Cerebral Blood Vessels and the Neurovascular Unit

The total surface of the capillary endothelial cells forms the major interface between the blood and brain. Other, less extensive, interface surfaces include choroid plexuses and arachnoid granulations (Table 59.2). At each of the BBB interfaces, high-resistance junctions between cells, which make the surface into an epithelial-like structure, restrict bulk transport. The epithelial sheets impede non–lipid-soluble substances, charged substances, or large molecules, whereas lipid-soluble substances, such as anesthetic gases, pass easily through the cells. Water has an anomalous structure that allows it to pass rapidly through endothelial cells but with slight restrictions.

Table 59.2 Characteristic Features of the Blood-Brain Interfaces

ATPase, Adenosine triphosphatase; CSF, cerebrospinal fluid; ISF, interstitial fluid.

In 1925, Cushing and Weed described the pathways involved in the movement of CSF and ISF through the ventricles and around the brain, recognizing that the CSF/ISF acted as a lymph-like fluid in brain. They compared that circulation to those of the blood and lymph. The importance of this circulation has grown with the discovery that ISF is formed by cerebral blood vessels, which have electrolyte pumps that make fluid in a fashion similar to that of the epithelial cells. Flowing around cells, ISF brings nutrients such as glucose and oxygen to neurons and astrocytes and removes the products of metabolism. ISF is absorbed either into the blood via terminal capillaries and venules or into CSF for eventual absorption through the arachnoid granulations (Fig. 59.1).

Fig. 59.1 Illustration of the third circulation. Cerebrospinal fluid (CSF) is formed by the choroid plexuses in the ventricles, and interstitial fluid (ISF) is formed by cerebral capillaries. At both sites, the action of the Na⁺/K⁺-ATPase pump creates the osmotic gradient that pulls water from the blood. Tight junctions (TJ) are found at each site of blood/brain interface. This includes the apical surface of the choroid plexus epithelial cells, the cerebral endothelial cells, and the arachnoid. Substances move between the brain and CSF across the gap junctions (GJ) on ependymal and pial surfaces.

Brain extracellular space comprises 15% to 20% of the total brain volume. Complex carbohydrates are found in the extracellular space, including hyaluronic acid, chondroitin sulfate, and heparan sulfate. Hyaluronic acid forms large water domains. These large extracellular matrix glycoproteins impede cell movement. After an injury, astrocytes secrete an extracellular molecule, hyaluron, which impedes movement of fluids in the extracellular space, slowing tissue repair. Treatment with hyaluronidase reduces hyaluron and improves regrowth of injured fibers (Back et al., 2005).

Proteases are secreted during development, angiogenesis, and neurogenesis to clear a path for the growing cells, similar to the secretion of proteases by spreading cancer cells (Candelario-Jalil et al., 2009; Yong, 2005). An important concept that has emerged over the past few years is that of the neurovascular (or gliovascular) unit. Neurons, astrocytes, and pericytes comprise the neurovascular unit (Neuwelt et al., 2008). On the abluminal surface of the endothelial cells is a thin layer of basal lamina composed of type IV collagen, fibronectin, heparan sulfate, laminin, and entactin. Entactin connects type IV collagen and laminin to add a structural element to the capillary. Fibronectin from the cells joins the basal lamina to the endothelium. Basal lamina provides structure through type IV collagen, charge barriers by heparan sulfate, and binding sites on the laminin and fibronectin molecules. Within the basal lamina reside the pericytes, which are a combination of smooth muscle and macrophage. Astrocyte foot processes surround the basal lamina. Glia limitans is found at the pial surface and at the interface between astrocytes and blood vessels (Owens et al., 2008). Neurons complete the group of cells that comprise the neurovascular unit (Girouard and Iadecola, 2006) (Fig. 59.2).

Fig. 59.2 The cerebral capillary is a fluid-secreting, epithelial-like cell with a high metabolic rate. The Na⁺/K⁺-ATPase pump on the apical surface forms cerebrospinal fluid. Tight junctions (TJ) between the endothelial cells maintain the electrical resistance. A large number of mitochondria are seen in the capillary. Amino acid and glucose transporters are present. Around the cell is a basal lamina composed of type IV collagen, laminin, fibronectin, and heparan sulfate. Astrocytic end-feet surround cells. Pericytes, which are embedded in the basal lamina, are macrophage-like cells that have macrophage and smooth-muscle functions in the perivascular space.

Cerebral blood vessels have very low permeability and high electrical resistance, making them more similar to epithelial cells than systemic capillaries, which are passive structures with low electrical resistance and fenestrations that permit passage of large protein molecules. In addition, cerebral blood vessels have highly selective molecular transport properties. During development, cerebral blood vessels acquire the characteristics that distinguish them from systemic capillaries. Astrocytes are critical in this differentiation process, which involves interactions between blood vessels and astrocytes. The critical nature of the astrocytes in this process was shown in transplantation studies involving chicken and quail cells, which can be separated histologically. Quail brain grafts from 3-day-old quails transplanted into the coelomic cavity of chick embryos become vascularized by chick endothelial cells and form a competent BBB. On the other hand, when avascular embryonic quail coelomic grafts are transplanted into embryonic chick brain, chick endothelial cells form leaky capillaries and venules (Stewart and Wiley, 1981). Subsequent studies showed that astrocytes induced BBB properties in non-neural endothelial cells in vivo (Janzer and Raff, 1987).

Electron microscopic studies with electron-dense tracers identified a major site of the BBB as the endothelial tight junctions (Reese and Karnovsky, 1967). Isolation of tight-junction proteins and cloning of the molecules permitted production of antibodies that identified their location. Zona occludins tether the tight-junction proteins to actin within the endothelial cells (Hawkins and Davis, 2005). Occludin and claudin form the actual tight junctions within the endothelial clefts (Furuse et al., 1993). Occludin attaches to the zona occludins, while claudins attach to occludin and protrude into the clefts between cells. The extracellular tails of claudins from adjacent cells self-assemble to form the tight junctions that are “zip-locked” together (Nitta et al., 2003).

Tight junctions between the endothelial cells create the unique membrane properties of the cerebral capillaries by greatly increasing the electrical resistance, which blocks transport of non–lipid-soluble substances (Box 59.1). Brain tissue has a very high demand for glucose and essential amino acids, which can be met by specialized molecules that transport glucose and amino acids across the BBB. Glucose transporters are densely distributed in the capillaries. At low levels of blood glucose, the carriers function at full capacity to meet metabolic needs, but at higher levels of blood glucose, the carriers are saturated and transport is dominated by diffusion rather than active transport. Several isoforms of the glucose transporter molecule have been isolated and cloned (Vannucci et al., 1997). High concentrations of one isoform, GLUT1, are found on cerebral blood vessels. GLUT3 is found on neurons and GLUT5 in microglia. GLUT2 is found predominantly in the liver, intestine, kidney, and pancreas. Impairment of glucose transport due to lack of transporters has been described in patients with Alzheimer disease and in other neurological disorders (Kalaria and Harik, 1989). Amino acid transporters carry essential amino acids into the brain. Competition for the amino acid transporters can lead to a deficiency state; serotonin uptake is decreased in patients with phenylketonuria, which competes for the transporter.

Steady-state levels of brain electrolytes are preserved by transport mechanisms at the BBB. Potassium is maintained at a constant level in the CSF and brain by the BBB (Katzman and Pappius, 1973). This prevents fluctuations of electrolyte levels in the blood from influencing brain levels. Calcium is similarly regulated. Glutamate, which is an excitotoxin, is excluded from the brain. Highly lipid-soluble gases such as carbon dioxide and oxygen are rapidly exchanged across the capillary. Anesthetic gases are effective because they readily cross the BBB and enter the brain.

The presence of the BBB creates a major impediment for the transport of drugs into the brain. For example, penicillin is restricted from entry into the brain, so high doses are needed to achieve therapeutic brain levels. Newer generations of antibiotics such as the cephalosporins penetrate more readily, making them better agents for treatment of brain infections. Chemotherapy of brain tumors has been hampered by the poor lipid solubility of most agents; to overcome this impediment, chemotherapeutic agents can be injected intrathecally or into permanent CSF catheters implanted into the ventricles, with injection bulbs buried beneath the scalp. Drugs of addiction are often modified to allow them to more readily cross the BBB. For example, heroin, which is derived from morphine, is modified to increase its lipid solubility to make it more easily transported into the brain. Similarly, other addictive substances such as nicotine and alcohol are highly lipid soluble and easily transported into brain.

Different rates for equilibration of various substances between blood and brain can cause paradoxical clinical situations. For example, to compensate for a metabolic acidosis, bicarbonate levels fall in both the blood and the brain. Metabolic acidosis is balanced by a respiratory alkalosis due to lowering of carbon dioxide by hyperventilation, which compensates for the acidosis. Carbon dioxide is reduced in both the blood and CSF compartments, since it readily crosses the BBB, while bicarbonate is much more slowly exchanged between the two compartments. This adjustment results in a stable, albeit pathological, situation. However, when the metabolic acidosis is corrected by intravenous infusion of bicarbonate, there is a rapid adjustment of Pco₂ as the hyperventilation stops and CO₂ builds up. Bicarbonate adjusts very slowly because of the limited transport across the BBB, and the CO₂ entering the brain causes a further fall in brain pH. This dangerous situation continues until the bicarbonate levels in the brain rise. Although treatment is necessary to correct the metabolic acidosis, patients may become worse due to brain acidosis if treatment is too rapid (Posner and Plum, 1967).

Production of Cerebrospinal Fluid and Interstitial Fluid

Production of brain fluids comes from multiple sources including the choroid plexuses within the ventricles, the electrolyte pumps on the abluminal surface of the cerebral capillaries, and metabolism. The main source is the choroid plexuses that form an important interface between CSF and blood. Choroid plexuses protrude into the cerebral ventricles; they are covered with a specialized type of ependymal cell that has tight junctions on the apical surface.

Choroid plexus capillaries are fenestrated. Substances from the blood can cross into the stroma next to the ependymal cells. They are blocked from entering the CSF by tight junctions that form at the apical surface of the ependymal cells. Choroid plexus ependymal cells are enriched with mitochondria, Golgi complexes, and endoplasmic reticulum—suggesting a high rate of metabolic activity—and are covered with microvilli that increase their surface area.

While choroid plexuses are the major source of CSF, capillaries contribute ISF to varying degrees, depending on the species. Production of CSF is constant across species when the volume of fluid formed is divided by the weight of the choroid plexus (Cserr, 1971). In humans, the volume of CSF in the ventricles is 140 mL, with a rate of CSF production of 0.35 mL/min or about 500 mL/day. Obstructive hydrocephalus can rapidly cause life-threatening symptoms when the rate of production remains constant but there is no absorption. On the other hand, removal of 20 mL of CSF at the time of LP for the treatment of idiopathic intracranial hypertension does not make physiological sense (although it seems to help at times, which may be due to the hole placed in the dura, with a slow leak of CSF).

CSF production occurs at both choroidal and extrachoroidal sites, and estimates of the proportion of CSF from each site vary, depending on the species and the method of measurement. Extrachoroidal production accounts for 30% of total CSF production in the cat (Rosenberg et al., 1980) and approximately 60% in the nonhuman primate (Milhorat, 1969). No measurements of the relative proportion of CSF produced at each source have been made in humans.

Higher levels of sodium, chloride, and magnesium and lower levels of potassium, calcium, bicarbonate, and glucose are found in CSF than are expected from a plasma ultrafiltrate, which suggests that the CSF is actively secreted. An ATPase pump on the apical surface of the choroidal cells secretes three sodium ions in exchange for two potassium ions; osmotic water follows the increased sodium gradient. Carbonic anhydrase converts carbon dioxide and water into bicarbonate, which is removed along with chloride to balance the sodium charge.

Production of CSF continues even when the ICP is high. Only acetazolamide, which inhibits carbonic anhydrase, can be used for the long-term reduction in CSF production. Experimentally, hypothermia, hypocarbia, hypoxia, and hyperosmolality have been shown to reduce production, but these are not practical to use for other than short periods. Osmotic agents such as mannitol and glycerol increase serum osmolality, lowering CSF production temporarily by about 50%. Agents that interfere with Na⁺/K⁺-ATPase reduce CSF production. Digitalis has an effect on the rate of CSF production, but ouabain, which is a more effective agent experimentally, is too toxic for use in patients. Recently, hypertonic saline has been shown to reduce CSF pressure; some of this effect may be due to a reduction in CSF production, but the mechanism of action remains to be clarified.

Capillaries, which have Na⁺/K⁺-ATPase on the abluminal surface, are a source of extrachoroidal ISF production. Gray matter has a dense neuropil that impedes the flow of water, whereas white matter, being more regularly arranged, is a conduit for normal flow of ISF as well as a route for movement of edema in pathological conditions. Normally the flow of ISF in the white matter is toward the ventricle, where it mixes with the CSF from the choroid plexus to be eventually drained across the arachnoid granulations that protrude into the sagittal sinus.

Water Molecules: Basis for Magnetic Resonance Imaging

Resonance signals detected by MRI are from water molecule protons. Since water is the most abundant source of protons in the brain, water protons dominate the signals. New rapid-acquisition pulse sequences are fast enough to show diffusion of water. Cytotoxic edema shrinks the extracellular space and restricts the diffusion of water (Moseley et al., 1990). The ability to monitor water diffusion by MRI has greatly improved our ability to diagnose an acute ischemic event. Water diffusion between cells in the extracellular space occurs normally. When there is cellular swelling and the extracellular space shrinks, the diffusion of water slows, and the apparent diffusion coefficient (ADC) shows a loss of signal, which appears black on the image. The diffusion-weighted image (DWI) has a bright signal. Because the DWI may show T2 shine-through that will be misinterpreted as reduced diffusion, both a darkened ADC and a bright DWI should be seen in the region of the infarct. In cerebral ischemia, the DWI is abnormal within minutes after the onset of the ischemia, making this an excellent diagnostic test for the presence of cerebral ischemia (Adami et al., 2002).

Diffusion tensor imaging (DTI) reveals the patterns of white matter tracts in three dimensions. Taking advantage of the directional flow of water protons along white matter, diffusion is measured in three planes, and the separate pathways for water movement between the fibers are traced. In patients with white-matter pathology, such as in vascular cognitive impairment and MS, injury patterns in the white matter can be revealed by DTI (Nitkunan et al., 2008).

Contrast agents are important in determining injury to the BBB. Iodine-containing contrast agents are used in CT scanning because they are radiopaque. When injected intravenously, contrast agents show the site of injury to the blood vessels by the appearance of the contrast agent on the scan. Iodine-containing contrast agents can cause anaphylactic reactions, however, particularly in individuals with allergy to shellfish. Contrast agents used in MRI studies are safer and more sensitive, making them the agents of choice. Gadolinium-containing compounds are used in MRI because they produce a paramagnetic effect. When they leak from the vessels into tissue, they cause a rapid relaxation of the protons that can be seen on T1-weighted images as a hyperintensity, compared to the precontrast scan.

Anatomical Sites of Central Nervous System Infection

The terminology used to describe various types of central nervous system (CNS) infections is anatomically based (Table 59.3). An infection limited to the subarachnoid space, with inflammation of the meninges, is called meningitis. Meningeal signs of headache, stiff neck, and photophobia are present without focal findings that would indicate spread into the parenchyma. When the infection spreads contiguously from the subarachnoid space through the pial surface or along Virchow-Robin spaces, crossing the gap-junctioned linings, the brain parenchyma is infected, and the term meningoencephalitis is used. In addition to meningeal signs, there are focal findings and possibly impaired consciousness and seizures. An infection in the brain tissue that is most likely spread via blood begins as a loose collection of invading cells referred to as a cerebritis; walling off of the infected brain tissue leads to an abscess. Finally, the term encephalitis is used to describe a more diffuse brain infection in both the gray and white matter, which is usually indicative of a viral infection. Occasionally the infection spreads in a potential space beneath the dura but outside the arachnoid; subdural empyema describes a life-threatening collection of pus over the brain surface that has often spread from an infected sinus through the venous plexus of the ethmoid or sphenoid sinuses into the subdural space. The presence of a subdural empyema should be suspected in a patient with sinus infection, fever, seizures, focal findings, and altered consciousness. Diagnosis of meningitis can be done by examination of CSF for signs of infection such as increased white blood cells or protein. Infections that invade the brain are best diagnosed with MRI, which can readily demonstrate a meningoencephalitis, cerebritis, abscess, or encephalitis. Use of contrast agents increases the potential of reaching a correct diagnosis based on site of infection. Subdural empyema is the most difficult condition to diagnose because it may only be a thin layer of pus on the surface of the brain and be obscured by the skull. Diagnosis can be missed on LP or CT.

Gap Junctions on Ependymal and Pial Surfaces

Lining the cerebral ventricles (other than over the choroid plexus) is a layer of ciliated ependymal cells connected by gap junctions. Pial cells lining the surface of the brain, which form the limiting glial membrane, the glial limitans, also have gap junctions. Fluid, electrolytes, and large protein molecules move through the gap junctions, allowing exchange between the CSF and ISF. Intrathecal administration of antibiotics and chemotherapeutic agents has been used to bypass the BBB.

Blood vessels penetrate the brain from the surface. As they enter the brain, they are invested with pia mater. The space between the penetrating blood vessels and the brain, prior to the point where only brain tissue surrounds the vessels, is called the Virchow-Robin space. After injection of substances intrathecally, the large proteins in the CSF space penetrate into the brain from the surface via the Virchow-Robin spaces. These perivascular routes may be involved in the spread of infection into the brain from the subarachnoid space in meningitis.

Arachnoid Granulations and Absorption of Cerebrospinal Fluid

Arachnoid granulations (pacchionian granulations) are the major sites for the drainage of CSF into the blood. They protrude through the dura into the superior sagittal sinus and act as one-way valves. As CSF pressure increases, more fluid is absorbed. When CSF pressure falls below a threshold value, the absorption of CSF ceases (Fig. 59.3). In this way, CSF pressure is maintained at a constant level, with the rate of CSF production as one determining factor.

Fig. 59.3 Schematic drawing of the relationship of cerebrospinal fluid (CSF) formation and absorption to pressure. CSF is formed at a constant rate of 0.35 mL/min. Absorption begins above a threshold value that varies from person to person. Once CSF absorption begins, it is linear, as seen in a one-way valve. When formation rate equals absorption rate, the steady-state CSF pressure is determined.

(Modified with permission from Cutler, R.W., Page, L., Galicich, J., et al., 1968. Formation and absorption of cerebrospinal fluid in man. Brain 91, 707-720.)

Although channels are seen in the arachnoid granulations, actual valves are absent. Tissue appears to collapse around the channel as the pressure falls, and the channels enlarge as pressure rises. Resistance to outflow across the arachnoid granulations leads to CSF pressure elevation. Substances can clog outflow channels and increase resistance to CSF absorption. Blood cells are trapped in the arachnoid villi, and subarachnoid hemorrhage causes a transient increase in CSF pressure and can occasionally lead to hydrocephalus. Similarly, white blood cells and increased protein from meningitis can block the arachnoid granulations and increase CSF pressure.

Cerebrospinal Fluid Pressure

Measurement of CSF pressure is a critical part of the LP. Pressures should be measured with the patient in the lateral recumbent position, and a narrow-bore spinal needle should be used to minimize CSF leakage. Performing the LP with the patient in the sitting position, although easier for the physician, eliminates the possibility of obtaining an accurate CSF pressure. Whenever CSF pressure is a critical piece of information, such as in the diagnosis of idiopathic intracranial hypertension, the sitting position should not be used.

The opening CSF pressure is measured with a manometer attached to the needle. Normal CSF pressure ranges from 80 to 180 mm H₂O but may go as high as 200 mm H₂O in obese patients or those who are not relaxed. Three components contribute to the measured pressure: volume of blood within the cranial cavity, amount of CSF, and the brain tissue. The CSF pressure recorded by the manometer represents the venous pressure transmitted from the right side of the heart through the venous sinuses. Small fluctuations from the cardiac systolic pulse and larger fluctuations from respirations can be seen in the column of fluid in the manometer. Pulsations in the manometer represent the fluctuations in the thin-walled veins (Davson, 1967). Arteries have thick elastic walls that dampen the pulsations from arteries. Deep respirations cause wide fluctuations in the CSF pressure, whereas changes in arterial pressure are barely visible. As ICP rises, tissue compliance falls and reserve capacity of the intracranial contents is lost. When tissue compliance is lost, small changes in fluid volume may lead to large increases in ICP.

Patients with increased ICP can be continuously monitored with indwelling catheters in the ventricles or with a sensor on the dura. Both procedures are invasive and only used in critically ill patients. Pathological elevations in ICP cause plateau waves that increase in steps to 50 mm Hg, where they persist for up to 20 minutes before returning to baseline. Treatment of patients with raised ICP can be monitored at the bedside with pressure monitors. Monitoring is used to gauge response to osmotic agents and to determine the severity of head injury. Treatment decisions can be made on the basis of the ICP.

In addition to complications of elevated CSF pressure, there are circumstances that lead to low pressure. Some patients have low pressure after LP owing to a persistent tear in the dura. Headaches occur with standing up and are often relieved with bedrest. Low-pressure headaches after LP generally resolve spontaneously. However, rarely they persist and require placement of a epidural blood-patch, which is accomplished by injecting the patient’s own blood to close the hole in the dura. Postsurgical and posttraumatic leakage of CSF can cause low-pressure headaches, and occasionally a spontaneous tear occurs. Diagnosis of a spontaneous leak can at times be difficult and may require injection of radiolabelled substances or contrast agents into the CSF (Schievink et al., 2008). Enhancement of the meninges can be seen on MRI in patients with low CSF pressure, but the cause of this abnormality is uncertain.

Composition of the Cerebrospinal Fluid