The Glymphatic System: A Beginner's Guide

Abstract

The glymphatic system is a recently discovered macroscopic waste clearance system that utilizes a unique system of perivascular tunnels, formed by astroglial cells, to promote efficient elimination of soluble proteins and metabolites from the central nervous system. Besides waste elimination, the glymphatic system also facilitates brain-wide distribution of several compounds, including glucose, lipids, amino acids, growth factors, and neuromodulators. Intriguingly, the glymphatic system function mainly during sleep and is largely disengaged during wakefulness. The biological need for sleep across all species may therefore reflect that the brain must enter a state of activity that enables elimination of potentially neurotoxic waste products, including β-amyloid. Since the concept of the glymphatic system is relatively new, we will here review its basic structural elements, organization, regulation, and functions. We will also discuss recent studies indicating that glymphatic function is suppressed in various diseases and that failure of glymphatic function in turn might contribute to pathology in neurodegenerative disorders, traumatic brain injury and stroke.

Keywords: Aging; Astrocytes; Cerebrospinal fluid secretion; Neurodegenerative diseases; Perivascular spaces; Sleep; The glymphatic system; Traumatic brain injury; Virchow–Robin spaces.

Fig. 1. Schematic representation of the brain's fluid compartments and barriers

The fluid compartments in the brain consist of intracellular fluid (ICF) (60-68%), interstitial fluid (ISF) (or extracellular fluid) (12-20%), blood (10%) and the cerebrospinal fluid (CSF) (10%) [5, 10]. The blood is separated from the CSF and interstitial fluid by the blood brain barrier (BBB) and blood-CSF barrier, respectively. Tight junctions between the blood endothelial cells constitute the BBB, restricting macromolecules to move freely from the blood to the brain parenchyma. Fluid and solutes in the perivascular space located between endothelial cells and astrocytic endfeet, expressing the water channel aquaporin-4 (AQP4) diffuses into the brain parenchyma. The blood-CSF barrier is formed by tight junctions between the choroid plexus epithelial cells. Macromolecules from the blood can move freely between the fenestrated endothelial cells to the interstitial fluid but is restricted by tight junctions in the choroid plexus epithelial cells, which therefore are believed to be the main players in determining CSF composition.

Fig. 2. Ion composition and transport across the choroid plexus epithelial cells

According to the classical hypothesis, ion transporters and channels in the choroid plexus epithelial cells account for the main part of cerebrospinal fluid (CSF) production. The apically expressed Na⁺/K⁺-ATPase, central to CSF secretion, creates the electrochemical gradient for Na⁺ that is imported via the Na⁺/H⁺ exchanger, NHE1 and/or the Na⁺-HCO₃^- co-transporter, NCBE in the basolateral membrane. Co-import of HCO₃^- via NCBE and hydration of CO₂ by carbonic anhydrase (C.A) increase the intracellular concentration of HCO₃^-, which creates an electrochemical gradient that drives the efflux of HCO₃^- via the basolateral-located Cl^-/HCO₃^- exchanger, AE2 and apical HCO₃^- channels. The operation of AE2 generates a rise in intracellular Cl^- driving the apical export of Cl^- through the NKCC1 and Cl^- channels. The final result of these processes at the choroid plexus epithelium is a net flux of Na⁺, HCO₃^- and Cl^- from the blood across the epithelium to the ventricles, which generates the osmotic gradient that makes water move through AQP1 thereby producing the CSF.

Fig. 3. The neurovascular unit

The structure and function of the neurovascular unit allow bidirectional communication between the microvasculature and neurons, with astrocytes playing intermediary roles. Pial arteries in the subarachnoid space bathed in CSF become penetrating arteries upon diving into the brain parenchyma. The perivascular space around penetrating arteries is termed the Virchow-Robin space. As the penetrating arteries branch into arterioles and capillaries the CSF-containing Virchow-Robin spaces narrow and finally disappear. However, the perivascular space extends to arterioles and capillaries to venules where it is made up by the basal lamina's extracellular matrix that provides a continuity of the fluid space between arterioles and venules. Astrocytic vascular endfeet expressing aquaporin-4 (AQP4) surround the entire vasculature and form the boundary of the perivascular spaces.

Fig. 4. In vivo two-photon imaging of glymphatic influx in the mouse cortex

A) Schematic representation of the imaging setup; Periarterial CSF influx of tracers injected in the cisterna magna into the subarachnoid CSF was assessed in vivo using two-photon microscopy through a closed cranial window. Lower bar: Imaging conducted at the cortical surface in 1-min intervals. B) 5 min after injection of small (TR-d3, dark blue) and large (FITC-d2000, green) molecular weight tracers and C-E) 100μm below the cortical surface 10, 20 and 30 min after injection of the tracers. Merge (light blue) indicates co-localization of TR-d3 and FITC-d2000. The cerebral vasculature was visualized with intra-arterial flourescent tracer (CB-d10), and arteries (A) and veins (V) were identified morphologically. Immediately after intracisternal injection, CSF tracer moved along the outside of the pial arteries, but not veins, and after 20-30 min they diffused into the brain parenchyma. Red circles, arterioles; blue circles, venules. Scale bars 100μm. Reprinted from [49] with permission.

Fig. 5. Model of glymphatic function in young, old and in Alzheimer's disease

A) In young and healthy people, CSF enters the brain parenchyma via periarterial pathways, washes out solutes from the interstitial space and empties along the veins. B) With aging, glymphatic function is reduced, possibly due to astrocytes becoming reactive and AQP4 de-polarized from the vascular endfeet to parenchymal processes. C) In Alzheimer's disease perivascular space of penetrating arteries are subject to accumulation of β-amyloid peptides. We hypothesize that accumulation of β-amyloid might be caused by impairment of the glymphatic system and that the perivascular pathways are further blocked by protein aggregates such as β-amyloid. In this model, the resulting changes in the perivascular environment lead to abnormal enlargement of perivascular space downstream, which further decreases glymphatic clearance.

Fig. 6. Abnormal perivascular spaces in Alzheimer's disease

2.5× magnification of hematoxylin and eosin staining of superior frontal gyrus and underlying white matter. A) The white matter of a 74-year old with no CNS diagnosis shows homogenously stained white matter with normal perivascular spaces. B) The white matter of an 80-year old Alzheimer's disease patient contains patches with paler staining and a large number of abnormally enlarged perivascular spaces. 100× magnification of normal C), slightly dilated D) and severely dilated E) perivascular spaces in an Alzheimer's disease patient. Reprinted from [112] with permission.