Table Of ContentAn Introduction to the Blood-Brain Barrier
An Introduction to
the Blood-Brain Barrier
HughDavson
Emeritus Professor,
Sherrington School of Physiology UMDS,
Guy' sand St Thomas' s Hospitals, London
Berislav ZIokovic
Associate Professor ofNeurosurgery,
Physiology and Biophysics,
USC School of Medicine, Los Angeles
Ljubisa Rakic
Professor of Neurobiology and Biochemistry,
School ofMedicine, Belgrade
and
Maleolm B. SegaI
Reader in Physiology,
Sherrington School ofPhysiology UMDS,
Guy' sand St Thomas' s Hospitals, London
M
MACMILLAN
© The authors 1993
Softcover reprint of the hardcover 1s t edition 1993
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First published 1993 by
THE MACMILLAN PRESS L TD
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andLondon
Companies and representatives
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ISBN 978-1-349-11884-7 ISBN 978-1-349-11882-3 (eBook)
DOI 10.1007/978-1-349-11882-3
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Contents
Preface vii
1. History and basic concepts 1
Classical experiments 1
The extracellular space of the brain 3
The cerebrospinal fluid 4
Permeability of the blood-brain barrier 11
Permeability: definition and measurement 14
Michaelis-Menten kinetics 23
Active transport 40
Application to the blood-brain barrier 56
The cerebrospinal fluid and the blood-CSF barrier 80
Transposition to the blood-brain barrier 101
Enzymatic contributions to the blood-brain barrier 109
Breakdown of the blood-brain barrier 110
Penetration of carrier-bound hormones 112
Penetration oflarge molecules 114
The broken-down barrier 121
Steady-state CSF concentrations 126
Complete morphology of the blood-brain barrier 128
2. Transport of glucose and amino acids in the central nervous
system 146
Glucose 146
Amino acids 170
3. Peptides and proteins 195
Peptides in the central nervous system 195
Functions of peptides in the brain 201
Peptide and protein interactions at the blood-brain barrier 218
Regulation of protein transport 242
Modulation of peptide transport 243
Enzymatic degradation of peptides 244
Peptide receptors in non-barrier regions 245
Therapeutic applications 249
Peptides and proteins in the CSF 253
4. Transport of some precursors of nucleotides and some
vitamins 273
Nucleotide precursors 273
Vitamins 285
v
vi Contents
5. Experimental models in the study of the pathology of the
blood-brain barrier 293
Introduetion 293
Amphetamine experimental psyehosis 294
Experimental allergie eneephalomyelitis 305
Cortieallesions 309
Subjea Index 323
Preface
The possibility of producing a short introduction to the physiology of the
blood-brain barrier was both discussed and agreed by the authors after a small
symposium on the subject had been held at the Serbian Academy of Arts and
Science in 1989. The present volume is the result; in it we have tried to present
lucidly the physical factors goveming transport from blood into the central nervous
tissue and the ccrebrospinal fluid, and then to describe some studies on specific
aspects, notably the transport of amino acids, sugars and peptides. Finally, in view
of the suspicion that some neurological diseases have as their basis a faHure or
impairment of the blood-brain barrier, we have included an account of some
attempts to establish animal preparations that might serve as experimental models
mimicking human pathology. If the reading of this book stimulates research on the
pathology of some central nervous diseases, it will have achieved its main purpose;
in addition, we trust that workers in the life sciences will find it a useful
introduction to a field that has expanded explosively since the early prejudices
against the concept of a blood-brain barrier were dispelled. We must conclude by
thanking the Wellcome Trust, the British Council and the Federal Yugoslav
Zavod for their financial help in promoting the co-operation between workers in
Britain and Yugoslavia that has culminated in the writing of this book.
June, 1991 H.D.
B.Z.
L.R.
M.B.S.
VIi
Chapter 1
History and Basic Concepts
Classical Experiments
Intravital Staining
The concept of the blood-brain barrier derives from the classical studies of the
pioneers in chemotherapy, such as Ehrlich, who administered dyestuffs parenter
ally in the hope that they would attack infective organisms. Thus Ehrlich observed
that many dyes, after intravenous injection, stained the tissues of practically the
whole body, while the brain was spared. Later, Lewandowsky (1900) showed that
the Prussian blue reagents (iron sah and potassium ferrocyanide) did not pass from
blood to brain, and he formulated clearly the concept of the blood-brain barrier
(Bluthirnschranke). The more definitive demonstration of the bafrier we owe to
Goldmann, who showed (1909) that, after intravenous injection with trypan blue,
the brain was unstained; the dye did not enter the cerebrospinal fluid (CSF),
although the choroid plexuses and meninges were stained. In a second paper
(Goldmann, 1913), he described experiments in which trypan blue was injected
into the CSF; in this event, the brain tissue was strongly stained, so that Goldmann
rightly concluded that there was, indeed, a barrier between blood, on the one
hand, and brain tissue on the other. Any argument that the failure to stain the
brain with trypan blue after intravenous injection was due to a peculiar staining
feature of the nervous tissue was negated by this fundamental 'second experiment',
the first experiment being the demonstration that nervous tissue was unstained
after intravenous injection.
Penetration ofOther Solutes
The blood-brain barrier, as initially revealed by studies on dyestuffs, was shown
not to be peculiar to these organic molecules, and, in general, it proved that
substances that usually failed to cross cell membranes also failed to cross the
blood-brain barrier (Krogh, 1946). Now, it was early demonstrated that penetra
tion into cells was govemed by the lipid-solubility of the compound being studied
measured by its oil-water partition coefficient,
concentration in oil
B=-------
concentration in water
Figure 1.1, for example, from the classical study ofCollander and Barlund (1933)
on single plant cells, shows that ease of penetration through the cell membrane,
measured by the molecule's permeability coefficient (p. 14), was directly related to
1
2 An Introduction to the Blood-Brain Barrier
10
!
Water
!
Water
• MRo < 15
® MRo 15-2
({g) MRo 22-3
~ MRo >30
0.0001 0.001 0.01 0.1
Figure 1.1 Permeability of Chara cells plotted against oiVwater partition coefficient.
Ordinates: permeability in cm/h XMI/2. Abscissae: oiVwater partition coefficient. From
Collander, Physiol. Plant. (1949)
its partition coefficient, relatively lipid-insoluble substances, such as sucrose and
glycerol, with partition coefficients in the region of 1 x 10-4-1 X 10-5, barely, if at
all, penetrating cells; when the partition coefficient was in the region of 0.01, there
was significant penetration, as with thiourea; with a partition coefficient of, say,
0.1, as with ethyl alcohol, penetration was very rapid. Early semiquantitative
studies of passage of solutes into the brain from blood showed that the barrier was
high for substances such as sucrose or mannitol but low for substances such as
ethyl alcohol or propyl thiourea. Thus, as Krogh emphasized in 1946, the
capillaries of the brain, or other lining membranes separating blood from the brain
tissue, behaved like single cells, in marked contrast to the capillaries of most other
tissues of the body, where it is found that passage from blood to the extracellular
space is rapid and virtually independent of the partition coefficient.
Early Objections to the Concept
If we look back on the subject now over the period, say, 1920 to 1960, it becomes
evident that Goldmann's second experiment was ignored, and it was repeatedly
argued that the blood-brain barrier, as described by Goldmann's first experiment,
History and Basic Concepts 3
was an artefact resulting from the failure of the dyestuff to be taken up by the
tissue after leaving the blood vessels. The strongest argument adduced in support
of this position was the appearance of the brain in electron-microscopical sections;
previously the histology of the brain had been examined by two basic procedures
that stained either the neurons (Golgi technique) or the glia. Pictures obtained by
either technique left room for the assumption that the nervous tissue had a
considerable extracellular space, comparable with that of other tissues. However,
the osmium-stained preparations of the electron microscope revealed both glia
and neurons, so tightly packed that it was argued that the extracellular space was
negligible, so that if trypan blue and other acidic dyestuffs passed out of the
capillaries, the staining would not be intense enough for observation in the
light-microscope.
There is no need to recapitulate in detail the experimental studies that these
claims stimulated. Since it was apparently insufficient to emphasize Goldmann's
second experiment and its significance, more direct experimental proof of a barrier
between blood, on the one hand, and the extracellular fluid of the central nervous
system, on the other, was necessary. First, it was important to determine the actual
size of the extracellular space; although the electron-microscopical studies
suggested that this would be very smalI, compared with that in muscle and other
connective tissues, nevertheless it must have an experimentally determinable
magnitude.
The Extracellular Space of the Brain
Experimental Measurement
The basis for determination of the extracellular space of a tissue, such as muscle,
has been the measurement of the concentrations of an 'extracellular marker' in the
tissue and in the medium surrounding the cells when the system is in diffusional
equilibrium with the marker. If the experiment is performed in vivo, the
concentration in the extracellular fluid may be equated with that in the blood
plasma, provided that there is readyequilibration between the two fluids. If the
experiment is carried out in vitro, then the concentration in the bathing medium
will be compared with that in the tissue. The requirement of the marker is that it
should not penetrate cells, being confined to the extracellular space. In this case
the extracellular space is given by:
concentration in tissue (mmoVg-tissue)
------------------~----~--~----xl00
concentration in plasma-water or saline (mmoVg-HzO)
With the in vivo experiment some of the marker will remain in the blood vessels,
but allowance for this may be made by measuring the 'blood-space' by infusing
labelIed red cells.
