Flow of cerebrospinal fluid (CSF) through perivascular spaces (PVSs) in the brain is important for clearance of metabolic waste. Arterial pulsations are thought to drive flow, but this has never been quantitatively shown. We used particle tracking to quantify CSF flow velocities in PVSs of live mice. CSF flow is pulsatile and driven primarily by the cardiac cycle. The speed of the arterial wall matches that of the CSF, suggesting arterial wall motion is the principal driving mechanism, via a process known as perivascular pumping. Increasing blood pressure leaves the artery diameter unchanged but changes the pulsations of the arterial wall, increasing backflow and thereby reducing net flow in the PVS. Perfusion-fixation alters the normal flow direction and causes a 10-fold reduction in PVS size. We conclude that particle tracking velocimetry enables the study of CSF flow in unprecedented detail and that studying the PVS in vivo avoids fixation artifacts.

Background The glial-lymphatic or glymphatic pathway is a fluid clearance pathway recently identified in the rodent brain. This pathway subserves the flow of cerebrospinal fluid (CSF) into the brain along arterial perivascular spaces and thence into the brain interstitium facilitated by aquaporin-4 (AQP4) water channels. The pathway then directs flows towards the venous perivascular and perineuronal spaces, ultimately clearing solutes from the neuropil into meningeal and cervical lymphatic drainage vessels. In rodents, the glymphatic pathway is primarily active during sleep, when the clearance of harmful metabolites such as amyloid β (Aβ) increases two-fold relative to the waking state. Glymphatic dysfunction has been demonstrated in animal models of traumatic brain injury (TBI), Alzheimer’s disease (AD) and micro-infarct disease, most likely in relation to perturbed expression of AQP4. The recent characterizations of the glymphatic and meningeal lymphatic systems calls for revaluation of the anatomical routes for CSF-ISF flow and the physiological role that these pathways play in CNS health. Recent developments Recent work has revealed that several features of the glymphatic and meningeal lymphatic systems are also present in humans. MRI imaging of intrathecally-administered contrast agent shows that CSF flows along pathways closely resembling the glymphatic system outlined in rodents. Furthermore, PET studies reveal that Aβ accumulates in the healthy brain after a single night of sleep deprivation, suggesting that the human glymphatic pathway might also be primarily active during sleep. Other PET studies have shown that CSF clearance of Aβ and tau tracers is reduced in patients with AD compared to healthy controls. The observed reduction in CSF clearance was associated with increasing grey matter Aβ levels in human brain, which is consistent with findings in mice showing that decreased glymphatic function leads Aβ accumulation. Altered AQP4 expression is also evident in brain tissue from AD or normal pressure hydrocephalus (NPH) patients; glymphatic MRI of NPH patients shows reduced CSF tracer entry and clearance. Where next? Future research is needed to confirm if specific factors driving glymphatic flow in rodents also apply to humans. Conducting longitudinal imaging studies to evaluate human CSF dynamics will determine if there is indeed a causal link between reduced brain solute clearance and the development of neurodegenerative diseases. Assessment of glymphatic function after stroke or TBI could identify if it correlates with neurological recovery. Gaining new insights into how behavior and genetics modify glymphatic function, and how this decompensates in disease should lead to the development of new preventive and diagnostic tools, as well as novel therapeutic targets.

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The glymphatic system is a brain-wide clearance pathway; its impairment contributes to the accumulation of amyloid-β. Influx of cerebrospinal fluid (CSF) depends upon the expression and perivascular localization of the astroglial water channel aquaporin-4 (AQP4). Prompted by a recent failure to find an effect of Aqp4 knock-out (KO) on CSF and interstitial fluid (ISF) tracer transport, five groups re-examined the importance of AQP4 in glymphatic transport. We concur that CSF influx is higher in wild-type mice than in four different Aqp4 KO lines and in one line that lacks perivascular AQP4 (Snta1 KO). Meta-analysis of all studies demonstrated a significant decrease in tracer transport in KO mice and rats compared to controls. Meta-regression indicated that anesthesia, age, and tracer delivery explain the opposing results. We also report that intrastriatal injections suppress glymphatic function. This validates the role of AQP4 and shows that glymphatic studies must avoid the use of invasive procedures.

Glymphatic transport, defined as cerebrospinal fluid (CSF) peri-arterial inflow into brain, and interstitial fluid (ISF) clearance, is reduced in the aging brain. However, it is unclear whether glymphatic transport affects the distribution of soluble Aβ in Alzheimer’s disease (AD). In wild type mice, we show that Aβ40 (fluorescently labeled Aβ40 or unlabeled Aβ40), was distributed from CSF to brain, via the peri-arterial space, and associated with neurons. In contrast, Aβ42 was mostly restricted to the peri-arterial space due mainly to its greater propensity to oligomerize when compared to Aβ40. Interestingly, pretreatment with Aβ40 in the CSF, but not Aβ42, reduced CSF transport into brain. In APP/PS1 mice, a model of AD, with and without extensive amyloid-β deposits, glymphatic transport was reduced, due to the accumulation of toxic Aβ species, such as soluble oligomers. CSF-derived Aβ40 co-localizes with existing endogenous vascular and parenchymal amyloid-β plaques, and thus, may contribute to the progression of both cerebral amyloid angiopathy and parenchymal Aβ accumulation. Importantly, glymphatic failure preceded significant amyloid-β deposits, and thus, may be an early biomarker of AD. By extension, restoring glymphatic inflow and ISF clearance are potential therapeutic targets to slow the onset and progression of AD.