiased transcriptomic analysis, we aimed to comprehensively elucidate the molecular mechanisms underlying these drug effects in Mouse Brain Microvascular Endothelial Cells (mBMEC) deficient in Ccm3. While propranolol shows limited efficacy in modulating the CCM transcriptomic phenotype in mBMEC, rapamycin demonstrates a higher impact. Rapamycin reverses gene expression changes induced by Ccm3 deficiency, restoring Klf2/4-dependent genes like Nos3, Adamts1, and Thbs1. Notably, we observed a reduction in KLF2 protein levels in Ccm3 KO cells treated with rapamycin. Critically, in vivo experiments demonstrate that a combination of rapamycin and lapatinib effectively reduces lesion volume in a chronic CCM model. This finding is particularly noteworthy as it suggests a potential treatment strategy for existing lesions. In summary, our work describes a new mechanism for the effects of rapamycin in Ccm3- deficient cells and identifies a new drug combination in the treatment of cavernomas.

BackgroundThis study examines whether quantifiable changes can be detected in ventricular volume in Idiopathic Normal Pressure Hydrocephalus (iNPH) patients that undergo ventriculo-peritoneal shunt procedures. There is no known metric that characterizes the change in ventricular volume for iNPH patients after shunt placement. MethodsTwo de-identified and independent datasets are studied: O_LI45 brain CT scans (24 diagnosed with iNPH and 21 normal elderly individuals) are used to evaluate the effectiveness of our proposed ventricular volume metric as a diagnostic tool for iNPH. The performance of our deep learning model-based metric is compared to the traditional Evans Index using ROC analysis. C_LIO_LI16 subjects with a total of 50 longitudinal CT scans taken before and after shunt surgery across different imaging centers are studied to quantify the impact of shunt treatment. Clinical symptoms of gait, balance, cognition, and bladder continence are studied with respect to the proposed metric. C_LI ResultsOur proposed metric achieves high accuracy (0.95), precision (0.96), and recall (0.96) in distinguishing between normal and iNPH subjects, surpassing the performance of the Evans Index. This metric allows us to track changes in ventricular volume before and after shunt surgery for 16 subjects. Notably, the 15 subjects with iNPH demonstrate a decrease in ventricular volume post-surgery and a concurrent clinical improvement in their iNPH symptomatology. ConclusionOur novel metric accurately quantifies changes in ventricular volume before and after shunt surgery for iNPH patients, serving as an effective radiographic marker for a functioning shunt in a patient with iNPH. O_LIWhat is already known on this topic - The diagnosis of iNPH involves both clinical and radiographic stigmata. Radiologists rely largely on visual examination of CT scans and provide qualitative evaluations about ventricular volume. C_LIO_LIWhat this study adds - Our study provides quantitative information about the patency and function of the shunt. C_LIO_LIHow this study might affect research, practice, or policy - The validated deep learning-based metric enhances iNPH diagnosis accuracy by tracking radiographic biomarkers. This facilitates decision-making regarding the efficacy of shunt surgery and the effect on brain compliance. We provide a web interface to apply the metric, its reliable performance across multiple institutional scanner types could be adapted to the real-time clinical evaluation of iNPH and improve treatment workflows. C_LI

Sensory perception and motor dexterity is coordinated by in part distinct anatomical centres in the spinal cord. Importantly the spinal cord is the first modulatory relay hub for coordinating sensory and motor inputs to allow control of an organisms response to a sensory experience and to orientate proprioceptive outputs. This is whilst communicating with higher centres within the brain to undertake greater complex neurophysiological function such as pain perception. This begins to outline the complexity of the nervous system communication. To allow this integral system to function efficiently neuronal homeostasis needs to be maintained with energy expenditure matched by proficient delivery of nutrients. This factor introduces the vascular system that extensively interacts in a multifaceted manner with differing aspects of the nervous system. Part of this multi-factoral interaction is through the heterogenic cellular makeup of the vascular network that delivers and modulates the molecular transport of such nutrients to spinal cord tissues, but also controlling penetration and migration of harmful pathogens and agents. Therefore the spinal cord is susceptible to any alterations in the microvessel integrity (e.g. vascular leakage) and/or function (e.g. cessated blood flow) of this vascular network, which principally occurs in times of pathology. Typically investigations into microvessel function have utilised histological and/or tracer based in-vivo assays. Methodologies such as evans blue extravasation have been used inconjunction with in-vitro cell biology assays such as transwell assays to determine microvessel integrity or function that only provides snapshots of developing vasculopathy. Adopting in-vivo imaging approaches, allow for real time functional measurements of the ongoing physiological function within the spinal cord, providing direct measurement of the vascular processes in play, including vascular architecture, blood flow and/or permeability. This technique in mouse allow for direct visualisation of cellular and/or mechanistic influence upon vascular function through utilising disease, transgenic and/or viral approaches. This combination of attributes allows for in depth real time understanding of the function of the vascular network within the spinal cord.

The pathways that run along the olfactory nerves crossing the cribriform plate and connecting to lymphatic vessels in the nasal cavity, have been identified as a crucial route for cerebrospinal fluid (CSF) outflow. However, the presence of a CSF efflux pathway through blood vessels in this region has yet to be clarified. This study aimed to elucidate the anatomical connections between the subarachnoid space and the bloodstream at the nasal epithelium and the venous drainage routes of the nasal epithelium in mice. Our findings demonstrated that CSF tracers could be drained not only through lymphatic vessels in the nasal cavity and cervical lymph nodes (CLNs), but also through the blood vessels in this area that extend to its venous drainage routes, including the facial and jugular veins. Additionally, we showed that ligation of CLNs neither impeded the influx and efflux of CSF tracers nor exacerbated Alzheimers disease (AD)-related pathology in AD mice. Our work reveals a previously unrecognized pathway for CSF drainage through blood vessels within the nasal mucosa. These findings provide insight into the efficient removal of waste products, facilitating optimal functioning of neural tissue within the susceptible tissue of our brains.

Ventriculoperitoneal (VP) shunts are effective at relieving hydrocephalus but are prone to malfunction. There are two hypotheses as to how shunts may malfunction independently of mechanical failure or blockage by debris from initial placement. The first is that the presence of a foreign object results in cells migrating into and colonising the shunt. The second is that the shunts contain either small numbers of live bacteria or residual bacterial products from manufacture or handling, triggering an inflammatory response that attracts cells to the site which go on to cause malfunctions. The presence of bacteria can be difficult to definitively rule in or out, given that they are capable of forming biofilms which poses challenges for isolation and microbiological culture. In this study, we measured 91 soluble immunological molecules and 91 soluble neurological molecules in CSF of patients with VP shunts and compared them to both patients without shunts and those with bacterial infection to determine whether there is an ongoing inflammatory response to shunting. We find that shunts elicit a soluble signature of neural wound healing and cell migration proteins that is distinct from the inflammatory signature of patients with neurological infection. This appears to represent a long-term response, persisting for at least 5 years in one patient.

Evidence strongly suggests that SARS-CoV-2, the cause of COVID-19, can enter the brain. SARS-CoV-2 enters cells via the S1 subunit of its spike protein, and S1 can be used as a proxy for the uptake patterns and mechanisms used by the whole virus; unlike studies based on productive infection, viral proteins can be used to precisely determine pharmacokinetics and biodistribution. Here, we found that radioiodinated S1 (I-S1) readily crossed the murine blood-brain barrier (BBB). I-S1 from two commercial sources crossed the BBB with unidirectional influx constants of 0.287 {+/-} 0.024 L/g-min and 0.294 {+/-} 0.032 L/g-min and was also taken up by lung, spleen, kidney, and liver. I-S1 was uniformly taken up by all regions of the brain and inflammation induced by lipopolysaccharide reduced uptake in the hippocampus and olfactory bulb. I-S1 crossed the BBB completely to enter the parenchymal brain space, with smaller amounts retained by brain endothelial cells and the luminal surface. Studies on the mechanisms of transport indicated that I-S1 crosses the BBB by the mechanism of adsorptive transcytosis and that the murine ACE2 receptor is involved in brain and lung uptake, but not that by kidney, liver, or spleen. I-S1 entered brain after intranasal administration at about 1/10th the amount found after intravenous administration and about 0.66% of the intranasal dose entered blood. ApoE isoform or sex did not affect whole brain uptake, but had variable effects on olfactory bulb, liver, spleen, and kidney uptakes. In summary, I-S1 readily crosses the murine BBB, entering all brain regions and the peripheral tissues studied, likely by the mechanism of adsorptive transcytosis. Graphical Abstract O_FIG_DISPLAY_L [Figure 1] M_FIG_DISPLAY C_FIG_DISPLAY

The blood-brain barrier (BBB) is a multifactorial and multicellular vascular interface separating the systemic environment from the central nervous system (CNS). It gates cerebral penetration of circulating molecules and cells and is the principal reason for low accumulation of many therapeutics in the brain. Low dose methamphetamine (METH) induces fluid phase transcytosis across the BBB in vitro and could therefore be used to enhance CNS drug delivery. Here we show, that low dose intravascular METH induced significant leakage exclusively via caveolar transport at the intact BBB in rodents ex vivo. Notably, METH-induced leakage was suppressed at 4{degrees}C and in Caveolin-1 (CAV1) knockout mice. Furthermore, METH strongly enhanced brain penetration of therapeutic molecules, namely doxorubicin (DOX), a small chemotherapeutic agent, and aflibercept (AFL), a ca. 100 kDA recombinant protein. Lastly, METH improved the therapeutic efficacy of DOX in a mouse model of human glioblastoma (GBM), as measured by a 25% increase in median survival time (p = 0.0024). Collectively, our data indicated that METH can facilitate preclinical assessment of novel experimental treatments and has the potential to enhance drug delivery to the diseased CNS.

Astrocyte endfeet form a near-continuous sheath around the brains vasculature, defining the perivascular spaces (PVS) that are crucial for brain fluid flow and solute transport. Yet, their precise physiological role remains poorly understood. Using 3D electron microscopy data, we created a high-fidelity poroelastic computational model of an arteriole segment with surrounding endfeet and parenchyma to investigate tissue displacement and fluid flow within the PVS, endfeet, and extracellular space (ECS) in response to blood vessel pulsations. Our model predicts that arteriole dilations compress the PVS while expanding the overall endfoot sheath volume due to tangential stretch. Moreover, fluid exchange primarily occurs through inter-endfoot gaps, driven by pressure differences, rather than across the aquaporin-4 (AQP4) rich endfoot membrane. PVS stiffness critically modulates these dynamics: increased stiffness of the PVS, for instance, due to vessel pathology or aging, would minimize or even reverse fluid exchange at the gliovascular interface. While AQP4 mediated water movement has a negligible impact on pulsation-driven mechanics, it significantly enhances osmotically driven fluid flow. Overall, our findings elucidate the complex balance of forces governing gliovascular mechanics and suggest that PVS composition strongly influences endfoot-parenchymal fluid exchange. SignificancePerivascular spaces, formed by astrocyte endfeet wrapping the vasculature, are high-conduit pathways for brain fluid flow and clearance. Vascular pulsations drive this flow, but the resulting mechanical interactions at the gliovascular interface remain largely unknown. We introduce a computational model of the solid and fluid mechanics here, using realistic geometries to capture intricate astrocyte morphology at the subcellular level. Our simulations reveal that changes in perivascular composition - associated with aging or neurodegenerative diseases - fundamentally alter mechanical coupling, potentially impeding fluid transport. This work provides a mechanistic framework for understanding brain clearance and constitutes a foundational model for computational studies of mechanical forces in the nervous system.

Objective: Shunt-dependent hydrocephalus is a common and costly complication of aneurysmal subarachnoid hemorrhage (aSAH), affecting up to 28% of survivors. Existing prediction tools, including the Chronic Hydrocephalus Ensuing from SAH Score (CHESS), have limited discriminative accuracy. We developed the CHECKMATE score, a clinically practical tool to improve prediction of ventriculoperitoneal shunt dependency after aSAH. Methods: In this multicenter retrospective cohort of 486 patients with aSAH from Mayo Clinic (January 1, 2006-December 31, 2021), we used multivariable logistic regression and machine learning to identify independent predictors of ventriculoperitoneal shunt placement. The CHECKMATE score was derived from 5 weighted variables: symptomatic hydrocephalus (10 points), intraventricular hemorrhage (5 points), SAH volume greater than 10 mL (3 points), neutrophil-to-lymphocyte ratio greater than 12 (2 points), and 10-year incremental age thresholds starting at older than 60 years (1 point each). Results: Of 486 patients (mean age, 56.3 years; 64.6% female), 137 (28.2%) required ventriculoperitoneal shunt placement. The CHECKMATE score achieved an area under the curve of 0.808 (compared to 0.737 for CHESS), with a sensitivity of 0.85, specificity of 0.67, and negative predictive value of 0.92 at the optimal cutoff of 14 points. Conclusions: The CHECKMATE score outperforms CHESS for predicting ventriculoperitoneal shunt dependency after aSAH and is easily used at the bedside. Its high negative predictive value helps identify low-risk patients who may benefit from earlier external ventricular drain weaning and shorter hospital stays.