The glymphatic system is a waste clearance system that utilizes perivascular channels, formed by astroglial cells, to promote efficient elimination of soluble proteins and metabolites from the central nervous system.
The glymphatic system also helps distribute non-waste compounds, such as glucose, lipids, amino acids, and neurotransmitters in the brain.
The glymphatic system functions mainly during sleep and is largely disengaged during wakefulness.
The biological need for sleep reflects the brain must enter a state of activity that enables elimination of potentially neurotoxic waste products, including β-amyloid.
Clearance of excess fluid and interstitial solutes is critical for tissue homeostasis.
In the peripheral tissues soluble material, proteins and fluid from the interstitial space are returned to the general circulation by the lymphatic system.
The lymphatic network extends to all parts of the peripheral tissues and the density of lymph vessels correlates with the rate of tissue metabolism.
The brain and spinal cord are characterized by a disproportionally high metabolic rate.
Synaptic transmission is sensitive to changes in their environment,
The central nervous system (CNS) completely lacks conventional lymphatic vessels.
The brain consists of four fluid compartments: cerebrospinal fluid (CSF), interstitial fluid, intracellular fluid, and the blood vasculature.
The blood is kept separate from the brain parenchyma and CSF by the blood-brain and blood-CSF barriers, respectively.
The blood is separated from the CSF and interstitial fluid by the blood brain barrier (BBB) and blood-CSF barrier, respectively.
These barriers are essential in maintaining the extracellular environment of the brain because they regulate the ionic and biochemical composition of the different fluid compartments.
The blood-brain barrier is comprised by blood vessel endothelial cells.
The blood-CSF barrier is formed primarily between the choroid plexus epithelial cells.
The plexus of capillary endothelial cells, are devoid of tight junctions, they are more permeable to macromolecules.
The choroid plexus epithelial cells, in turn, are connected by tight junctions.
At the choroid plexus, the trans-epithelial transport of macromolecules is regulated by epithelial transporters determining which macromolecules that enter CSF from the blood.
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%).
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, expresses 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.
It is hypothesize that ion transporters and channels in the choroid plexus epithelial cells account for the main part of cerebrospinal fluid (CSF) production.
The ion channel/transporters
processes at the choroid plexus epithelium results in 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.
CSF comprises 10% of the total fluid volume within the cranial cavity
The CSF flows through the four ventricles that are linked by channels or foramina into the subarachnoid space of the cortex and spinal cord
CSF fom the cortical subarachnoid space it penetrates the brain parenchyma perivascularly and bathes the brain before it exits the CNS and drains into the lymphatic system.
CSF is thought to be produced primarily by the choroid plexuses, which are expansions of the ependymal epithelium, lining the lateral, third, and fourth ventricles.
The choroid plexuses are highly folded and vascularized structures consisting of a single layered cuboidal or low cylindrical epithelium residing on a basement membrane.
The luminal surface area of the choroid plexus epithelial cells is covered by microvilli and possess either one primary cilia or small tufts of motile cilia.
CSF production at the choroid plexus is mediated by exchange and transport of ions, especially Cl-, Na+ and HCO3, across the epithelial cells, which generates an osmotic gradient that drives the movement of water from the blood to the ventricle lumen.
The Na+/K+-ATPase localized in the apical membrane of the choroid plexus epithelial cell is central for CSF production.
Ouabain, which inhibits the Na+/K+-ATPase, reduces CSF production by 50-60%.
The Na+/K+-ATPase actively pumps out Na+ from the epithelial cell to the CSF in the ventricular lumen, thereby keeping the intracellular Na+ concentration low (45 mM compared to the extracellular 141-152 mM)
There us a transmembrane gradient, which drives the import of Na+ across the basolateral membrane possibly via the Na+-dependent HCO3- co-transporter, NCBE [13] and/or the Na+/H+ exchanger NHE1.
HCO3- and its transcellular exchange with Cl- have also been demonstrated to be important for CSF production.
Application of acetazolamide or, inhibitors of carbonic anhydrases and anion exchange, respectively, reduces CSF formation by 30-50%
Overall there is a net movement of Na+, Cl- and HCO3- from the blood across the choroid plexus epithelium to the ventricles.
This outward movement of Na+, Cl- and HCO3- generates the osmotic gradient that drives water in the same direction across the apical membrane.
Water fluxes across the choroid plexus epithelium take place mainly through the highly water permeable channel, AQP1.
AQP1, located primarily in the choroid plexus epithelial cells.
Overall, the net result of ion and water movement across the choroid plexus epithelium is production of CSF that, compared to the blood, is lower in protein and K+, and higher in Na+, Cl- and Mg2+ and has a 99% water content compared to a water content of 92% in plasma.
CSF is continuously produced.
It is renewed approximately four times each 24hours, and the total CSF volume of 150-160 mL in humans
Its volume is kept constant by removal of CSF.
CSF is drained into the peripheral lymphatic system by efflux via the olfactory bulb and along cranial and spinal nerves.
Efflux along cranial and spinal nerves and the olfactory route are among the most important efflux pathways for CSF.
The choroid plexuses alone are responsible for the vast majority (80-90%) of CSF formation.
That CSF formation occurs by filtration and flux of fluid through the capillary walls, and the respective volumes of CSF and interstitial fluid mainly depend on hydrostatic and osmotic forces between the CSF and brain parenchyma created by gradients of proteins and inorganic ions across the capillary membrane.
In physiological conditions, water is filtered from capillaries with high capillary pressure, to the interstitial fluid and CSF.
In these capillaries, the permeability of plasma electrolytes such as Na+ and Cl− is low and the electrolytes are therefore retained.
The CSF and interstitial fluid are continuously interchanging and the volume occupied by each compartment depends on hydrostatic and osmotic forces.
The arterial cerebral circulation consists of an anterior cerebral circulation and posterior cerebral circulation supplied by the internal carotid arteries and the vertebral arteries, respectively.
The anterior circulation, which includes the middle and anterior cerebral arteries, communicates with the posterior circulation, the basilar artery and posterior cerebral arteries, via anterior and posterior communicating arteries at the Circle of Willis.
From the Circle of Willis, the anterior circulation perfuse the younger parts of the brain including the neocortex of the cerebral hemispheres, while the posterior circulation supplies the brainstem and cerebellum.
At the cortical surface, cerebral arteries extend into pial arteries running through the CSF-containing subarachnoid space and the subpial space.
As pial arteries dive down into the brain parenchyma they transition into penetrating arterioles and create a perivascular space, known as the Virchow-Robin space.
The Virchow-Robin spaces are filled with CSF and bordered by a leptomeningeal cell layer on both the inner wall facing the vessel and on the outer wall facing perivascular astrocytic endfeet.
The CNS vasculature is unique in that all arterioles, capillaries, and venules within the brain parenchyma are surrounded by astrocytic vascular endfeet.
The vascular endfeet of the astrocytes create the outer wall of the perivascular space surrounding the vasculature.
As arterioles go deeper down in the brain parenchyma, the Virchow-Robin spaces become continuous with the basal lamina.
Thus, the Virchow-Robin space disappears before the capillary level where the perivascular space consists solely of basal lamina.
The basal lamina is a thin sheet of extracellular matrix.
The basal lamina is composed primarily of laminin, fibronectin, type IV collagen, heparin sulfate proteoglycan and other extracellular matrix components.
The basal lamina separates endothelial cells, pericytes and astrocytes.
Endothelial cells, pericytes and astrocytes, smooth muscle and neurons comprise the neurovascular unit which is tightly linked to the extracellular matrix of the basal lamina by adhesion molecules, including integrins and dystroglycan.
Due to the porous structure of extracellular matrix the basal lamina provides minimal resistance to CSF influx.
The neurovascular unit allows 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.
The penetrating arteries branch into arterioles and capillaries the CSF-containing Virchow-Robin spaces narrow and finally disappear.
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.
The perivascular spaces constitute a low resistance pathway for influx of CSF, and are also important sites for delivery of energy substrate and regulation of blood flow.
With pathological conditions, such as stroke, the innate inflammatory response and edema formation is initiated in the perivascular spaces.
From the cerebral capillaries blood continues into the post-capillary venules where the basement membranes of endothelial cells and astrocytes enlarge to again provide a CSF-drained perivascular space.
In general, blood from the brain’s interior, including the deep white and gray matter surrounding the lateral and third ventricles flows into the larger central/deeps veins and exits the cerebral cortex and subcortical white matter via the cortical veins that extend to the brain surface as pial veins.
The territories drained by the central veins and cortical veins reveal a marked degree of overlapping.
The superficial cortical veins anastomose with the deep veins and empty into the superior sagittal sinus.
Cerebral venous blood from the superior sagittal sinus and the deep veins leave the brain via a confluence of sinuses draining into the sigmoid sinuses and jugular veins.
The CSF and interstitial fluid (ISF) continuously interchange, and
The exchange is facilitated by influx of CSF along the periarterial space.
From the subarachnoid space, CSF is driven into the Virchow-Robin spaces by arterial pulsatility, respiration, and CSF pressure gradients and the loose fibrous matrix of the perivascular space can be viewed as a low resistance highway for CSF influx.
The transport of CSF into the brain parenchyma is facilitated by AQP4 water channels expressed in a highly polarized manor in astrocytic endfeet that ensheathe the brain vasculature.
CSF movement into the parenchyma of the brain drives interstitial fluid fluxes within the tissue toward the perivenous spaces surrounding the large deep veins.
The interstitial fluid is collected in the perivenous space from where it drains out of brain toward the cervical lymphatic system.
This system of convective fluid fluxes with rapid interchange of CSF and interstitial fluid is entitled the glymphatic system based on its similarity to the lymphatic system in the peripheral tissue in function, and on the important role of glial AQP4 channels in the convective fluid transport.
The macroscopic clearance mechanism of interstitial solutes may be of particular importance for neurodegenerative diseases including Alzheimer’s disease, which is characterized by the accumulation of proteins, including amyloid plaques and tau tangles.
The paravascular glymphatic pathway driven by AQP4-dependent bulk flow constitutes a major clearance pathway of interstitial fluid solutes from the brain’s parenchyma.
The human brain weighs 2% of the body’s total weight but contains 25% of the cholesterol in the human body.
Despite the brain being highly enriched in cholesterol, the blood-brain-barrier prevents influx of lipids and lipoproteins, including cholesterol, from the blood to the brain.
Unlike peripheral tissues, which obtain blood-borne cholesterol secreted by the liver, the brain synthesizes all its cholesterol de novo.
Excess cholesterol is eliminated from the brain by hydroxylation of cholesterol to 24-OH cholesterol.
80% of the 24-OH cholesterol in the body is found in the brain and the circulatory system acts as sink for excess cholesterol produced in the brain.
The brain’s internal lipid transport is via its own supply of lipid carrier particles of the high density lipoprotein type secreted by astrocytes
Secretion of high density lipoprotein particles from astrocytes is dependent on lipoproteins, mainly Apolipoprotein E and J, and allows delivery of lipids such as cholesterol to neurons.
Apolipoprotein carriers also mediate clearance of excess hydroxylated cholesterol and β-amyloid.
The Apolipoprotein E allele 4 is a major genetic risk factor for Alzheimer’s disease, and this lipid carrier is important for maintaining homeostasis necessary for a healthy environment of the brain.
Apolipoprotein E is concentrated in astrocytic processes at the pial surface and around the blood vessels
In addition, the choroid plexus and cells in the wall of the third ventricle also produce Apolipoprotein E.
Thus, Apolipoprotein E production is co-localized with CSF production sites and transport pathways suggesting that lipids are transported by the glymphatic system.
The glymphatic system is comparable to the peripheral lymphatic system that transports the dietary fat, incl. cholesterol, absorbed in the intestine.
Only the <1 kDa lipophilic tracers diffuse into the brain parenchyma whereas the larger tracers were largely confined to the perivascular routes, suggesting that the glymphatic system plays a central role in macroscopic distribution of lipids in the brain and that medium to large lipid soluble molecules might require carrier particles in order to be delivered via the CSF.
Astrocytes thus play a key role in lipid synthesis and lipid distribution by releasing lipid carrier proteins, such as Apolipoprotein E, and in maintaining the highway for distribution, the glymphatic system.
Glymphatic transport of CSF along the periarterial spaces, followed by convective flow through the brain parenchyma, and exit of interstitial fluid (ISF) along the perivenous space to the cervical lymph system, is requires energy driven by multiple mechanisms.
The production of CSF by the choroid plexus creates a pressure that dictates the direction of the fluid flow through the ventricular system to the subarachnoid space.
Respiration is instrumental in movement of CSF through the aqueduct.
Entry of CSF along the perivascular space is facilitates glymphatic ISF-CSF exchange and clearance function.
Glymphatic activity is driven, in part, by arterial pulsatility and explains why perivascular influx occurs preferentially around pulsating arteries and not cerebral veins.
The glymphatic system is turned on during sleep, compromising alertness.
Sleep enhances memory consolidation.
The basic biological need for sleep is unclear.
Brain energy metabolism only declines by 25% during sleep suggesting that sleep does not simply serve to conserve energy
Glymphatic activity is dramatically enhanced during sleep while its function is suppressed during wakefulness.
The sleep state is conducive to convective fluid fluxes and thereby to clearance of metabolites.
A major function of sleep is thought to be that the glymphatic system is turned on and that the brain clears itself of neurotoxic waste products produced during wakefulness.
This suggests that it is the differences in the sleep versus wakeful state and not daily circadian rhythms that regulate glymphatic activity.
Norepinephrine also is a key regulator of glymphatic activity and that it might be responsible for suppression of glymphatic during wakefulness.
The burst release of norepinephrine during arousal increases the cellular volume fraction resulting in a decrease in the interstitial space.
Norepinephrine also acts directly on choroid plexus epithelial cells and inhibits CSF production.
Removal of norepinephrine signaling, mimicking the sleep state, enhances CSF production.
Norepinephrine acts via different mechanisms on both fluid availability and convective fluxes to suppress glymphatic function and norepinephrine can therefore be considered both a key regulator of the switch between the sleep and wakeful state and solute clearance from the brain.
Glymphatic activity decreases sharply during aging.
Reactive gliosis contributes to the age-related decline in glymphatic function.
AQP4 plays a central role in facilitating CSF-ISF exchange along periarterial influx pathways as well as interstitial solute clearance through perivascular drainage pathways
The genetic deletion of AQP4 was previously shown to impair CSF-ISF exchange by ∼65% and reduce the clearance of β–amyloid by ∼55%
The vascular polarization of astrocytic AQP4 is partly lost in reactive astrocytes in old brains.
Aging is associated with loss of perivascular AQP4 polarization, along penetrating arterioles, and that presence of cortical parenchymal AQP4 correlates with CSF-ISF exchange, suggests that the age related decline in glymphatic function might be in part attributable to dysregulation of astroglial water transport.
Other factors perhaps contributing to the reduction of glymphatic activity with aging are the decline in CSF production by 66% and CSF pressure by 27% percent.
Aging accompanied by stiffening of the arterial wall leads to a reduction in arterial pulsatility, which is one of the drivers of glymphatic influx.
The failure of the glymphatic system in aging might thus contribute to the accumulation of misfolded and hyperphosphorylated proteins and thereby render the brain more vulnerable to developing a neurodegenerative pathology or perhaps escalate the progression of cognitive dysfunction.
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.
With aging, glymphatic function is reduced, possibly due to astrocytes becoming reactive and AQP4 de-polarized from the vascular endfeet to parenchymal processes.
In Alzheimer’s disease perivascular space of penetrating arteries are subject to accumulation of β-amyloid peptides.
It is hypothesized 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 resulting in changes in the perivascular environment lead to abnormal enlargement of perivascular space downstream, which further decreases glymphatic clearance.
Neurodegenerative diseases are characterized by accumulation of aggregated proteins.
Toxic protein monomers, oligomers, and aggregates are present in the interstitial fluid and CSF misfolded β–amyloid and fibrillary tangles of tau in Alzheimer’s disease, misfolded α-synuclein in Parkinson’s disease and misfolded superoxide dismutase in mouse models of amyotrophic lateral sclerosis.
β-amyloid production is highest during the awake state when neuronal activity is highest.
Neurologic cells produce β-amyloid and in particular oligodendrocytes and their precursor cells, which, at least in part, might account for the myelin disorders observed in Alzheimer’s disease.
The production and turnover of β-amyloid is rapid.
In healthy, young people 8.3% of total β-amyloid is cleared each hour via the CSF.
Interstitial β-amyloid in Alzheimer’s disease and perivascular spaces are a sites for both amyloid deposits and Alzheimer’s disease pathology.
The clearance by the glymphatic system in combination with transport across the blood-brain barrier are necessary and sufficient to remove of extracellular β-amyloid until the end of the reproductive lifespan around which time failure in adequate CSF bulk flow leads to accumulation of β-amyloid.
It is likely low activity of the glymphatic system could be a major risk factor for development of neurodegenerative diseases.
β-amyloid accumulates predominantly at cerebral arteries.
Abnormal enlargement of the perivascular space is more frequently observed in Alzheimer’s disease, suggesting a spiral of protein accumulation, deformation of glymphatic routes and further reduction in protein clearance.
Abnormalities at the perivascular space are also prominent in non-Alzheimer’s dementia.
Vascular dementias are the second most common cause of dementia and these diseases are also characterized by deformation of the perivascular space.
Changes in or surrounding cerebral blood vessels due to hypertension, atherosclerosis or hereditary diseases can alcause vascular dementia.
Vascular dementia is often caused by pathology in small cerebral blood vessels and capillaries, collectively termed small vessel disease.
In small vessel disease enlargement of the perivascular space is frequently observed.
Traumatic brain injury, increases the risk of premature dementia and Alzheimer’s disease.
Multiple studies have shown that repeated traumatic events, and even single events of moderate to severe head trauma, can lead to progressive neurodegeneration.
Traumatic brain injury induces release of β-amyloid peptide and of C-tau, which is a highly abundant intracellular microtubule protein in axons.
C-tau is a biomarker of brain injury since it is released in vast quantities and correlates with severity of traumatic brain injury.
A large increase in interstitial tau leads to cellular uptake and initiation of fibrillary aggregates, which attracts additional tau leading to formation of neurofibrillary tangles ultimately resulting in a prion-like spread of the pathology.
Traumatic brain injury is linked to large astroglial scars and persistent activation of innate neuroinflammation.
With repetitive moderate traumatic brain injury, the influx of CSF into the brain and CSF-mediated clearance is impaired in the ipsilateral hemisphere at day 1 post injury.
The reduction of glymphatic function persists until at least 28 days post injury.
The decrease in glymphatic function is associated with glial scars characterized by hypertrophic GFAP-positive processes in the ipsilateral hemispheres.
CSF-mediated removal of tau by glymphatic routes is crucial for limiting secondary neuronal damage following traumatic brain injury.
Head trauma, such as subarachnoid hemorrhage, severely impairs glymphatic function.
Cerebral hemorrhage can cause a widespread inhibition of glymphatic function.
Biomarkers of traumatic brain injury exit the brain via the glymphatic system.
Inhibition of glymphatic activity by four mechanistically distinct manipulations, including sleep deprivation, cisterna magna puncture, inhibition of CSF production using acetazolamide, or genetic deletion of AQP4 resulted in marked reduction of astrocytic proteins.
Acute injury, including traumatic brain injury, subarachnoid bleeding or stroke, profoundly impact glymphatic function and impair convective fluid flow.
The impairment of glymphatic function could further exacerbate injury due to accumulation of both normal metabolic waste as well as injury-induced debris.
Accumulating evidence suggests related decline in glymphatic clearance in significant ways contributes to accumulation of protein aggregation.