Current Understanding of the Anatomy, Physiology, and Magnetic Resonance Imaging of Neurofluids: Update From the 2022 "ISMRM Imaging Neurofluids Study group" Workshop in Rome.

Neurofluids is a term introduced to define all fluids in the brain and spine such as blood, cerebrospinal fluid, and interstitial fluid. Neuroscientists in the past millennium have steadily identified the several different fluid environments in the brain and spine that interact in a synchronized harmonious manner to assure a healthy microenvironment required for optimal neuroglial function. Neuroanatomists and biochemists have provided an incredible wealth of evidence revealing the anatomy of perivascular spaces, meninges and glia and their role in drainage of neuronal waste products. Human studies have been limited due to the restricted availability of noninvasive imaging modalities that can provide a high spatiotemporal depiction of the brain neurofluids. Therefore, animal studies have been key in advancing our knowledge of the temporal and spatial dynamics of fluids, for example, by injecting tracers with different molecular weights. Such studies have sparked interest to identify possible disruptions to neurofluids dynamics in human diseases such as small vessel disease, cerebral amyloid angiopathy, and dementia. However, key differences between rodent and human physiology should be considered when extrapolating these findings to understand the human brain. An increasing armamentarium of noninvasive MRI techniques is being built to identify markers of altered drainage pathways. During the three-day workshop organized by the International Society of Magnetic Resonance in Medicine that was held in Rome in September 2022, several of these concepts were discussed by a distinguished international faculty to lay the basis of what is known and where we still lack evidence. We envision that in the next decade, MRI will allow imaging of the physiology of neurofluid dynamics and drainage pathways in the human brain to identify true pathological processes underlying disease and to discover new avenues for early diagnoses and treatments including drug delivery. Evidence level: 1 Technical Efficacy: Stage 3.

ApoE4 Astrocytes Secrete Basement Membranes Rich in Fibronectin and Poor in Laminin Compared to ApoE3 Astrocytes.

The accumulation of amyloid-ß (Aß) in the walls of capillaries and arteries as cerebral amyloid angiopathy (CAA) is part of the small vessel disease spectrum, related to a failure of elimination of Aß from the brain. Aß is eliminated along basement membranes in walls of cerebral capillaries and arteries (Intramural Peri-Arterial Drainage-IPAD), a pathway that fails with age and ApolipoproteinEε4 (ApoE4) genotype. IPAD is along basement membranes formed by capillary endothelial cells and surrounding astrocytes. Here, we examine (1) the composition of basement membranes synthesised by ApoE4 astrocytes; (2) structural differences between ApoE4 and ApoE3 astrocytes, and (3) how flow of Aß affects Apo3/4 astrocytes. Using cultured astrocytes expressing ApoE3 or ApoE4, immunofluorescence, confocal, correlative light and electron microscopy (CLEM), and a millifluidic flow system, we show that ApoE4 astrocytes synthesise more fibronectin, possess smaller processes, and become rarefied when Aß flows over them, as compared to ApoE3 astrocytes. Our results suggest that basement membranes synthesised by ApoE4 astrocytes favour the aggregation of Aß, its reduced clearance via IPAD, thus promoting cerebral amyloid angiopathy.

Vascular basement membrane alterations and ß-amyloid accumulations in an animal model of cerebral small vessel disease.

Non-amyloid cerebral small vessel disease (CSVD) and cerebral amyloid angiopathy (CAA) may be interrelated through the damaged basement membranes (BMs) and extracellular matrix changes of small vessels, resulting in a failure of ß-amyloid (Aß) transport and degradation. We analyzed BM changes and the pattern of deposition of Aß in the walls of blood vessels in spontaneously hypertensive stroke-prone rats (SHRSP), a non-transgenic CSVD model. In 45 SHRSP and 38 Wistar rats aged 18 to 32 weeks: (i) the percentage area immunostained for vascular collagen IV and laminin was quantified; (ii) the capillary BM thickness as well as endothelial and pericyte pathological changes were analysed using transmission electron microscopy (TEM); and (iii) the presence of vascular Aß was assessed. Compared with controls, SHRSP exhibited a significantly higher percentage area immunostained with collagen IV in the striatum and thalamus. SHRSP also revealed an age-dependent increase of the capillary BM thickness and of endothelial vacuoles (caveolae) within subcortical regions. Endogenous Aß deposits in the walls of small blood vessels were observed in the cortex (with the highest incidence found within fronto-parietal areas), striatum, thalamus and hippocampus. Vascular ß-amyloid accumulations were frequently detected at sites of small vessel wall damage. Our data demonstrate changes in the expression of collagen IV and of the ultrastructure of BMs in the small vessels of SHRSP. Alterations are accompanied by vascular deposits of endogenous Aß. Impaired ß-amyloid clearance along perivascular and endothelial pathways and failure of extracellular Aß degradation may be the key mechanisms connecting non-amyloid CSVD and CAA.

Inhibition of Aquaporin-4 Improves the Outcome of Ischaemic Stroke and Modulates Brain Paravascular Drainage Pathways.

Aquaporin-4 (AQP4) is the most abundant water channel in the brain, and its inhibition before inducing focal ischemia, using the AQP4 inhibitor TGN-020, has been showed to reduce oedema in imaging studies. Here, we aimed to evaluate, for the first time, the histopathological effects of a single dose of TGN-020 administered after the occlusion of the medial cerebral artery (MCAO). On a rat model of non-reperfusion ischemia, we have assessed vascular densities, albumin extravasation, gliosis, and apoptosis at 3 and 7 days after MCAO. TGN-020 significantly reduced oedema, glial scar, albumin effusion, and apoptosis, at both 3 and 7 days after MCAO. The area of GFAP-positive gliotic rim decreased, and 3D fractal analysis of astrocytic processes revealed a less complex architecture, possibly indicating water accumulating in the cytoplasm. Evaluation of the blood vessels revealed thicker basement membranes colocalizing with exudated albumin in the treated animals, suggesting that inhibition of AQP4 blocks fluid flow towards the parenchyma in the paravascular drainage pathways of the interstitial fluid. These findings suggest that a single dose of an AQP4 inhibitor can reduce brain oedema, even if administered after the onset of ischemia, and AQP4 agonists/antagonists might be effective modulators of the paravascular drainage flow.

Targeting lysyl-oxidase (LOX) may facilitate intramural periarterial drainage for the treatment of Alzheimer's disease.

Alzheimer's disease is the commonest form of dementia. It is likely that a lack of clearance of amyloid beta (Aß) results in its accumulation in the parenchyma as Aß oligomers and insoluble plaques, and within the walls of blood vessels as cerebral amyloid angiopathy (CAA). The drainage of Aß along the basement membranes of blood vessels as intramural periarterial drainage (IPAD), could be improved if the driving force behind IPAD could be augmented, therefore reducing Aß accumulation. There are alterations in the composition of the vascular basement membrane in Alzheimer's disease. Lysyl oxidase (LOX) is an enzyme involved in the remodelling of the extracellular matrix and its expression and function is altered in various disease states. The expression of LOX is increased in Alzheimer's disease, but it is unclear whether this is a contributory factor in the impairment of IPAD in Alzheimer's disease. The pharmacological inhibition of LOX may be a strategy to improve IPAD and reduce the accumulation of Aß in the parenchyma and within the walls of blood vessels.

Taxifolin: A Potential Therapeutic Agent for Cerebral Amyloid Angiopathy.

Cerebral amyloid angiopathy (CAA) is characterized by the accumulation of ß-amyloid (Aß) in the walls of cerebral vessels, leading to complications such as intracerebral hemorrhage, convexity subarachnoid hemorrhage and cerebral microinfarcts. Patients with CAA-related intracerebral hemorrhage are more likely to develop dementia and strokes. Several pathological investigations have demonstrated that more than 90% of Alzheimer's disease patients have concomitant CAA, suggesting common pathogenic mechanisms. Potential causes of CAA include impaired Aß clearance from the brain through the intramural periarterial drainage (IPAD) system. Conversely, CAA causes restriction of IPAD, limiting clearance. Early intervention in CAA could thus prevent Alzheimer's disease progression. Growing evidence has suggested Taxifolin (dihydroquercetin) could be used as an effective therapy for CAA. Taxifolin is a plant flavonoid, widely available as a health supplement product, which has been demonstrated to exhibit anti-oxidative and anti-inflammatory effects, and provide protection against advanced glycation end products and mitochondrial damage. It has also been shown to facilitate disassembly, prevent oligomer formation and increase clearance of Aß in a mouse model of CAA. Disturbed cerebrovascular reactivity and spatial reference memory impairment in CAA are completely prevented by Taxifolin treatment. These results highlight the need for clinical trials on the efficacy and safety of Taxifolin in patients with CAA.

Cerebral Vessels: An Overview of Anatomy, Physiology, and Role in the Drainage of Fluids and Solutes.

The cerebral vasculature is made up of highly specialized structures that assure constant brain perfusion necessary to meet the very high demand for oxygen and glucose by neurons and glial cells. A dense, redundant network of arteries is spread over the entire pial surface from which penetrating arteries dive into the cortex to reach the neurovascular units. Besides providing blood to the brain parenchyma, cerebral arteries are key in the drainage of interstitial fluid (ISF) and solutes such as amyloid-beta. This occurs along the basement membranes surrounding vascular smooth muscle cells, toward leptomeningeal arteries and deep cervical lymph nodes. The dense microvasculature is made up of fine capillaries. Capillary walls contain pericytes that have contractile properties and are lined by a highly specialized blood-brain barrier that regulates the entry of solutes and ions and maintains the integrity of the composition of ISF. They are also important for the production of ISF. Capillaries drain into venules that course centrifugally toward the cortex to reach cortical veins and empty into dural venous sinuses. The walls of the venous sinuses are also home to meningeal lymphatic vessels that support the drainage of cerebrospinal fluid, although such pathways are still poorly understood. Damage to macro- and microvasculature will compromise cerebral perfusion, hamper the highly synchronized movement of neurofluids, and affect the drainage of waste products leading to neuronal and glial degeneration. This review will present vascular anatomy, their role in fluid dynamics, and a summary of how their dysfunction can lead to neurodegeneration.

Cerebral Amyloid Angiopathy Presenting as Massive Subarachnoid Haemorrhage: A Case Study and Review of Literature.

Cerebral amyloid angiopathy (CAA) is characterised by the progressive accumulation of ß-amyloid (Aß) in the walls of cerebral capillaries and arteries representing a major cause of haemorrhagic stroke including lobar intracerebral haemorrhage (ICH) and convexity subarachnoid haemorrhage (SAH). Haemorrhaging from CAA predominantly involves smaller arteries rather than arterial aneurysm. Restricted bleeding into the subarachnoid space in CAA results in asymptomatic or mild symptomatic SAH. Herein, we present an autopsied case of massive SAH related to CAA. An 89-year-old male with a history of mild Alzheimer's disease (AD) and advanced pancreatic cancer with liver metastasis developed sudden onset of coma. Head CT illustrated ICH located in the right frontal lobe and right insula, as well as SAH bilaterally spreading from the basal cistern to the Sylvian fissure, with hydrocephalus and brain herniation. He died about 24 h after onset and the post-mortem examination showed no evidence of arterial aneurysm. The substantial accumulation of Aß in the vessels around the haemorrhagic lesions led to the diagnosis of ICH related to CAA and secondary SAH, which may have been aggravated by old age and malignancy. This case suggests that CAA can cause severe SAH resembling aneurysmal origin and thus may be overlooked when complicated by atypical cerebral haemorrhage.

Retinal Vascular Pathology in a Rat Model of Cerebral Small Vessel Disease.

Introduction: The initial disease stages of hypertensive arteriopathy (HA) and cerebral amyloid angiopathy (CAA), the two main forms of sporadic human cerebral small vessel diseases (CSVD), are too subtle to be detectable on clinical routine imaging. Small vessel disease (SVD) is a systemic condition, affecting not only the brain, but also other organs. The retina appears as an ideal marker for the early detection of incipient CSVD. We therefore investigated the retinal microvasculature of the spontaneously hypertensive stroke-prone rat (SHRSP), an animal model of sporadic CSVD. Materials and Methods: The brains and retinas of 26 male SHRSP (18-44 weeks) were examined histologically and immunohistochemically for the presence of HA phenomena (erythrocyte thrombi, small perivascular bleeds) and amyloid angiopathy (AA). Results: CAA and AA in the retina showed a significant correlation with age (CAA: rho = 0.55, p = 0.005; AA: rho = 0.89, p < 0.001). The number of erythrocyte thrombi in the brain correlated with the severity of retinal erythrocyte thrombi (rho = 0.46, p = 0.023), while the occurrence of CAA correlated with the appearance of AA in the retina (rho = 0.51, p = 0.012). Retinal SVD markers predicted CSVD markers with good sensitivity. Conclusions: These results indicate that SVD also occurs in the retinal microvasculature of SHRSP and the prediction of cerebral erythrocyte thrombi and CAA might be possible using retinal biomarkers. This underlines the important role of the investigation of the retina in the early diagnosis of CSVD.

Immune complex formation impairs the elimination of solutes from the brain: implications for immunotherapy in Alzheimer's disease.

BACKGROUND: Basement membranes in the walls of cerebral capillaries and arteries form a major lymphatic drainage pathway for fluid and solutes from the brain. Amyloid-ß (Aß) draining from the brain is deposited in such perivascular pathways as cerebral amyloid angiopathy (CAA) in Alzheimer's disease (AD). CAA increases in severity when Aß is removed from the brain parenchyma by immunotherapy for AD. In this study we investigated the consequences of immune complexes in artery walls upon drainage of solutes similar to soluble Aß. We tested the hypothesis that, following active immunization with ovalbumin, immune complexes form within the walls of cerebral arteries and impair the perivascular drainage of solutes from the brain. Mice were immunized against ovalbumin and then challenged by intracerebral microinjection of ovalbumin. Perivascular drainage of solutes was quantified following intracerebral microinjection of soluble fluorescent 3kDa dextran into the brain at different time intervals after intracerebral challenge with ovalbumin. RESULTS: Ovalbumin, IgG and complement C3 co-localized in basement membranes of artery walls 24 hrs after challenge with antigen; this was associated with significantly reduced drainage of dextran in immunized mice. CONCLUSIONS: Perivascular drainage along artery walls returned to normal by 7 days. These results indicate that immune complexes form in association with basement membranes of cerebral arteries and interfere transiently with perivascular drainage of solutes from the brain. Immune complexes formed during immunotherapy for AD may similarly impair perivascular drainage of soluble Aß and increase severity of CAA.