Table Of ContentBiolog y an d
Patholog y o f
Nerv e Growt h
Ç . ME I LI U
Division of Biology and Medicine
Brown University and Miriam Hospital
Providence, Rhode Island
1981
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Liu, H.M ei.
Biolog y an d patholog y of nerv e growth .
Bibliography : p.
Include s index .
1. Nerves—Growth . 2. Developmenta l neurology .
3. Nervou s system—Regeneration . 4 . Nerves—Wounds
and injuries . I . Title . [DNLM: 1. Nervou s system —
Pathology . WL1 0 1L 783b ]
QP363.5 L58 612'.8 1 81-363 0
ISBN 0-12-452960- 7 AACR2
PRINTE D IN THE UNITE D STATE S OF AMERIC A
81 82 83 84 9 8 7 6 5 4 3 2 1
To the memory of my parents,
Liu Shan-Tung and Hsia Shee-Chou,
for the wisdom, the encouragement, and
dreams they have bestowed upon me.
Prefac e
Before discussin g the goal of this book, I shall first give an accoun t of my
philosophica l positio n in writin g it. I am takin g fundamentalis t and inter -
disciplinaris t point s of view. My main aim is to presen t a coheren t pictur e of
the basic pathophysiologica l chain reaction s that sustai n the life of the nervou s
system .
The immens e complexitie s of the nervou s syste m in the highe r animal s have
necessitate d the division of neuroscienc e into subdisciplines . Investigator s tend
to develop a one-side d viewpoin t dependin g on the kinds of tools or approache s
they use. The resul t is that we are confronte d with an enormou s amoun t of data
that are often disjointe d and at times contradictory . Never have we had so much
informatio n on somethin g we understan d so little . We have lost sight of the fact
that the nervou s system , despit e its complexities , exists as a single continuou s
entit y and that ther e is intens e interdependenc e and interactio n betwee n neura l
and non-neura l cells. The behaviora l pattern s of the nervou s tissu e carr y an in-
terna l logic, which is determine d by geneti c and environmenta l factors , and as
such can be understoo d only by examinin g the nervou s syste m from the earlies t
beginnin g to the time of demis e of the cells.
In this book, I set myself an immens e task of delvin g into what is known of the
biochemica l basis of cell interactio n in general , and neuron-gli a interactio n in
particular . How do cells presen t themselve s to their environmen t and become
determine d in return ? What are the molecula r mechanism s of nerve growth and
maturation ? How is informatio n receive d and transmitted , and throug h what
kind of networ k do defensiv e and reparativ e activitie s take place? Using data
derive d from embryological , tissu e cultural , chemical , histological , and
regenerativ e studies , a unifie d concept of neura l growth , behavior , and
organizatio n is beginnin g to emerge . I believe we can take a new look at the
ix
÷ PREFAC E
whole pictur e becaus e remarkabl e new information , some of which may seem to
be unrelate d to neuroscienc e at a superficia l glance , permit s new, far-reachin g
insights .
The book begins with a brief discussio n of the embryologica l development .
This is a prerequisit e to the understandin g of specific chemica l labels acquire d
by neuron s and supportin g cells as a resul t of differentiation . The cell labels that
mediat e cell recognitio n and interactio n are thus criticall y determine d by
cellula r and molecula r event s that take place before the formatio n of the nervou s
system . The second and thir d chapter s examin e the proces s of neurona l matura -
tion, myelination , and integratio n with targe t tissues . The controversia l issue s of
guidanc e and the natur e of the factor s that determin e the matchin g of nerve
with muscl e are explore d from morphologica l and biochemica l point s of view.
The second half of the book center s on the issue of reactio n to injury , and it is
precede d by a discussio n of the basic principl e of regeneratio n as seen in lower
animals . Many lines of scientifi c evidenc e have shown that despit e our unique -
ness in body and mind , huma n beings , at the basic level of existence , have not
evolved very far from our lower ancestors . These trait s havin g thus arisen , per-
sist into highe r forms on accoun t of their economy , simplicity , and surviva l
value . I am attemptin g to show that two of the centra l issue s of neuroscience ,
namel y neura l organizatio n (which is controlle d by cell interaction ) and reactio n
to injur y (which is necessar y for survival) , are interrelate d at the cellula r and
molecula r levels; both operat e on the same biophysica l principle s across all
kinds of anima l and plan t species .
The centra l purpos e of this book is to presen t a them e of biologica l coherenc e
and continuity : thus , the emphasi s is on the complet e picture . Individua l factor s
are considere d to the exten t that we can understan d the entir e process , in order
to see how the component s are designe d to work togethe r in a marvelou s way for
the benefit of the total being. I have purposel y avoide d the telegraphi c style of
citatio n of references , but relevan t histor y on variou s topics will be briefly
reviewed .
The phenomen a of nerve growth , regeneration , and disease s as presente d in
this book do not differ significantl y from those alread y known . But the reade r
will find in man y area s my own interpretation , which may differ radicall y from
some currentl y accepte d and taugh t views. These unconventiona l views are the
outgrowt h of attempt s at revealin g the rationa l continuit y that is built into the
biological systems . It is probabl y true that "every major progres s is achieve d by
the overthro w of dogma " (Lewis Thomas) . I hope that this new approac h may
provide an initia l step in bringin g new and interestin g developmen t to the field
of neuroscience .
H. Mei Liu
Acknowledgmen t
I am indebte d to my teacher s in pathology , Dr. Milton Bohrod and Dr. Edit h
Potter , who conveyed to me their enthusiasm , their libera l approac h towar d in-
vestigation , and the resolutio n to be accurat e in presentin g facts and to be
forever critica l of onesel f and others . I wish to expres s gratitud e to Dr. Sumne r
Zacks for providin g the kind of environmen t in his departmen t which made the
writin g of this book possible . I owe man y thank s to my colleague , Dr. Michae l
Sheff, for hour s of delightfu l discussio n on chemistr y and cuisine . Dr. Richar d
Goss has been most helpfu l in providin g informatio n relatin g to regeneration .
The histologica l and electro n microscop e preparation s were the collective efforts
of man y technologist s with whom I have worked throughou t the years , but I
would like to give specia l thank s to Mrs. Laila Ovadi a for the hundred s of seria l
section s of chick embryo s and salamande r arm s which she patientl y prepared , to
Mrs. Sand y Kunt z whose expertis e in scannin g electro n microscop y has given the
illustration s a specia l artisti c flavor, and to Ms. Kath y Moos for spendin g man y
hours typin g the manuscript .
Specia l thank s also go to Dr. William Windle , who has provide d continuou s
advice and encouragement . Finally , this monograp h and recen t relate d researc h
work were made possibl e by a gran t from the Rhode Islan d Foundation .
xi
1
Embryologica l
Developmen t
What enable s the nervou s syste m to perfor m such fantasti c tasks ? The power
lies in the uniqu e propertie s of each individua l neuro n and in the complex but
orderl y connection s formed by million s of neuron s amon g themselve s and with
the targe t tissues . Becaus e of the intensiv e studie s undertake n durin g recen t
years , we are beginnin g to get a glimps e of the intricat e phenomen a of the
developmen t and architectura l design of the nervou s syste m in variou s species of
animals .
Scientist s involved in the studie s of neuropatholog y and neura l regeneratio n
usuall y show no interes t in embryology , partl y becaus e of its complexity . It is im-
portan t to realiz e that the embryoni c origin of a given cell tells somethin g of its
later behavio r and is therefor e of far reachin g significance . The close
parallelis m betwee n embryolog y and regeneratio n make s it necessar y for those
intereste d in nerve regeneratio n to have some basic understandin g about the
origin and developmenta l proces s of cells in the nervou s syste m and of those
associate d with them . It is even more apparen t that knowledg e of neuro -
embryolog y is a prerequisit e to the understandin g of developmenta l neuro -
pathology . As regard s developmenta l neurobiolog y per se, the curren t unsettle d
issues concernin g the guidin g factor s of nerve connection , the origin of sheat h
cells, and their allege d relationshi p to the neura l crest will continu e to be
shroude d in myster y unles s one take s a panorami c view of the life histor y of the
organis m as a whole.
1
2 1. EMBRYOLOGICA L DEVELOPMEN T
Becaus e of the obstacle s in dealin g with the nervou s syste m of highe r animals ,
most of the embryologica l studie s have been carrie d out in amphibian s and
avian species . A large volume of informatio n has been accumulate d durin g the
past centur y with regar d to the morphogeneti c steps of embryoni c develop -
ment . Experimenta l embryologists , usin g delicat e microsurgery , have produce d
amazin g result s in isolatio n and combinatio n of embryoni c parts . These pionee r
experimentalist s have contribute d immensel y to our understandin g of the
dynami c natur e of the embryologica l process .
The inventio n of the tissu e cultur e techniqu e by Harriso n has opene d a new
dimensio n to the biologica l investigators . It reduce s the complex nervou s syste m
to the level wher e a limite d numbe r of neuron s may be observe d while alive and
unde r controlle d conditions . The basic knowledg e of growth behavio r of
neurons , their interrelatio n with the sheat h cells, and their respons e to humora l
growth and inhibitor y factor s have been elucidated .
Despit e the enormou s advantag e offered by the tissu e cultur e method , one
must at the same time be awar e of its limitations . The major setbac k is that cells
isolate d from the body do not behav e in exactl y the same manne r as they do in
vivo. Neuron s in cultur e will aggregat e and grow; they respon d to growth factor s
and inhibitors ; they even form synapse s and become myelinated . However , ther e
is one thin g that neuron s fail to do in culture , that is to form a functiona l brai n
or spina l cord. The ingredien t missin g in tissu e cultur e is that mysteriou s force
called organization ; withou t it, neuron s would scatte r about like a pile of bricks
rathe r than a house . We perceiv e the presenc e of an organizin g force indirectl y
by the marvelousl y consisten t patter n of morphogeneti c steps observe d durin g
embryologica l study , but still the force elude s us as it has generation s of in-
vestigators .
In the following chapter , we shall examin e the bares t minimu m of the essentia l
facts about the developmenta l histor y of the early embry o and the histogeneti c
change s of the variou s cellula r element s of the nervou s system , to make specula -
tions and explor e alternativ e explanation s which could make the conflictin g da*ta
somewha t more plausible . We are more intereste d in integratin g the man y facets
of the known developmenta l steps rathe r than the detail s and differences . The
use and limitation s of the tissu e cultur e method s and pitfall s of some of the ex-
perimenta l procedure s used in the past will be criticall y examined . Hopefully ,
this will provid e us with the necessar y backgroun d for later discussion .
In view of the confusio n which may arise as a resul t of inconsistenc y in ter-
minology regardin g the nonneura l cells in the nervou s system , the definitio n
listed below describe s the use of words in this book.
Glial cells: A genera l term for supportin g cells in the nervou s system ; the cen-
tral glia includ e astrocytes , oligodendrocytes , and microglia , and the periphera l
glia includ e neurolemm a cells, endoneura l and perineura l fibroblasts .
I. THE EARL Y EMBRY O 3
Satellite cells: Cells that surroun d the perikaryo n of neuron s in the centra l
nervou s syste m and in periphera l nerve ganglia .
Neurolemma cells: Cells that surroun d axons of periphera l nerve s and form
myelin sheaths ; they are known as Schwan n cells or sheat h cells.
I. THE EARL Y EMBRY O
A. Th e Ovu m
At the time of fertilization , the ovum appear s deceptivel y simple . The
cytoplas m contain s the usua l set of organelle s as well as variou s amount s of yolk
materia l dependin g on the species . Except for its large r size ther e is hardl y
anythin g eithe r morphologicall y or chemicall y that could distinguis h an ovum
from other cells, yet this cell has the potentia l of giving rise to all of the tissue s in
the body, hence the term ''totipotent. " It is to be distinguishe d from "multi -
potent " which mean s that the cell can give rise to man y but not all kinds of cells.
Even before fertilization , importan t developmenta l change s are takin g place
in the ovum. In the frog egg, the cytoplasmi c component s underg o a period of
shiftin g which leads to asymmetrica l distributio n of the cytoplasmi c material ,
and the egg is said to be polarized . The anima l pole contain s a highe r concentra -
tion of RNA and glycogen , while the vegeta l pole contain s a high concentratio n
of yolk material . The poles of the eggs are distinguishabl e grossl y by the pig-
ment patter n which is characteristi c for each species . In the frog egg, the anima l
pole is heavil y pigmented , and the vegeta l pole is light in color. The sea urchi n
egg has a band of light pigmen t encirclin g an area just beneat h the equato r
towar d the vegeta l pole (Chen , 1967; Eber t and Sussex , 1970; Graha m and Ware -
ing, 1976). The regiona l differenc e of the egg is the reaso n why blastomere s or
daughte r cells arisin g from one pole of the egg are qualitativel y differen t from
those at the other pole after the first few divisions .
The mechanis m which bring s about the establishmen t of the polarit y of the
egg can be trace d to externa l factors . We learne d that the polarit y of the
seawee d Fucus egg may be influence d by unilatera l light and pH in the medium ;
the eggs germinat e towar d the acidic side and away from the light source (see
p. 156). When man y eggs are placed together , germinatio n occurs on the side
facing the other eggs (Fig. 1). Evidently , a diffusabl e chemica l substanc e pro-
duced by the Fucus egg is responsibl e for causin g the cytoplasmi c streamin g
and the establishmen t of polarity . The ova of birds and mammal s are protecte d
from sunligh t and seawate r by the oviduct . The factor s which influenc e the
polarit y chang e may be postulate d to exist in the chemica l substance s secrete d
by the mucosa l linin g of the oviduct .
4 1. EMBRYOLOGICA L DEVELOPMEN T
Fig. 1. Germinatio n of a grou p of seawee d Fucus eggs showin g orientatio n towar d one
another—grou p effect .
B. Earl y Division s
At the outset , the ovum is marke d by invisibl e zones, each destine d to form a
specialize d tissue . Earl y embryologist s have take n advantag e of the fact that sea
urchi n and amphibia n eggs have characteristi c pattern s of pigmentatio n which
serve to differentiat e the poles of the egg. In addition , vital dyes have been used
to mark specific region s of the egg so that the fate and final location of the
embryoni c cells could be trace d in the later stages .
Durin g fertilizatio n of the sea urchi n egg, the sperm enter s the anima l pole
where pigmen t is present . The plan e of the first division is vertica l and run s
throug h the animo-vegeta l axis (Fig. 2). The blastomere s or daughte r cells are,
therefore , quantitativel y and qualitativel y equal . The second division follows the
same plan e as the first division ; each of the four blastomere s acquire s the same
nuclea r and cytoplasmi c constituents . When isolate d at this stage , each cell is
Fig. 2. Earl y division s of frog egg. The first and secon d division s are vertica l and run throug h
the animo-vegeta l axis , the thir d divisio n is horizonta l and result s in eigh t cells , four at the anima l
pole and four at the vegeta l pole .
