Cerebrospinal Fluid, Blood-Brain Barrier and Blood-CSF Barrier_

Formation and function of the cerebrospinal fluid

Liquor (cerebrospinal fluid – CSF, liquor cerebrospinalis) is a clear, colourless fluid found in CNS
either intracerebrally in the ventricular system of the brain (making up 20 % of the total CSF
volume) or extracerebrally in the subarachnoid space (the remaining 80 % of the total volume).

The total volume of cerebrospinal fluid is approximately 150 ml and it is produced at a rate of
450 ml per day (thus replacing itself three times a day). Liquor is synthesized through two
processes. Secretion (producing around 50 to 70% of the volume) occurs in the cells of the
choroid plexus and ventricular ependyma. The rest of the fluid is synthesized by the
ultrafiltration of blood plasma through choroidal capillaries.

CSF circulates from the lateral ventricles through interventricular foramina (foramina of Monro)
into the third ventricle. Mesencephalic duct (aqueduct of Sylvius) connects the third and fourth
ventricle and the liquor flows further into the subarachnoid space by means of three foramina:
single median aperture (foramen of Magendie) and a pair of lateral apertures (foramen of
Luschka). Liquor located in the subarachnoid space surrounds the brain and spinal cord. Portion
of liquor leaves the fourth ventricle to enter the central canal of the spinal cord.

The resorption of liquor takes place in the arachnoid granulations (arachnoid villi, Pacchioni’s
granulations). These are the protrusions of the arachnoid through the dura mater, projecting
further into the intracranial venous sinuses enabling the flow of liquor back to the blood
circulation.

It is important to maintain a continuous equilibrium between the secretion, circulation and

resorption of the CSF. In the case of imbalance, the fluid accumulates in the system causing a
condition called hydrocephalus. If the CSF accumulates in the ventricular system, the
hydrocephalus is termed internal. In the case of its accumulation in the subarachnoid space, we
call the hydrocephalus external.
CSF performs the following functions:

1) Mechanical and supportive

The brain is essentially fully immersed in the cerebrospinal liquid causing a reduction of is real
weight (around 1500g) to an equivalent of about 25 g. This mechanism protects the brain
against the damage caused by its own weight.

2) Protective

CSF acts as a shock absorber, protects the brain from a sudden pressure or temperature
changes and the components of the immune system present in the fluid (leukocytes,
immunoglobulins etc.) provide a protection against various pathogens as well.

3) Metabolic

Liquor helps to maintain the correct composition of the environment surrounding nervous
tissue cells (homeostasis). It also partially provides the supply of nutrients and disposal of the
metabolic waste products and forms a medium through which a diffusion of a various signal
molecules (like neurotransmitters) takes place.

It is important to realize that the volume of all tissues and fluids within the skull is limited by
the volume of their bone container. Monroe-Kelly’s doctrine says that the intracranial space
volume remains constant and its individual components (blood, cerebrospinal fluid and the
nervous tissue) exist in a state of volume equilibrium. Any increase in the volume of one of the
component (caused for example by an intracranial bleeding, hydrocephalus or tumor growth) is
compensated by a reduction in the volume of other components. CSF acts within this system
partially as a buffer (though not having a large capacity) – it is the first system to compensate
the increase in volume of the other two compartments. Brain circulation can act in a similar
way but to a much lesser extent, because a restriction in blood supply increases a risk of a
hypoxic damage to the brain tissue.
Composition of the cerebrospinal fluid

1) Biochemistry: ions, glucose, lactate, proteins including ELFO

2) Cytology

3) Microbiology: cultivation, possibly PCR

4) One backup sample (stored at 4 °C)

During the sample collection we can measure the pressure of the cerebrospinal fluid as well.
The value in the case of lumbar puncture in patient lying on side is approximately 8-15 mmHg
(10-18 cm H2O) or in the case of sitting patient around 16-24 mmHg (20-30 cm H2O).

Physiological findings are summarized in the following tables:

1) Ions
pH
7.28-7.32

Osmolarity
285 mosm/l

Na+ K+ Ca2+ Cl- etc.

2) Nutrients and proteins

Glucose (glycorrhachia)
Proteins (proteinorrhachia)
(small proteins mostly, immunoglobulins – IgG, IgM)
Lactate
Urea
Lipids

3) Cells

Erythrocytes
0 / mm3
Lymphocytes
0-5 / mm3

Bacteria
0 / mm3

Blood-brain barrier and blood-CSF barrier

Blood-brain barrier (BBB, hematoencephalic barrier – HEB)

The term blood-brain barrier denotes the barrier separating the brain tissue from the blood
circulation. The structural basis consists of three parts:

1) Layer of endothelial cells interconnected through tight junctions and not containing
fenestrations (normally present in endothelium of other tissues)

2) Basal membrane consisting of basal lamina of astrocytes and basal lamina of endothelial cells

3) Protrusions of astrocytes – so-called pedicles, which anneal on the basal membrane

The main function of BBB is the protection of CNS tissue (especially neurons) against different
noxious substances, which would be otherwise able to penetrate to the CNS. The selectivity of
BBB prevents many harmful substances from passing into the CNS, but on the other hand
allows other (necessary for the brain function) to enter. Generally, hydrophilic substances do
not pass BBB until the endothelial cells and astrocytes express appropriate transport channels.
The hydrophobic nature of BBB on the other hand allows the penetration of lipophilic
substances.

1) Freely, by a passive diffusion, penetrate the BBB:

a) Small molecules: H2O, O2, CO2, NH3, ethanol

b) Lipophilic substances: steroid hormones

2) Selective transporters (facilitated diffusion or active transport) are typical for:

a) Glucose: GLUT-1

b) Amino acids

3) Some of the macromolecules use pinocytosis to pass through BBB

The presence of BBB has some significant clinical implications. Apart form being important
protection mechanisms against microorganisms, reducing the risk of CNS infection, BBB also
prevents the passage of antibodies and antibiotics, which can seriously impede body’s ability to
fight an infection. Beside the above- mentioned antibiotics, there exist a number of other drugs
and substances, which are unable to pass the BBB. One of the examples is dopamine, whose
insufficiency is characteristic of a Parkinson’s disease. We cannot use dopamine to treat the
disease, as it will not get to the basal ganglia (where the shortage occurs). Instead we have to
use its precursor L-DOPA, which penetrates BBB and turns into dopamine in the brain tissue.

Encephalopathy caused by an unconjugated hyperbilirubinemia (called kernicterus) may

develop in newborns due to underdeveloped blood-brain barrier that does not prevent the
bilirubin from entering the brain tissue. It is however very rare in adults with fully developed
BBB.

Some regions of the brain do not have the protection of BBB. These are the chemoreceptor
zones with an important function of controlling the composition of blood. Examples include:

1) Subfornical organ: contains osmoreceptors regulating the secretion of ADH

2) Organum vasculosum laminae terminalis (OVLT): contains osmoreceptors regulating the

secretion of ADH and triggers the feeling of thirst

3) Area postrema: contains chemoreceptors, which are in control of the vomiting

Blood-brain Barrier Maintains the Constancy of the Brain's Internal Environment

The blood-brain barrier (BBB) protects the neural tissue from variations in blood composition
and toxins. Elsewhere in the body the extracellular concentrations of hormones, amino acids
and potassium undergo frequent fluctuations, especially after meals, exercise or stressful
times. Since many of these molecules regulate neuronal excitability, a similar change in the
composition of interstitial fluid in the CNS can lead to uncontrolled brain activity. The
endothelial cells forming the blood-brain barrier are highly specialized to allow precise
control over the substances that enter or leave the brain.

Discovery of blood-brain barrier. The discovery of the BBB dates back more than 100 years
when, in the 1880s, Paul Ehrlich observed that intravenous administration of certain dyes (e.g.
trypan blue) stained all organs except the brain and the spinal cord. He concluded that the
dyes had a lower affinity for binding to the nervous system as compared to other tissues. In
1913, Edwin Goldman, an associate of Ehrlich, demonstrated the very same dyes, when
directly injected into the cerebrospinal fluid (CSF), readily stained nervous tissue but not
other tissues. The term “blood-brain barrier” was coined, however, by Lewandowsky in 1898,
after he and his colleagues had performed experiments to demonstrate that neurotoxic
agents affected brain function only when directly injected into the brain but not when
injected into the vascular system. It took an additional 70 years until Reese and colleagues
localized the barrier to the capillary endothelial cells within the brain by electron-microscopic
studies.

All areas of the brain do not have a blood-brain barrier. The structures located at strategic
positions in the midline of the ventricular system and lack the BBB are collectively referred to
as circumventricular organs (CVOs). In these non-barrier regions, the tight junctions between
endothelial cells are discontinuous thus allowing entry of molecules. Many of these areas
participate in hormonal control.

Areas of brain without a blood-brain barrier:

Pituitary gland
Median eminence
Area postrema
Preoptic recess
Paraphysis
Pineal gland
Endothelium of choroid plexus
Mitosis, a process of cell duplication, or reproduction, during which one cell gives rise to two
genetically identical daughter cells. Strictly applied, the term mitosis is used to describe the
duplication and distribution of chromosomes, the structures that carry the genetic information.

Mitosis has four stages: prophase, metaphase, anaphase, and telophase.

Prior to the onset of mitosis, the chromosomes have replicated and the proteins that will form
the mitotic spindle have been synthesized. Mitosis begins at prophase with the thickening and
coiling of the chromosomes. The nucleolus, a rounded structure, shrinks and disappears. The
end of prophase is marked by the beginning of the organization of a group of fibres to form a
spindle and the disintegration of the nuclear membrane.

The chromosomes, each of which is a double structure consisting of duplicate chromatids, line
up along the midline of the cell at metaphase. In anaphase each chromatid pair separates into
two identical chromosomes that are pulled to opposite ends of the cell by the spindle fibres.
During telophase, the chromosomes begin to decondense, the spindle breaks down, and the
nuclear membranes and nucleoli re-form. The cytoplasm of the mother cell divides to form two
daughter cells, each containing the same number and kind of chromosomes as the mother cell.
The stage, or phase, after the completion of mitosis is called interphase.

Mitosis is absolutely essential to life because it provides new cells for growth and for
replacement of worn-out cells. Mitosis may take minutes or hours, depending upon the kind of
cells and species of organisms. It is influenced by time of day, temperature, and chemicals.
Spermatogenesis is initiated in the male testis with the beginning of puberty. This
comprises the entire development of the spermatogonia (former primordial germ cells) up to
sperm cells. The gonadal cords that are solid up till then in the juvenile testis develop a lumen
with the start of puberty. They then gradually transform themselves into spermatic canals that
eventually reach a length of roughly 50-60 cm. They are termed convoluted seminiferous
tubules (Tubuli seminiferi contorti) and are so numerous and thin that in an adult male testicle
their collective length can be 300 to 350 meters. They are coated by a germinal epithelium that
exhibits two differing cell populations: some are sustentacular cells (= Sertoli's cells) and the
great majority are the germ cells in various stages of division and differentiation.

The development of the germ cells begins with the spermatogonia at the periphery of the
seminal canal and advances towards the lumen over spermatocytes I (primary spermatocytes),
spermatocytes II (secondary spermatocytes), spermatids and finally to mature sperm cells.

Developmental stages of spermatogenesis

In the course of spermatogenesis the germ cells move towards the lumen as they mature. The
following developmental stages are thereby passed through:

A-spermatogonium

B-spermatogonium

Primary spermatocyte (= spermatocyte order I)

Secondary spermatocyte (= spermatocyte order II)

Spermatid

Sperm cell (= spermatozoon)

The spermatogenesis can be subdivided into two successive sections:

The first comprises the cells from the spermatogonium up to and including the secondary
spermatocyte and is termed spermatocytogenesis.

The second one comprises the differentiation/maturation of the sperm cell, starting with the
spermatid phase and is termed spermiogenesis (or spermiohistogenesis).

The temporal course of spermatogenesis

The approximate 64 day cycle of the spermatogenesis can be subdivided into four phases that
last differing lengths of time: