Microvasculature as a Key Regulator of Health and Disease in the Brain and Beyond

Manipulating the peripheral nervous system to affect microvascular perfusion and metabolic health

Diabetes and the associated hyper- and hypo-glycemia damage blood vessels in the brain, worsening cognitive function and increasing the risk of developing dementia. Type 2 diabetes mellitus is characterized by insulin resistance. The “resistance” to insulin is largely a result of poor insulin delivery to the target cells with blood flow being the primary regulator of insulin mediated glucose update. Blood flow to skeletal muscle is tightly regulated by both the vasoconstrictive, sympathetic and vasodilatory, parasympathetic nervous systems.

We are taking advantage of the interorgan communication facilitated by the peripheral nervous system to manipulate blood flow to improve glucose homeostasis.

The baroreceptor reflex is defined as the sensing of pressure in the aortic arch and compensatory changes in vascular tone and heart rate to maintain blood pressure. Using a mouse with improved cardiac contractility, we activated the baroreceptor reflex, to induce vasodilation and improve glucose homeostasis in diet-induced obese mice.

Fatty liver is associated with hypertension and insulin resistance. We previously established that the fatty liver produces GABA, which inhibits activity of the vagal nerve resulting in hyperinsulinemia and insulin resistance. Inhibiting liver GABA production improves insulin sensitivity, prevents obesity and angiotensin II induced hypertension, and increases vasodilatory cGMP in skeletal muscle. GABA signaling onto the hepatic vagal nerve increases peroneal nerve activity and sympathetic tone at skeletal muscle. This established the key role of hepatic vagal afferent signaling in regulating insulin and glucose delivery to skeletal muscle.

Together our studies establish the potential to target the afferent peripheral nervous system signaling to modulate microvascular blood flow, systemic blood pressure, and metabolic health.

Mind over MASLD: Unravelling brain dysfunction in steatotic liver disease

Metabolic dysfunction–associated steatotic liver disease (MASLD) is a chronic multisystem disease affecting approximately 30% of the general population and more than 80% of individuals with obesity. It frequently coexists with cardiovascular disease, diabetes, and cancer, contributing to a growing global burden of multimorbidity. While hepatic encephalopathy is classically associated with advanced liver disease, accumulating evidence indicates that neurocognitive impairment and accelerated brain ageing also occur in earlier stages of MASLD. However, the mechanisms driving these cerebral alterations, and whether they are reversible following disease resolution, remain largely unknown.

In this talk, data will be presented from male C57BL/6NTac mice fed a control diet (CD) or a high-fat, high-cholesterol diet (HFD) for 26 weeks to induce MASLD. To assess the impact of ageing and dietary intervention, a subset of HFD-fed mice was subsequently switched to a low-fat diet (LFD) and followed until 18 months of age. All animal procedures were carried out in accordance with the UK Animals (Scientific Procedures) Act 1986 and its associated guidelines, and complied with the ARRIVE guidelines. The study was conducted under a UK Home Office Project Licence and approved by the local Animal Welfare and Ethical Review Body (AWERB). All efforts were made to minimise animal suffering and to reduce the number of animals used.

Behavioural analyses revealed anxiety-like behaviour in both HFD and aged HFD-LFD mice, while memory impairment was evident in aged CD-LFD and HFD-LFD groups.Measurements of brain oxygenation showed reduced partial pressure of oxygen and tissue oxygen saturation in HFD, aged HFD-LFD, and CD-LFD mice at baseline, assessed under anaesthesia using fluorescence-based methods and optoacoustic tomography. Cerebrovascular reactivity to systemic hypercapnia (10% CO₂) was preserved. Structural analyses demonstrated altered vascular complexity and reduced pericyte coverage in HFD and aged HFD-LFD mice compared to controls. These changes were accompanied by increased cortical microglial density, coverage, and volume, consistent with a reactive inflammatory phenotype.

Together, these findings identify altered brain oxygenation and inflammation as potential contributors to MASLD-associated brain dysfunction. Importantly, dietary fat reduction in later life did not reverse MASLD or its associated cerebral alterations, suggesting that brain vulnerability may persist or even be exacerbated with ageing. Ongoing studies in mice of both sexes are currently exploring alternative dietary strategies and mechanisms acting along the gut-liver–brain axis. This work aims to define the long-term impact of MASLD on brain health, even after liver disease improvement, and to understand how metabolic liver disease interacts with the natural ageing process.

Cross-sectional Associations Between Sleep Quality and Glucose Regulation in Older Adults

Introduction

Sleep quality, glucose regulation, and age-related metabolic changes converge on physiological pathways linked to cognitive decline across the lifespan. More than 500 million adults worldwide meet criteria for impaired glucose regulation, highlighting the scale of metabolic vulnerability in aging populations¹. Aging is also associated with altered sleep architecture, increased fragmentation, and higher apnea burden, all of which may contribute to metabolic and neurovascular risk². Yet how sleep architecture (macro and microarchitecture) and sleep-disordered breathing relate to glucose regulation in aging, and whether these relationships differ by sex, remains incompletely understood.

Aims/Objectives

We investigated the relationship between traditional sleep architecture, sleep depth (Odds Ratio Product; ORP), and glucose regulation in community-dwelling middle-aged to older adults. We further examined whether these associations differed by sex.

Methods

The study conformed to the standards set by the Declaration of Helsinki. Eighty-six adults (n=56 women; age 63.8±5.9 years) from the Brain in Motion II study completed overnight in-home Level II polysomnography and fasting blood sampling. Glucose status was classified as normoglycemic (n=45; ≤5.6 mmol/L) or glucose-dysregulated (n=40; >5.6 mmol/L). Groups were comparable for age and sex.

Results
Individuals with glucose dysregulation had higher body mass index (30.8±4.9 vs. 27.3±5.1 kg/m²; p=0.002), higher body fat percentage (38.1±7.1 vs. 35.1±7.2%; p<0.001), and lower V̇O₂max (24.9±5.3 vs. 27.9±4.7 mL/kg/min; p=0.007). Glucose-dysregulated participants displayed higher sleep efficiency (76.5±10.1 vs. 70.2±10.4%; p=0.002), shorter sleep-onset latency (25.3±36.2 vs. 42.6±45.6 min; p=0.037), and less full wakefulness (ORP decile 10: 8.9±6.4 vs. 13.7±7.1% of total sleep time; p=0.002). Total sleep time (346.0±65.0 vs. 327.6±72.8 min; p=0.278) and wake after sleep onset (137.1±58.3 vs. 125.4±51.7 min; p=0.241), did not differ significantly between groups. No group differences were observed in other ORP deciles or NREM stages.

Sex-based analyses showed that females had higher sleep efficiency (75.0±10.4 vs. 69.8±10.9%; p=0.012), less time awake (115.4±52.4 vs. 141.0±60.5 min; p=0.018), and greater NREM stage 3 sleep (55.1±40.1 vs. 17.7±26.6 min; p<0.001) compared with males. Sex-by-glucose interactions indicated that differences in sleep efficiency and wakefulness by glucose status were evident in females but not males.

Overall, apnea–hypopnea index (AHI) did not differ by glucose group or sex; however, glucose-dysregulated females exhibited higher AHI than normoglycemic females (26.1±16.8 vs. 15.7±11.9 events·h⁻¹, p=0.046). When categorized by OSA severity, glucose-dysregulated participants showed a greater burden of moderate-to-severe OSA, whereas normoglycemic participants were more likely to fall within no or mild OSA categories. Hypoxia-related indices were also worse in glucose-dysregulated participants, particularly among females.

Conclusion

Our findings suggest that the relationship between glucose regulation and sleep in older adults is more nuanced than expected. Glucose-dysregulated participants exhibited patterns consistent with increased homeostatic sleep pressure, including higher sleep efficiency and reduced wakefulness, potentially reflecting early physiological adaptations preceding overt sleep disruption. The paradoxically “better” sleep outcomes may be influenced by higher BMI and a greater burden of undiagnosed mild-to-moderate OSA, with chronic sleep fragmentation, REM disruption, and nocturnal hypoxia contributing to compensatory sleep drive. This underscores the importance of considering sleep-disordered breathing when evaluating sleep–glucose interactions and may inform early preventive strategies targeting both metabolic and sleep pathways to support healthy aging.

Acute Intestinal Inflammation Alters Cerebral Hemodynamic Stability and Functional Connectivity in Mice

Introduction

Inflammatory bowel disease (IBD), primarily Crohn’s disease and ulcerative colitis, is an immune-mediated inflammatory disorder of the gastrointestinal tract. Beyond intestinal pathology, individuals with IBD frequently experience extra-intestinal manifestations and have an increased risk of cerebrovascular events, potentially driven by systemic inflammation and prothrombotic mechanisms. Using a dextran sulphate sodium (DSS)-induced colitis model, we investigated whether acute intestinal inflammation is associated with systemic inflammatory changes and alterations in cerebral haemodynamics and functional connectivity assessed using functional ultrasound (fUS).

Methods

CD1 mice aged 4–8 weeks were allocated to acute DSS-induced colitis (5% w/v, 40 kDa DSS in drinking water for 3 days; n=20, 10 female) or control conditions (standard drinking water; n=20, 10 female). Disease activity index (DAI) was assessed daily using body weight change, stool consistency, and faecal occult blood testing. Following the 3-day exposure, mice underwent 15 minutes of resting-state fUS imaging (Iconeus One) under isoflurane anaesthesia. Blood was collected for plasma, and mice were perfusion-fixed with paraformaldehyde.

Faecal calprotectin and serum inflammatory/coagulation-related markers (IL-6, von Willebrand factor antigen [VWF:Ag], functional VWF activity [VWF:CBA], and ADAMTS13) were quantified using ELISA.

fUS was used to map cerebral blood volume (CBV) and functional connectivity across the brain. Twenty regions of interest (ROIs) were manually defined with reference to the Allen Mouse Brain Atlas. Baseline CBV was defined as the mean of the first 40 seconds; relative change (ΔCBV) was computed as (CBV − baseline)/baseline. ROI-wise ΔCBV time courses were extracted. Functional connectivity assessment was estimated from low-frequency (<0.1 Hz) fluctuations following detrending and confound removal. Group comparisons used t-tests, with family-wise error rate control via permutation testing (10,000 permutations); p<0.05 was considered significant. All procedures complied with the UK Animals (Scientific Procedures) Act 1986 and were approved by the University of Manchester ethical review board.

Results

DSS exposure induced clear intestinal and systemic inflammatory changes. DAI scores increased from day 1 and remained elevated through day 3 in the DSS group relative to controls (Figure 1a). Faecal calprotectin levels were significantly higher in DSS-treated mice, alongside increased serum IL-6 (Figure 1b-c). In contrast, coagulation-related markers (VWF:Ag, VWF:CBA, and ADAMTS13) did not differ significantly between groups (Figure 1d-h).

Resting-state fUS revealed a consistent but modest increase in mean ΔCBV in DSS-treated mice; however, no ROI survived permutation-based FWER correction. Notably, temporal association, auditory, and ectorhinal cortices showed significantly greater temporal variability of ΔCBV (higher standard deviation over time), suggesting increased haemodynamic fluctuation (Figure 2). ROI-to-ROI connectivity analyses further identified significantly increased functional connectivity between auditory and ectorhinal regions in the DSS group. (Figure 3)

Conclusion

Acute DSS-induced colitis produced robust intestinal inflammation accompanied by systemic cytokine elevation and measurable alterations in resting-state cerebral haemodynamic dynamics. Although mean ΔCBV differences did not survive stringent multiple-comparison correction, increased temporal variability and enhanced connectivity between auditory and ectorhinal regions suggest that peripheral inflammation may modulate neurovascular dynamics. Further mechanistic work is warranted to clarify inflammation–brain coupling in IBD and its potential relevance to cerebrovascular vulnerability.

Does systemic microvascular function mediate the association between large artery stiffness and cognitive function in older adults?

INTRODUCTION: Previous research identified a link between increased large artery stiffness and adverse brain health, supporting the concept that the penetration of excess pulsatility into cerebral circulation damages its small vessels and, when persisted, leads to cognitive decline. Although assessing cerebral microvasculature non-invasively at bedside poses a major challenge, systemic microvascular function such as that of skin can be utilised to investigate the link. We determined whether large artery stiffness and systemic microvascular function were associated with cognitive function in older adults, and also determined whether systemic microvascular function mediated an association between large artery stiffness and cognitive function. METHODS: Older adults with diverse cardiovascular risk (n=435, 162 females) participated in this study. Estimated pulse wave velocity (ePWV) as an indirect estimate of large artery stiffness was calculated from age and mean arterial pressure as described elsewhere.¹ Systemic microvascular function was assessed by responses to pharmacological stimulation of forearm skin blood vessels using a solid-state laser Doppler imager (LDI2, Moor Instruments, Axminster, UK) as previously described.² Endothelium-dependent (acetylcholine: ACh) and -independent (sodium nitroprusside: SNP) vasodilators were delivered trans-dermally via iontophoresis, and forearm skin perfusion was measured as the area under the response curve of each vasodilator (ACh_auc and SNP_auc, respectively) normalised to resting perfusion. Validated cognitive tasks [Addenbrooke’s Cognitive Examination Revised (ACE-R), Trail Making Test Part-A (TMT-A) and Part-B (TMT-B)] were administered to assess participants’ global cognitive function, speed of processing and executive function, respectively. A higher total score for ACE-R and a shorter completion time for TMT-A and TMT-B indicate better functions. All procedures accorded with current UK legislation. RESULTS: One standard deviation (SD) increase in ePWV was positively associated with TMT-A [β=0.339 (0.226, 0.467), p<0.001] and TMT-B [β=0.342 (0.232, 0.451), p<0.001] after accounting for conventional cardiovascular risk factors. Additionally, one SD increase in ACh_auc was inversely associated with TMT-B after adjusting for conventional cardiovascular risk factors [β=-0.113 (-0.201, -0.024), p=0.013]. A mediation analysis revealed that the association between ePWV and TMT-B was partially mediated by ACh_auc explaining about 3% of the total effect in the unadjusted model (p<0.05), but this did not persist in the adjusted model. CONCLUSIONS: Large artery stiffness and systemic microvascular function were associated with cognitive function in older adults, but these vascular measures may independently contribute to cognitive function without systemic microvascular function mediating the association between large artery stiffness and cognitive function in our cohort.

Effects of ageing, obesogenic diet, and the flavonoid Fisetin on small artery function in mice

In humans, ageing and metabolic stress involving a sedentary lifestyle and excess caloric intake are significant risk factors for developing cardiovascular, cerebrovascular, and neurodegenerative diseases. The pathophysiology of these diseases involves hypertension and/or dysregulation of blood flow in critical organs. Small (resistance) arteries (<300 µm) regulate tissue blood flow and systemic vascular resistance. Endothelial dysfunction, hypercontractility, deficient autoregulation, structural remodeling, and arterial stiffening are hallmarks of vascular dysfunction. The flavonoid fisetin has known cardioprotective activities, but it is unknown whether it can alleviate vascular dysfunction in ageing and metabolic stress. The primary objectives of this study were to investigate small artery dysfunction in young vs. old mice with or without diet-induced metabolic stress, and to test whether Fisetin can reverse vascular dysfunction. A secondary objective was to test if Rho-kinase is involved in age- or diet-induced vascular dysfunction.

Animal treatments and procedures were approved by the relevant authorities. Wildtype C57BL/6NTac male mice were divided into 8 groups evenly distributed to intermittent Fisetin or Vehicle treatment (4 weeks; 20 mg/kg 2 days/week). Young mice (3.5 months at study end) were fed either normal diet (N=16) or a high-fat diet (60% saturated fat) with fructose (10%) in drinking water (N=16) for 10 weeks. Old mice (retired breeders; 17 months at study end) were fed either normal chow (N=15) or a high-fat/high-fructose diet (N=15) for 10 weeks. At study end, mice were humanely sacrificed, and 3^rd order mesenteric arteries (150-200 µm) were quickly dissected free from fat and connective tissue. Using a pressure myograph system, arteries were cannulated under physiological pressure (60 mm Hg) and bathed in Krebs buffer (37°C; pH 7.40). We measured the following vascular responses: vasoconstriction to high-KCl (75 mM K⁺) and noradrenaline (NA, 1 µM); dilation to the endothelium-dependent vasodilator acetylcholine (ACh, 10 µM); myogenic tone (MT; pressure range 60-120 mm Hg); structural remodeling (passive lumen diameter, wall cross-sectional area, wall/lumen-ratio); and vascular stiffness (Young’s Modulus). We also tested the effects of Rho-kinase inhibition (Ripasudil, 400 nM) on MT. We performed factor analysis with age and diet as factors (plus age x diet interaction) in vehicle- vs. Fisetin-treated mice, using two- or three-way Anova with multiple comparisons. A P-value <0.05 was considered significant.

Ageing was associated with increased body weight (P<0.0001) and NA contractility (P<0.05), reduced endothelial function (P<0.05), and hypertrophic remodeling (P<0.05; P<0.01). Obesogenic diet was associated with increased body weight (P<0.0001), reduced endothelial function (P<0.05), reduced MT (P<0.001), and structural remodeling (P<0.05; P<0.01). An age x diet interaction was observed for MT (P<0.001) and vascular stiffness (P<0.001). Fisetin treatment was associated with a reduced age-dependent difference in NA contractility, reduced endothelial dysfunction, reduced MT in young mice, and reduced structural remodeling. Rho-kinase inhibition reduced MT in the young vehicle-treated mice (both diets) and reduced MT in all Fisetin-treated mice (young/old, both diets). This is consistent with a loss of Rho-kinase activity in ageing,¹ which was partially restored by Fisetin treatment.

Overall, Fisetin treatment may have beneficial effects on aspects of the function of small arteries governing blood pressure and -flow.

Using microfluidic models to explore vascular responses in CSVD and stroke

Introduction

Stroke and cerebral small vessel disease (CSVD) are major causes of death and disability worldwide. Cerebral Small Vessel Disease is a group of conditions that affects the stability of cerebral micro-vessels, manifesting in a variety of clinical syndromes, including vascular cognitive impairment, dementia, and stroke. CSVD accounts for a quarter of the 12 million global stroke cases, and ~45% of the 35–42 million new cases of dementia each year (Bos et al, 2018). Despite the world-wide importance of CSVD and stroke there are no current treatments beyond risk factor modification and restoration of blood flow. A major barrier to developing new therapies is a lack of relevant human in vitro models that capture the form and function of brain blood vessels. Our lab is developing new models of both stroke and CSVD that enable the study of cerebral blood vessel function and dysfunction.

Aims

To develop a suit of microfluidic cell culture models that replicate blood vessel dysfunction in CSVD and stroke.

Methods

We have used soft-lithography micro-fabrication techniques to produce microfluidic cell culture devices that can pattern a cell laden hydrogel and stimulate vasculogenesis to create a capillary network mimetic. To model stroke, human primary brain endothelial cells, pericytes and astrocytes were co-cultured and allowed to form vessel like structures before being subjected to oxygen glucose deprivation. Vessel integrity was measured using fluorescent tracer dyes. For CSVD models, induced pluripotent stem cells were derived from patients with monogenic forms of CSVD and differentiated to endothelial cells, pericytes and astrocytes for co-culture in microfluidic devices. Barrier integrity of the resulting in vitro vessels was measured using fluorescent tracers and compared to isogenic disease corrected controls.

Results

Both stroke mimicking conditions and CSVD causing mutations disrupted barrier function of in vitro brain capillary networks.

Conclusions

Microfluidic cell culture approaches can be used to create brain capillary mimetics that recapitulate barrier disruption relating to stroke and CSVD.

Menstrual endocrine fluctuations increase dynamic cerebrovascular functions

Introduction

Recent scientific effort has started to delineate how ovarian hormones, which regularly fluctuate during the menstrual cycle, influence neural and cerebrovascular tissue. This may underlie the relationships between later hormonal decline and neurodegenerative disease risk¹. To fully profile how hormones interact with cerebrovascular function, it is vital to consider, not just resting physiology, but also dynamic aspects of cerebrovasculature that support neural activity.

This study uses hypercapnic cerebrovascular reactivity (CVR) and visually-evoked haemodynamic response function (HRF) to investigate the influence of menstrual-related changes in oestradiol and progesterone on dynamic aspects of the cerebrovascular system.

Methods

21 menstruating females (age mean[SD]= 22.9[3.91] years) completed a 3T MRI (Siemens MAGNETOM Prisma) scanning session during the early follicular (EFP; day 1-4; N=17), late follicular (LFP; day 10-12; N=17), and mid-luteal (MLP; day 20-22; N=18) phase of their menstrual cycle (14 completed all three). All took place at the same time of day, after fasting for 4-6 hours. Circulating hormones were measured via blood samples. All procedures were compliant with ethical standards.

Simultaneous blood oxygen level dependant (BOLD)-CVR and cerebral blood flow (CBF)-CVR data were collected using a pseudocontinuous arterial spin labelling (pCASL) acquisition during periods of hypercapnia (5% CO₂) with PETCO₂ monitoring (maximum repetition time [TR]=3600ms, echo time [TE] 1=10ms, TE2=30ms, in-plane resolution=3.4×3.4, slice thickness=6.5mm, GRAPPA acceleration factor=3). The HRF was estimated using a whole brain EPI scan during 100% contrast checkerboard presentation (TR=2s, TE=30ms, resolution=2mm³, multi-band acceleration factor=4).

The HRF timeseries was fit with a gamma variate model. Three metrics were extracted– peak amplitude (PA), time-to-peak (TTP), and full-width-half-max (FWHM). PETCO₂ traces were used in modelling CVR maps. BOLD-CVR (∆%BOLD/mmHgCO₂) and CBF-CVR values (∆ml/100g/min/mmHgCO₂) were extracted across the whole brain, while HRF metrics were extracted from visually-responsive regions. Mixed linear models investigated the contribution of hormones to vascular outcomes across phases. Progesterone was regressed against oestradiol to allow investigation of independent variation (hereby called resProgesterone).

Results

Oestradiol accounted for a significant amount of global CVR variance, both for CBF-CVR (χ²(1)= 50.92; p<0.001; average variation attributed to oestradiol across a cycle in this sample=1.01∆ml/100g/min/mmHgCO₂) and for BOLD-CVR (χ²(1)=59.14; p<0.001; average variation attributed to oestradiol across a cycle in this sample=0.1∆%BOLD/mmHgCO₂). ResProgesterone however, was only significantly associated with global CBF-CVR (χ²(1)=26.71; p<0.001; average variation attributed to resProgesterone=0.6∆ml/100g/min/mmHgCO₂).

For HRF, increasing oestradiol was associated with slightly higher PA (χ²(1)=20.55; p<0.001; average variation attributed to oestradiol across a cycle in this sample=0.1%BOLD) and earlier TTP (χ²(1)=10.036; p=0.002; average variation attributed to oestradiol across a cycle=0.4 seconds). Increasing resProgesterone was associated with narrower FWHM (χ²(1)=28.71; p<0.001; average variation attributed to resProgesterone=0.5 seconds). No significant regional effects were found.

Conclusions

This study found evidence that dynamic cerebrovascular functions are associated with menstrual-related ovarian hormones. These effects were small but notable within the context of the regular menstrual cycle and raise questions about larger hormonal variations, such as within pregnancy or menopause. This may be a potential mechanism underlying menstrual symptomatology and has implications for fMRI studies that assume intact neurovascular coupling processes in women, regardless of menstrual staging, to make inferences about neural activity.

Astrocyte-to-pericyte signalling underlies serotonin-mediated cerebral blood flow regulation

Cerebral blood flow (CBF) is tightly coupled to neuronal activity. Excitatory glutamatergic activity induces local vessel dilation, while monoamines such as noradrenaline and serotonin (5-HT) stemming from brainstem neurons induce vessel constriction to maintain vascular tone. Despite its well-described role as a vasoconstrictor, the exact mechanism through which 5-HT affects blood flow remains largely understudied. Here, we show that serotonergic nerve terminals are in direct contact with capillary pericytes and astrocytic endfeet. Applying 5-HT to brain slices and in vivo induces potent pericyte constriction by activating the 5HT_2C receptor, which is expressed in neurons and astrocytes. Exogenous 5-HT application induces a significant increase in the frequency, duration and amplitude of calcium transients in astrocytes and pericytes. Importantly, blocking production of 20-HETE, a potent arachidonic acid derivative that is released by astrocytes to constrict vessels, abolished the 5-HT-mediated pericyte constriction in brain slices, suggesting that 5-HT modulates pericyte contraction through astrocytes. Altogether, these data suggest that in homeostasis, 5-HT exerts its vasoactive effects on CBF by regulating astrocyte-to-pericyte communication.

In the context of Alzheimer’s disease (AD), the association between serotonergic axons, pericytes and astrocytes is altered, owing to the development of markedly enlarged neuronal processes, termed axonal spheroids (AxS), that are found near Aβ plaques. AxS have been primarily studied in excitatory neurons and their development in monoaminergic and cholinergic axons remains largely unexplored. Here, we show that monoaminergic and cholinergic AxS have striking anatomical differences to glutamatergic AxS, with far greater axon swelling and distinct molecular constituents. We classify amyloid plaques based on the type of affected axons in their vicinity and explore its association with plaque compaction. Relevantly, using in vivo two-photon microscopy in anaesthetized animals expressing GRAB_5-HT sensors, we show that some cortical plaques are linked to dynamic fluctuations in local 5-HT levels, possibly reflecting altered neurotransmitter release or diffusion through AxS. These findings suggest that the peri-plaque environment could be shaped by the affected axon type and raise the possibility that acetylcholinesterase inhibitors may exert their therapeutic effect by restoring peri-plaque neurotransmitter balance. We are currently investigating how the presence of cholinergic and monoaminergic AxS might influence local inflammatory responses and blood flow.

Inflammation before and after the bleed: A dual role for cholesterol in intracerebral haemorrhage?

Intracerebral haemorrhage (ICH) is a severe form of stroke caused by spontaneous blood vessel rupture and bleeding into the brain tissue, resulting in significant disability and death. Our research, using in vitro systems, post-mortem brain tissue, zebrafish, and mouse models, has identified Cholesterol 25-hydroxylase (Ch25h) as a critical molecule influencing both the risk of ICH and the brain’s response to injury after a bleed.

Systemic infections and inflammation can compromise neurovascular integrity, increasing the risk of blood vessel rupture and ICH in some people (1). While infection-induced hypertension and endothelial dysfunction are known contributors, hypocholesterolemia—low levels of LDL cholesterol—is a less understood but clinically relevant risk factor (2). Cells respond to infection by temporarily reducing cholesterol availability, which can impair the formation and function of adherens junctions and may destabilise the neuroendothelium (3, 4). We hypothesise that this infection-driven cholesterol dysregulation can, under certain conditions, lead to neurovascular instability and ICH.

After ICH occurs, the influx of blood into the brain triggers widespread inflammation and cellular damage. Reactive oxygen species, iron from haemoglobin breakdown, and cellular debris activate microglia and recruit peripheral leukocytes, amplifying injury. Cholesterol metabolism plays a dual role in this context: disrupted lipid handling can intensify inflammation, while efficient cholesterol clearance supports tissue repair. Cholesterol is also essential for membrane synthesis and remyelination, making its regulation a delicate balance between injury and recovery.

Ch25h is an enzyme that converts cholesterol into 25-hydroxycholesterol (25HC), an oxysterol involved in immune regulation. We found that antiviral signalling increases Ch25h expression in zebrafish and human foetal brain tissue, correlating with ICH risk. In brain endothelial cell cultures and zebrafish models, Ch25h and 25HC disrupt vascular integrity by remodelling cholesterol metabolism (5). This suggests a mechanistic link between infection, cholesterol dysregulation, and neurovascular dysfunction. As well as its role in ICH risk, Ch25h also influences recovery after brain haemorrhage. In both mouse and human brain, Ch25h is strongly upregulated in microglia and macrophages after ICH. Loss of Ch25h in mice worsens outcomes, while treatment with 25HC enhances blood clearance and improves acute recovery. We also show 25HC regulates lipid droplet accumulation in macrophages following phagocytosis of red blood cells, indicating a broader role in lipid handling during haematoma clearance.

In summary, Ch25h and its product 25HC are central to both the onset and resolution of ICH. They mediate neurovascular dysfunction during infection and regulate immune and lipid responses during brain injury and recovery. Targeting this pathway may offer new preventative and therapeutic strategies to reduce ICH risk and improve outcomes, respectively.

Early Flow Dysregulation and Rarefaction in an Angiotensin II Infusion-Recovery Model

Loss of microvascular blood flow is a critical feature of hypoperfusion across neurovascular diseases, particularly hypertension-associated vascular dementia. However, the early mechanisms underlying capillary flow dysregulation (pre-rarefaction), stalling (functional rarefaction), and eventual microvessel loss (anatomical rarefaction), remain poorly defined and difficult to model. Here, we examine these microvascular pathologies in an Angiotensin II (AngII) infusion-recovery model in which mice receive a pressor dose of AngII (800 ng/kg/min) or vehicle saline for 28 days followed by a 14-day recovery period.

Blood pressure and heart rate were monitored by tail cuff plethysmography and microvascular flow was quantified at two-week intervals through an acute cranial window using two-photon line scan imaging and Radon transform analysis. Flow velocity, temporal variability and stall frequency were assessed in the first four capillary branch orders adjoining precapillary arterioles. Brains were subsequently collected for immunohistochemical analyses of endothelial cells, pericytes and tissue hypoxia.

AngII induced severe hypertension by week two (131±4 mmHg AngII vs. 100±3 mmHg saline; N=8 vs. N=8; p=0.0002) which persisted through week four (136±3 mmHg AngII vs. 111±3 mmHg saline; N=8 vs. N=8; p=0.0002) and normalized by week six (105±4 mmHg AngII vs. 101±3 mmHg saline; N=8 vs. N=8; p=0.8706). Capillary flow velocity slowed considerably by week four (0.79±0.07 mm/s AngII vs. 1.13±0.09 mm/s saline; N=4 vs. N=5; p=0.0125) and remained reduced at week six (0.87±0.06 mm/s AngII vs. 1.18±0.10 mm/s saline; N=6 vs. N=6; p=0.0389) despite normalized blood pressure. Flow variability, measured as temporal coefficient of variation (CV) excluding stalls, was unchanged during infusion but increased at recovery week six (AngII CV=0.28±0.02, saline CV=0.18±0.01; N=6 vs. N=6; p=0.0021). Stalling rose progressively, peaking at recovery week six (AngII 24.2±2.6% vs. saline 9.9±1.9%; N=6 vs. N=6; p<0.0001).

These functional disturbances were accompanied by anatomical rarefaction. Microvessel loss was pronounced in the hippocampus of six-week AngII recovery mice (5.9±0.4% AngII vs. 8.2±0.4% saline CD31+ vessel density; N=6 vs. N=6; p=0.0005) with concurrent pericyte loss (2.1±0.3% AngII vs. 3.8±0.2% AngII PDGFRb+ pericyte density; N=6 vs. N=6; p=0.0002). This coincided with substantial tissue hypoxia (17.0±0.8% AngII vs. 0.9±0.1% saline HypoxyProbe+ tissue area; N=6 vs. N=6; p<0.0001). By contrast, the cortex displayed a trend towards microvessel loss (9.2±0.3% AngII vs. 10.0±0.4% saline CD31+ vessel density; N=6 vs. N=6; p=0.2528), significant pericyte loss (2.6±0.3% AngII vs. 4.5±0.3% saline PDGFRb+ pericyte density; N=6 vs. N=6; p<0.0001) and less pronounced hypoxia than the hippocampus (4.1±0.8% AngII vs. 0.7±0.1% saline HypoxyProbe+ tissue area; N=6 vs. N=6; p=0.0009).

Together, these findings show that four-week AngII infusion initiates early microvascular rarefaction, producing persistent microvessel loss even after a two-week return to normotension. The hippocampus exhibits the greatest anatomical vulnerability, whilst the cortex shows marked flow dysregulation and stalling consistent with early-stage rarefaction. These early disturbances likely precede broader neurological consequences observed in extended AngII models. This preliminary data outlines the early steps linking hypertension to hypoperfusion, regional hypoxia, and cognitive decline, laying the groundwork for future efforts to define the mechanisms driving microvascular rarefaction in vascular dementia.

Amyloid-β driven neuronal hyperexcitability creates energy imbalance in an Alzheimer’s mouse model, particularly affecting the hippocampus

Introduction

Active neurons dilate nearby blood vessels, ensuring sufficient oxygen availability, fuelling this activity. Neurovascular coupling (NVC) mechanisms facilitating this functional hyperaemia may be disrupted early in Alzheimer’s Disease (AD) pathogenesis through direct actions of Beta-Amyloid (Aβ) on neurovasculature and/or induction of chronic neuronal hyperexcitability, impairing the ability of healthy NVC to be sustained. The hippocampus is affected early in AD which may be partly driven by its lower vascularisation and NVC response compared to cortex, making it particularly vulnerable to further NVC disruption. APOE4, the strongest risk allele for sporadic AD, may also confer compromised NVC in carriers, and is an integral link between systemic cardiovascular health and AD.

Aims

In the current study we aim to unpick contributions of Aβ accumulation and APOE genotype to early-AD NVC impairment, comparing the hippocampus to the visual cortex.

Methods

To do this we have employed a novel mouse model combining humanised APOE (3/3 or 4/4) and doxycycline-inducible APPSwe/Ind transgenes, as well as the CaMKII driven GCaMP6f neuronal calcium indicator. By measuring, in vivo, brain blood flow and oxygenation, neuronal activity, and vascular dilatory responses, before and after induction of Aβ expression we aim to untangle the relative impact of low level Aβ on different aspects of neurovascular physiology, and to understand how these changes could feed into gradual neurodegeneration.

Statistical Analyses

Data were analysed using linear mixed models to determine the effects of genotype, time off doxycycline and brain region, with animal ID specified as the random factor (n=7-18 mice per condition).

Ethics

All experimental procedures were approved by the UK Home Office, in accordance with the 1986 Animal (Scientific Procedures) Act, and the University of Sussex animal welfare ethical review board.

Results

Preliminary data suggests that soluble but not aggregated Aβ increases in both cortex hippocampus over 3 months, causing cortical neurons to become hyperexcitable. Over the same timescale, cortical blood flow increases from baseline, possibly to counter this hyperexcitability, but decreases in the hippocampus. However, blood oxygenation decreases in both the cortex and hippocampus, suggesting any increase in blood flow is insufficient to counter increased oxygen consumption by neurons.

Conclusions & Future Directions

Ongoing analyses will interrogate influences of APOE genotype, characterise any effect on hippocampal neurons and cortical and hippocampal NVC.

Redox regulation of neurovascular coupling in health and disease

The functional integrity of the brain depends on a tight coupling between neuronal activity and local cerebral blood flow, a process known as neurovascular coupling (NVC). This mechanism ensures the timely delivery of oxygen and metabolic substrates to active neuronal circuits. Nitric oxide (NO), produced through the glutamate–NMDAr–nNOS–sGC pathway, is a key mediator of communication between neurons and the cerebral vasculature. Under physiological conditions, tightly controlled redox signaling preserves NO bioavailability and supports efficient NVC. Conversely, disruption of redox homeostasis reduces NO bioavailability which is linked to NVC impairment and cognitive decline in different pathological conditions (e.g. Alzheimer’s disease, vascular cognitive impairment and type 2 diabetes). Understanding how redox mechanisms regulate NO signaling within the neurovascular unit is therefore critical for identifying strategies capable of preserving cerebrovascular function and cognitive health. Among these, dietary inorganic nitrate—abundant in vegetables such as beetroot and green leafy vegetables—has emerged as a promising approach to enhance NO bioavailability.

To explore the interplay between redox regulation, NO signaling, neurovascular coupling and cognition, we investigated rodent models of vascular cognitive impairment, including chronic cerebral hypoperfusion (2VO) and type 2 diabetes. Using a multimodal approach combining behavioral testing, in vivo assessment of NVC and cerebral blood flow dynamics, magnetic resonance imaging, and molecular analyses of vascular remodeling and oxidative stress pathways, we demonstrate that these models exhibit marked neurovascular dysfunction accompanied by oxidative stress–driven redox imbalance and impaired spatial learning and memory. Importantly, boosting NO bioavailability through dietary nitrate was able to improve NVC responses, improved cerebral blood flow regulation, attenuated pathological vascular remodeling, and improved cognitive performance. Together, these findings highlight the central role of redox-dependent regulation of NO signaling in maintaining neurovascular coupling in health and its disruption in disease.

Characterising the Effect of NLRP3 Inflammasome-Induced Vascular Neuroinflammation on Blood Brain Barrier Integrity

Various studies have implicated neuroinflammatory NLRP3 inflammasome activation in a host of neurodegenerative diseases with neurovascular aspects, though the interaction between the inflammasome and the blood-brain barrier has yet to be deciphered. This project aims to assess the effect of NLRP3 inflammasome activation on the integrity of the neural endothelium, independent of cell death, and investigate the underlying pathways. Furthermore, this project aimed to elucidate the electrophysiological effects of endothelial TLR4 activation by LPS, with potassium efflux being a key regulator of NLRP3 inflammasome activation.

hCMEC/D3 cells were cultured in EGM-2 Basal Medium and SingleQuots supplements 5% foetal bovine serum, 0.4% human fibroblast growth factor-B, 0.04% hydrocortisone and 0.1% ascorbic acid along with 1% Pen-Strep. LPS treatments were conducted over 6 hours, or 5 hours + hour 10µM nigericin with or without an hour of 20µM MCC950 treatment. Permeability assays were conducted on endothelial monolayers cultured on 0.4µm Transwell membranes, followed by ion permeability assessment by a Voltohmeter, and 2000kDa Dextran-Fluorescein flow determined by ID3 Spectramax plate reader. Protein for western blotting was obtained by scraping with RIPA buffer and membranes were read using a LICOR ODESSEY. Immunofluorescence was imaged using a Zeiss AxioObserver7 confocal microscope and analysed by ImageJ.

Whole-cell currents were recorded using a MultiClamp 700B amplifier for data acquisition (n=6-10 cells per condition) and analysis was performed with Clampex 11.2 (Molecular Devices) software. Whole-cell currents were recorded in HEPES buffered standard solution at a perfusion rate of 4ml/min and ∼35ºC.

All graphed data is presented as mean ± SEM. Statistical analysis was performed using GraphPad Prism 10 software and used one- or two-way ANOVA followed by Tukey’s multiple comparisons, or unpaired t-tests. Significance was set at *p < 0.05.

Following treatment, Dextran permeability increased by 5.89×10^-12±1.513×10^-12mol/cm²×S (p=0.0078) and TEER decreased by 42.68±7.944% (p=0.0003) in comparison to the healthy controls. MCC950 prior to nigericin addition showed a 36.97±10.26 % increase (p=0.013) in ion permeability compared with the LPS + nigericin treated groups (n=3-7 independent experiments). Western blotting confirmed inflammasome activation and immunofluorescence showed inflammasome localisation, a shift in morphology and a reduction of tight and adherens junction localisation to the cell membranes in NLRP3 inflammasome activated cells (3 replicates). Interestingly, LPS treatment reduced whole-cell currents at 50mV by -100.2±31.37pA (p=0.0085, N=6-7) compared with untreated cells and V ¹/² K⁺ by 10.99±4.302mV (p=0.0268, N=6-7).

These findings imply that NLRP3 activation is disruptive to the BBB by altering the morphology and junction protein expression of microvasculature endothelial cells, though further research is needed to decipher the underlying pathways involved. The effect of NLRP3 inflammasome inhibitor MCC950 in reversing some of these effects could show some future therapeutic potential for rescuing inflammatory-associated vascular dysfunction in AD in conjunction with anti-amyloid treatments. The electrophysiology studies show a novel effect of TLR4 activation that may influence K⁺ efflux for inflammasome activation.

Acute Microvascular Effects of Cilostazol and Isosorbide Mononitrate Reveal Regional Redistribution of Cerebral Blood Supply in Wild-Type Mice

Background:

White matter (WM) is preferentially damaged in cerebral small vessel disease (cSVD), likely due to its susceptibility to chronic hypoperfusion (Wardlaw et al., 2013). Cilostazol (CIL) and isosorbide mononitrate (ISMN) have been shown in clinical LACI trials to improve memory and reduce stroke risk (Wardlaw et al., 2023); however, the acute cerebrovascular mechanisms and the vascular compartments at which these drugs act remain unclear. We investigated whether CIL and ISMN induce age- and region-dependent acute changes in microvascular haemodynamics.

Methods:

Wild-type C57BL/6J mice of both sexes underwent cranial window implantation at approximately 12 weeks of age over WM or cortical grey matter (GM). For WM windows, ~800 µm of cortex was aspirated over the corpus callosum and a custom 3D-printed window with a glass base was implanted. GM windows were placed over the visual cortex without aspiration, preserving pial arteries. Mice were first recorded at 4 months of age; the same WM mice were re-recorded after 8 months of ageing (1 year old).

Using a blinded crossover design, mice received vehicle or combined CIL+ISMN with ≥6-day washout. Haemodynamics were recorded at baseline, 30 min, 1 h, 2 h, 4 h, 6 h, and 24 h using combined laser Doppler flowmetry and haemoglobin spectroscopy. Measures included red blood cell (RBC) flux, RBC speed, concentration of moving RBCs, oxygen saturation (sO₂), total haemoglobin (HbT), and derived cerebral metabolic rate of oxygen (CMRO₂). Awake, head-fixed recordings were obtained during resting periods. Linear mixed-effects models were used with drug and timepoint as fixed effects and animal ID as a random effect, with Dunnett post-hoc tests. Drug doses were scaled from the LACI-2 trial using body-surface-area conversion.

Results:

In deep tissue containing WM of young mice (n = 8), CIL+ISMN significantly reduced RBC flux (treatment effect p = 0.022; timepoint × treatment p = 0.032), driven by a marked reduction at 6 h (estimate −0.22 ± 0.06, p = 0.0003). RBC concentration was also reduced (p = 0.0072), while RBC speed was unchanged. CMRO₂ decreased significantly (p = 0.016), most prominently at 6 h (p = 0.0007), without significant changes in sO₂.

In the same mice re-examined longitudinally after 8 months of ageing (n = 4), WM responses to CIL+ISMN were preserved, with significant reductions in RBC flux (p = 0.013), RBC concentration (p = 0.041), and CMRO₂ (p = 0.007).

In contrast, GM showed a significant increase in moving RBC concentration (p = 0.003), a modest reduction in HbT (p = 0.044), and a trend toward increased sO₂ (p = 0.082), with no change in flux or speed. Blood pressure under sedation did not differ between baseline, vehicle, or drug conditions.

Conclusions:

Our data indicate region-specific microvascular responses to cilostazol and isosorbide mononitrate, with preserved cortical grey matter microcirculation and reduced red blood cell delivery and oxygen consumption in deep tissue containing white matter. These findings support an acute redistribution of microvascular resources rather than a uniform microvascular response. Ongoing two-photon imaging and chronic hypoperfusion models will determine the vascular origin and pathological relevance of this redistribution under chronic treatment conditions.

BACE1: A novel regulator of the microcirculation from head to toe

Metabolic disease (Type 2 diabetes and obesity), neurodegeneration and auto-immune diseases are all characterised by chronic inflammation. We are interested in the vascular consequences of inflammation by understanding the molecular mechanisms involved. Vascular inflammation is a complex immune response occurring in blood vessels, characterised by endothelial dysfunction, activation of immune cells and perivascular adipose tissue and the release of proinflammatory chemokines and cytokines. Damaged blood vessels increased risk of developing cardiovascular diseases which is a precursor to vascular complications including atherosclerosis, heat attack and stroke.

The protease β-site APP-cleaving enzyme 1 (BACE1) is associated with Alzheimer’s disease development, via the production of β-amyloid peptides. However, our work using proteomics and RNAseq analysis has shown increased BACE1 activity drives endothelial dysfunction. Furthermore, we have identified genetic associations with hyperlipidaemia and hypertension. Here we explore the hypothesis that BACE1 could be a novel drug target for vascular disease.

Results: BACE1 is highly expressed in endothelial and smooth muscle cells in human atherosclerotic lesions. With elevated expression occurring in the early phases of lesion development.

Four-week treatment of a BACE1 inhibitor (Merck-3) in Western diet fed ApoE KO mice reduced atherosclerotic plaque load by 50% both in the whole aorta and aortic root (P<0.05). Furthermore, BACE1 inhibition lowered vascular inflammation (P< 0.01), serum triglyceride (P< 0.01) and LDL cholesterol (P< 0.05) levels and reduced hypertension (P< 0.05) compared to vehicle controls. Mechanistically, BACE1 inhibition restores endothelial cells and vascular function.

Conclusions: Taken together our data demonstrate that elevated BACE1 activity is a driving factor of atherosclerotic lesion development via induction of endothelial dysfunction. Thus, targeting this with available BACE1 inhibitors could be a novel therapy for atherosclerosis and vascular disease. These findings have the potential to have wide ranging impact as the interrelation of blood vessels, inflammation and immunity is a fundamental physiological process while also underlying several diseases, ranging from infections to autoimmune diseases, metabolic syndrome, dementia.

Systemic inhibition of nitric oxide synthesis alters vascular tone, vasomotion and brain oxygen availability and reduces network-level coordination of cerebrovascular physiology

Nitric oxide (NO) is a critical regulator of vascular tone and cerebral blood flow, and reduced NO bioavailability is implicated in both cardiovascular and neurodegenerative disease. Although inhibition of nitric oxide synthase (NOS) is known to alter cerebral haemodynamics and vasomotion, its effects are typically assessed using individual physiological measures in isolation. Here, we applied a multivariate systems-level approach to characterise how NOS inhibition reshapes coordinated cerebrovascular and systemic physiology.

Multiple indices of systemic and cerebrovascular function were recorded simultaneously in awake mice under control conditions and following administration of the non-specific NOS inhibitor L-NG-nitroarginine methyl ester (L-NAME; 75 mg kg⁻¹, s.c.). Cerebral blood flow, blood oxygenation, total haemoglobin and derived oxygen metabolism (CMRO₂) were measured in visual cortex and hippocampus alongside peripheral heart rate, respiratory rate, arterial oxygen saturation, pulse distension and blood pressure. Vasomotion and neurovascular coupling were quantified from these signals.

L-NAME reduced heart and respiratory rates and increased blood pressure, with minimal effects on peripheral oxygen saturation. In contrast, cerebral blood flow, oxygenation and CMRO₂ were markedly reduced in both regions, while blood volume was relatively preserved. Vasomotion power and frequency increased substantially, particularly in the hippocampus, whereas microvascular pulsatility increased similarly across regions and neurovascular coupling remained largely intact. Multivariate analyses combining principal component analysis with mixed-effects modelling revealed dissociable physiological axes reflecting direct effects of NOS inhibition and secondary effects mediated by reduced cerebral oxygenation. Notably, changes in vasomotion were more strongly associated with reductions in cerebral oxygenation than with drug condition itself, indicating that some effects commonly attributed to loss of NO signalling may instead arise from secondary hypoxia. These findings suggest that NOS inhibition produces complex cerebrovascular changes that cannot be assumed to reflect direct NO-dependent mechanisms alone.

Hypercapnia, a potential biomarker for vascular dementia, elicits increased cortical cerebral blood flow through connexin channels.

Introduction:

The human brain produces a staggering 75 litres of CO₂/day. We recently published data demonstrating CO₂ mediates neurovascular coupling, affecting cerebral blood flow (CBF) (Hosford et al., 2022). A meta-analysis published by our group suggested that nitric oxide and cyclooxygenase (COX) are key modulators of this response (Hosford et al., 2022). However, the mechanisms behind this phenomenon and the implications for cerebrovascular disease remain incompletely understood.

Aims

In this study, we aim to characterise the molecular mechanism behind hypercapnia induced vasodilation of the cerebrovasculature. We also aim to characterise this effect on cerebrovascular reactivity in the hyperhomocysteinemia (HHcy) mouse model of vascular dementia.

Methods

Vessel diameter changes in cortical arterioles were recorded from ex vivo rat brain slices, pre-constricted with L-phenylephrine (200µM) in artificial cerebral-spinal-fluid (aCSF). Isohydric hypercapnia (pH 7.35) was induced by bubbling aCSF with 13.5% CO₂ containing 80mM sodium bicarbonate. Ex vivo and in vivo dye loading experiments were carried out on brain slices and open cranial windows on 10-week-old male C57BL6 mice using aCSF containing 200µM carboxyfluorescein. Hypercapnia in in vivo studies was delivered by free-breathing of 10% CO₂. The diet-induced HHcy model of vascular dementia, known to develop cognitive impairment and cerebral microbleeds (Sudduth et al., 2013), was carried out on C57BL6 mice of both sexes (n=20). CBF and cerebrovascular reactivity to hypercapnia were quantified using arterial-spin-labelling MRI (9.4T Bruker) under medetomidine/isoflurane anaesthesia.

Results

Recording of cortical arteriolar diameter changes in these slices confirmed CO₂induced vasodilation independent of CO₂induced acidification (p<0.05, n=9, t-test). Vasodilation was prevented by COX-1 inhibition (p<0.001, n=6, one-way-ANOVA) and was affected by neuronal nitric oxide synthase (NOS) inhibition (p<0.001, n=6, one-way-ANOVA), but not endothelial NOS inhibition (p>0.05, n=6, one-way-ANOVA) or neuronal activity inhibition (TTX) (p>0.05, n=6, one-way-ANOVA). Additionally, we found that hypercapnia-induced dilation depends on purinergic signalling (n=6) and is modulated by connexin/pannexin channel availability (p<0.01, n=6, one-way-ANOVA). We confirmed the role of these channels through loading of carboxyfluorescein, a connexin/pannexin channel permeable dye. Concentration dependent CO₂ ex vivo dye loading in brain slices was blocked by connexin channel inhibitors. This was confirmed in vivo in the mouse somatosensory cortex, using hypercapnia (p<0.01, n=5, one-way-ANOVA), and CO₂ release induced by forepaw stimulation (p<0.01, n=5, one-way-ANOVA).

In the HHcy model of vascular dementia, no differences in basal CBF or cognitive impairment were detected after 10 weeks on the HHcy inducing diet. However, cerebrovascular reactivity to hypercapnia was significantly reduced in mice on the HHcy diet compared to those on the control diet (p<0.05, n=20, t-test). This is suggestive of early disrupted cerebrovascular reactivity, a translational biomarker which could precede cognitive deficits and changes to basal perfusion.

Conclusion

In conclusion, our studies indicate that hypercapnia-induced cerebrovascular dilation is dependent upon neuronal nitric oxide release, purinergic signalling, and CO₂induced connexin channel availability. Further studies are underway to understand the mechanisms driving the impairment of cerebrovascular reactivity to hypercapnia in the HHcy mouse model, highlighting its potential to be used as a much-needed early biomarker of vascular dementia.

Advanced MRI shows subtype-specific vascular characteristics in Gliomas

INTRODUCTION
Gliomas generate an additional blood supply to meet their increased need for nutrients and oxygen, altering normal physiology and tissue metabolism^1,2. As proliferating cells demand more blood flow, the structure and function of the brain’s vasculature are disrupted. Vascular proliferation is a pathological hallmark of gliomas, leading to abnormal and dysfunctional vessels³. Considerable heterogeneity exists within tumours regarding vascular morphology. For example, high microvessel density is characteristic of astrocytic gliomas, and high-grade tumours exhibit pronounced neovascularization⁴.

The formation of new blood vessels to support tumour growth has clinical consequences, most notably the development of vasogenic edema, this can lead to an increase in intracranial pressure due to leakage of the blood–brain barrier⁵.

AIM
This study used advanced MRI techniques to characterize vascular differences across glioma subtypes.

METHODS
The study was approved by the NHS Research Ethics Committee, Wales, UK and Cardiff University’s Ethics Committee, School of Psychology, Cardiff, UK.

Quantitative susceptibility maps (QSM) were acquired from 27 patients (mean age 45.5 ± 8.8 years; 20 male) diagnosed with glioma. Vascular parameters—including vessel density, vessel diameter, and oxygen extraction fraction (OEF)—were extracted from the QSM maps.
Pre- and post-contrast T1-weighted images (gadolinium-enhanced), T2-weighted images, and T2-FLAIR were used to segment brain tissue into tumour core (enhancing vs. non-enhancing), edema, necrotic area, cavity, and residual healthy grey matter. (Figure 1)

ANOVAs were conducted to evaluate differences in OEF (within skeletonized vessels), vessel diameter, and vascular density in edema, tumour core, and residual healthy grey matter. Gender was included as a covariate.

RESULTS
Structurally, we observed increased vessel diameter in edema and tumour core regions compared with residual healthy grey matter (F_(2,38)= 11.08; p < 0.01) (Figure 2-A). Vascular density was higher in enhancing tumour tissue compared with non-enhancing tumour regions and healthy grey matter (F_(2,38)= 16.81; p < 0.01). (Figure 2-B)

Physiologically, OEF was reduced in non-enhancing tumour regions compared with healthy grey matter and enhancing tumour tissue. This result may reflect infiltration or hypoxia, with altered OEF and modest vascular changes consistent with metabolic stress or tissue loss (F_(2,38)= 7.16; p < 0.01) (Figure 2-C).

Cerebral blood flow (CBF) was reduced in both enhancing and non-enhancing tumour areas compared with healthy grey matter, indicating vascular–metabolic decoupling in enhancing cores likely related to abnormal vessel structure and shunting (F_(2,38)= 22.07; p < 0.01) (Figure 2-D).

CONCLUSION
Our findings demonstrate the potential of QSM to identify distinct metabolic and vascular characteristics across glioma subtypes, consistent with evidence that more aggressive tumours develop their own blood supply, leading to altered vascular parameters.

Integrating QSM into routine glioma assessment may enable non-invasive hypoxia evaluation, differentiation from non-tumour abnormalities, and support individualized treatment planning. This approach could help guide surgical and radiotherapy strategies and facilitate longitudinal monitoring of treatment response.

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Thursday 16 April - Friday 17 April 2026

Microvasculature as a Key Regulator of Health and Disease in the Brain and Beyond

Date

Location

09:45 SA01 Manipulating the peripheral nervous system to affect microvascular perfusion and metabolic health

10:15 SA02 Mind over MASLD: Unravelling brain dysfunction in steatotic liver disease

10:45 C01 Cross-sectional associations between sleep quality and glucose regulation in older adults

11:00 C02 Acute intestinal inflammation alters cerebral hemodynamic stability and functional connectivity in mice

11:45 Cardiovascular system around the clock: The role of circadian rhythms and redox signalling in ageing and β-adrenergic-induced cardiac remodelling

12:15 C03 Does systemic microvascular function mediate the association between large artery stiffness and cognitive function in older adults?

12:30 C04 Effects of ageing, obesogenic diet, and the flavonoid fisetin on small artery function in mice

13:45 Role of incretin hormones in regulation of cerebrovascular circulation

14:15 SA03 Using microfluidic models to explore vascular responses in CSVD and stroke

14:45 C05 Menstrual endocrine fluctuations increase dynamic cerebrovascular functions

15:00 C06 Astrocyte-to-pericyte signalling underlies serotonin-mediated cerebral blood flow regulation

15:45 SA04 Inflammation before and after the bleed: A dual role for cholesterol in intracerebral haemorrhage?

16:15 C07 Early flow dysregulation and rarefaction in an angiotensin II infusion-recovery model

16:30 C08 Amyloid-β driven neuronal hyperexcitability creates energy imbalance in an Alzheimer’s mouse model, particularly affecting the hippocampus

09:30 Microglial tunneling nanotubes as helplines during neurodegeneration

10:00 SA05 Redox regulation of neurovascular coupling in health and disease

10:30 C10 How does a mild decrease in brain oxygen supply constrain hippocampal function?

C14 Astrocyte enrichment of 3D cortical constructs enhances brain repair following traumatic brain injury

11:45 SA06 Cerebrovascular dynamics in dementia: Exploring dysfunctional brain cell interactions

12:15 C11 Characterising the effect of NLRP3 inflammasome-induced vascular neuroinflammation on blood brain barrier integrity

12:30 C12 Acute microvascular effects of cilostazol and isosorbide mononitrate reveal regional redistribution of cerebral blood supply in wild-type mice

14:00 Endothelial Nrf2 signalling under physiological oxygen levels: Implications for improved clinical translation

14:30 SA07 BACE1: A novel regulator of the microcirculation from head to toe

15:00 C09 Systemic inhibition of nitric oxide synthesis alters vascular tone, vasomotion and brain oxygen availability and reduces network-level coordination of cerebrovascular physiology

15:15 C14 Targeting the blood-brain barrier: New frontiers in neurodegeneration and brain health

16:00 SA08 Blood flow changes and myelin damage as early mechanisms leading the dementia

16:30 C15 Hypercapnia, a potential biomarker for vascular dementia, elicits increased cortical cerebral blood flow through connexin channels

16:45 C16 Advanced MRI shows subtype-specific vascular characteristics in gliomas

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