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THE GERM-CELL CYCLE IN ANIMALS

THE MACMILLAN COMPANY

NEW YORK • BOSTON • CHICAGO - DALLAS
ATLANTA • SAN FRANCISCO

MACMILLAN & CO., Limited

LONDON • BOMBAY • CALCUTTA
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TORONTO

THE GERM-CELL CYCLK
IN ANIMALS

ROBERT W. HEGNER, Ph.D.

ASSIST.AJNT PROFESSOR OF ZOOLOGY IN THE UNIVERSTTY

OF IVnCHIGAN

AUTHOR OF "an IXTRODUCTIOX TO ZOOLOGY
AND "college zoology"

Neb) gorfe

THE MACMILLAN COjMPANY

1914

Set up and elcctrotyped. Published September, 1914.

J. S. Gushing Co. — Berwick & Smith Co.
Norwood, Mass,, U.S.A.

PREFACE

This book is the result of a course of lectures
delivered during the past school year before a class
in Cellular Biology at the University of INIichigan.
Many of the most important recent additions to
our knowledge of heredity have resulted from, the
study of the germ cells, especially those of animals.
This study is now recognized as one of the chief
methods of attacking certain problems in genetics
and must be employed in correlation with animal
breeding before we can hope to obtain an adequate
explanation of the results of hybridization. For-
tunately the cytological studies of the germ cells,
both observational and experimental, have kept
pace with the rapid advances in our knowledge of
plant and animal breeding which have been made
since the rediscovery of Mendel's investigations in
1900. The term "Germ-Cell Cycle" is meant to
include all those phenomena concerned with the ori-
gin and history of the germ cells from one genera-
tion to the next generation. The writer has, with
few exceptions, limited himself to a consideration of
the germ cells in animals because the cycle is here
more definite and better known than in plants.

It is obvious to any one familiar with this subject
that only a few of the many interesting phases of

1161S2

vi PREFACE

the problems involved can be considered in a work
of this size, and those for which space can be found
must be limited in their treatment. For this reason
some periods in the germ-cell cycle are only briefly
mentioned, whereas others are more fully discussed.
The latter are naturally those in which the writer
is most interested and with which he is best ac-
quainted. Furthermore, the attempt is made to
present the data available in such a way as to make
it intelligible to those who have not been able to
follow in detail the progress of cytology during the
past few years. This can only be accomplished by
introducing many facts that are well known to
cytologists and zoologists in general, but are neces-
sary for the presentation of a complete account of
the subject.

Much of the recent cytological work done on germ
cells has emphasized the events which take place
during the maturation of the eggs and spermatozoa,
that is, the periods of oogenesis and spermatogenesis.
These are, of course, very important phases of the
germ-cell cycle, but they should not be allowed to
overshadow the rest of the history of the germ cells.
Contrary to the usual custom, the period that is
emphasized in this book is not the maturation of
the germ cells, but the segregation of the germ cells
in the developing egg and the visible substances
(keimbahn-determinants) concerned in this process.

It has been impossible to include in this book as
much illustrative material as desirable, but the bib-
liography appended indicates what data exist and

PREFACE vii

where they may be obtained. This list of publica-
tions has been arranged according to the method
now in general use among zoologists; the author's
name and the date of the appearance of the contri-
bution in question are bracketed in the text wher-
ever it has been considered necessary, and reference
to the list at the end of the book will reveal the
full title and place of publication of the work, thus
avoiding cumbersome footnotes. The figures that
have been copied or redrawn are likewise referred
in every case to the original source. Many of them
have been taken from the writer's previous publica-
tions and a few have been made especially for this
work. The writer has likewise drawn freely upon the
text of his original investigations already published.

Ann Arbor, Michigan,
April 16, 1914.

TABLE OF CONTENTS

CHAPTER I

PACK

Introduction j

The Cell, 2 ; Cell Division, 13 ; Methods of Repro-
duction, 17; The Germ Cells, 19; The Life Cycles of
Animals, 22.

CHAPTER II

General Account of the Germ-cell Cycle in Ani-
mals C25

Protozoa, 25 ; Metazoa, 28.

CHAPTER III

The Germ-cell Cycle in the P^dogenetic Fly,

Miastor 51

CHAPTER IV

The Segregation of the Germ Cells in Sponges, Cce-

lenterates, and vertebrates . . . c9

1. Porifera, 69. 2. Coelenterata, 80. 3. Verte-
brata, 98.

CHAPTER V

The Segregation of the Germ Cells in the Artiiro-

PODA lOG

1. The Keimbahn in the Insects, 106 ; Diptera, 107 ;
Coleoptera, 109 (In Chrysomelid Beetles, 109; Origin
of Nurse Cells, 119; Cyst Formation in Testis, 125;
Amitosis, 133; Differentiation of Nuclei in Egg, 141);
Hymenoptera, 143. 2. The Keimbalui in tlie Crusta-
cea, 163.

X TABLE OF CONTENTS

CHAPTER VI

PAGE

The Segregation of the Germ Cells in Nematodes,

Sagitta, and Other Metazoa . . . 174i

1. The Keimbahn in the Nematodes, 174. 2. The
Keimbahn in Sagitta, 179. 3. The Keimbalin in Other
Animals, 183.

CHAPTER Vn

The Germ Cells of Hermaphroditic Animals . . 189

CHAPTER VIII

Keimbahn-Determinants and their Significance . 211

A. The Genesis of the Keimbahn-Determinants,
211 (a. Nuclear, 213 ; b. Cytoplasmic, etc., 224 ; c. Dis-
cussion, 228) . B. The Localization of the Keimbahn-
Determinants, 235. C. The Fate of the Keimbahn-
Determinants, 240.

CHAPTER IX

The Chromosomes and Mitochondria of Germ Cells 245
The Chromosome Cycle in Animals, 245. The Mito-
chondria of Germ Cells, 275.

CHAPTER X

The Germ-plasm Theory 290

References to Literature . . . . . .311

Index of Authors ......•• 337

Index of Subjects 341

THE GERM-CELL CYCLE IN ANIMALS

GERM-CELL CYCLE IN ANIMALS

CHAPTER I

INTRODUCTION

Since the enunciation by Harvey of the aphorism
Omne vivum ex ovo in the seventeenth century, the
statement has frequently been made that every
animal begins its individual existence as an egg.
While this is not strictly true, since no eggs occur in
the life history of many one-celled animals (Pro-
tozoa), and a large number of multicellular animals
(Metazoa) are known to develop from buds or by
fission, still the majority of animals arise from a single
cell — the egg (Fig. 4, A) . In most cases this egg,
or female sex-cell, is unable to develop in nature
unless it is penetrated by a spermatozoon or male
sex-cell (Fig. 4, B). The single cell resulting from the
fusion of an egg and a spermatozoon is known as a
zygote. One of the most remarkable of all phenom-
ena is the development of a large, com})lex organisui
from a minute, and apparently simple, zygote.

According to the older scientists, a miniature of the
adult individual was present in the egg, and (k*v(0-
opment consisted in the growth and exi)ansion of

B 1

D. H. HILL LIBRARY
North Carolina State College

2 GERM-CELL CYCLE IN ANIMALS

rudiments already preformed. This belief could not
continue to exist after Caspar Wolff's brilliant
researches proved that adult structures arise grad-
ually from apparently undifferentiated material ; that
is, development is epigenetic. Epigenesis, however,
does not explain development; it simply maintains
that it occurs.

During the years since the theory of epigenesis
was proposed a new theory of preformation has
entered into our conception of development, a theory
which we may designate as predetermination. We
know from our microscopical studies that the germ
cells possess a certain amount of organization, and
that the zygote contains certain structures con-
tributed by the egg and other structures brought into
the egg by the spermatozoon. Hence, to a certain
extent, development is predetermined, since the initial
structure of the zygote determines the characteristics
of the individual that arises from it. On the other
hand, development is also epigenetic, and our modern
conception includes certain features of each theory.

The Cell. A brief account of the structure,
physics, and chemistry of the cell will serve to give us
some idea of the condition of the zygote from which
the individual arises, and will help us to understand
certain events in the germ-cell cycle to be discussed
later.

The cell is the simplest particle of matter that is
able to maintain itself and reproduce others of its
kind. The term *ceir was applied by Hooke in 1665
to the cell-like compartments in cork. Cells filled

INTRODUCTION 3

with fluid were slightly later described by Malpighi.
In 1833 Robert Brown discovered nuclei in certain
plant cells. What is known now as the Cell Theory
is usually dated back to the time of the botanist
Schleiden (1838) and the zoologist Schwann (1839),
whose investigations of the cellular phenomena in
animals and plants added greatly to the knowledge
of these units of structure. At this time the cell-
wall was considered the important part of the cell,
but continued research proved this idea to be erro-
neous. Schleiden called the substance within the cells
plant slime. Later (1846) von Mohl gave the term
protoplasm to the same substance. The substance
within the animal cell was named sarcode by Du-
jardin. The similarities between the protoplasm of
plants and the sarcode of animals were noted by
Cohn, and animal cells without cell-walls were
observed by Kolliker (1845). It was not, however,
until 1861 that Max Schultze finally established the
fact that plant protoplasm and animal sarcode are
essentially alike, and defined the cell as a mass of
protoplasm containing a nucleus. Schultze's re-
searches serve as the starting point for modern
studies of cellular phenomena, but the definition
furnished by him must be modified slightly, since we
now know that many cells exist without definite
nuclei. These cells, however, are provided with
nuclear material scattered throughout the cell body
(the so-called distributed nucleus). Our definition
must be changed to read, a cell is a mass of proto-
plasm containing nuclear material. Changes like-

4 GERM-CELL CYCLE IN ANIMALS

wise have taken place in the Cell Theory; we no
longer consider cells as isolated units and the multi-
cellular animal as equivalent to the sum of its con-
stituent cells, but recognize the influence of the cells
upon one another, thus reaching the conclusion that
the metazoon represents the sum of the individual
cells plus the results of cellular interaction.

Cells vary considerably in size, ranging from those
we call Bacteria, which may be no more than 2 5 otto
of an inch in length, to certain egg cells which are
several inches long; the latter, however, owe their
enormous size to the accumulation of nutritive sub-
stances within them. An average cell measures
about 2 5V0 of ^n inch in diameter. Cells vary in
shape as well as in size; egg cells are frequently
spherical, but most cells are not, since they are sur-
rounded by other cells which press against them.
A diagram of a typical cell is shown in Fig. 1.

Authorities are not agreed as to the structure of
protoplasm; to some it appears, as shown in Fig. 1,
to consist of a network of denser fibers called spon-
gioplasm (s) traversing a more liquid ground
substance, the hyaloplasm. Others consider proto-
plasm to be alveolar in structure, thus resembling
an emulsion, whereas another group of zoologists
maintain that while protoplasm may appear to be
fibrillar or alveolar, its essential basis consists of
multitudes of minute granules. Wilson's view is
the one usually adopted at the present time ; that
is, the protoplasm of the same cell may pass suc-
cessively "through homogeneous, alveolar, and

INTRODUCTION 5

fibrillar phases, at different periods of growth and
in different conditions of physiological activity,"
and that "apparently homogeneous protoplasm is a
complex mixture of substances which may assume

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^^G. 1. — Diagram of a cell, as = attraction-sphere; c = centrosome;
ch = chromatin reticulum; cr = chromidia; ec — ectoplasm; en = en-
doplasm; A: = karyosome; Z = linin; m = mitochondria; we = meta-
plasm; nm = nuclear membrane; p = plastid; pi = plasmosome or
nucleolus; s = spongioplasm ; v = vacuole.

various forms of visible structure according to its
modes of activity."

The physical properties of protoplasm are not well
known, since most of our studies have been made with
fixed material. We know that protoplasm may
exist as a gel or a sol, and that it is intermediate
between true solids and true liquids, with many of

6 GERM -CELL CYCLE IN ANIMALS

the properties of each and a number of properties
pecuHar to itself. No doubt the protoplasm differs
in its physical nature in different cells. In the egg
of the starfish, Asterias, Kite (1913) has shown that
the cytoplasm is a translucent gel of comparatively
high viscosity and is only slightly elastic ; pieces
become spherical when separated from the rest of
the egg. Scattered throughout this gel are minute
granules (microsomes) about xoVo^ mm. in diameter
which cannot be entirely freed from the matrix.
What appear to be alveoli contain globules which
possess many of the optical properties of oil drops;
these are suspended in the living gel. The cyto-
plasm of the starfish egg is not therefore alveolar in
structure as usually stated, but is rather of the
nature of a suspension of microsomes and globules
in a very viscous gel. The nuclear membrane is a
highly translucent, very tough, viscous solid, and
not a delicate structure as ordinarily conceived.
The nucleolus is a quite rigid, cohesive, granular gel
suspended in the sol which makes up the rest of the
nuclear material. Dividing male germ cells of cer-
tain insects (squash bugs, grasshoppers, and crickets)
revealed the fact that the chromosomes are the most
highly concentrated and rigid part of the nuclear
gel ; that the spindle fibers are elastic, concentrated
threads of nuclear gel ; and that the metaphase
spindle fibers seem to be continuous with the ends of
the chromosomes.

The ground substance of the nucleus is a sol termed
nuclear sap or karyolymph. In the so-called *rest-

INTRODUCTION 7

ing' nucleus a network of fibers may be observed
similar to the spongioplasm in the cytoplasm ;
these consist of a substance named linin because it
usually occurs in threads (Fig. 1, /). Distributed
along the linin fibers are granules of a substance which
stains deeply with certain dyes, and for this reason is
known as chromatin (ch). These chromatin gran-
ules may unite to form larger spherical masses, the
karyosomes or chromatin-nucleoli (A'), and during
mitotic nuclear division constitute the chromosomes
(Fig. 3, C). In many cells one or more bodies
resembhng the karyosomes somewhat, but differing
from them chemically and physiologically, are pres-
ent; these are the true nucleoli or plasmosomes
(Fig. 1, pi). Embedded in the cytoplasm near
the nucleus may often be seen a granular body, the
centrosome (c), which is thought to be of great
importance during mitotic cell division. The pro-
toplasm surrounding the centrosome is usually a
differentiated zone, the attraction-sphere (as), con-
sisting of archoplasm. The chromatin whicli may be
seen in the cytoplasm of certain cells is as a rule
in the form of granules called chromidia (cr). Cer-
tain other cytoplasmic inclusions that have attractiMJ
considerable attention within the past fifteen years
exist as granules, chains, or threads, and are known as
mitochondria, chondriosomes, plastosomes, etc. (m).
Various sorts of plastids (^), such as chloroplastids
and amyloplastids, may be present, besides a varying
number of solid or liquid substances, collectively
designated as metaplasm (me) or para])lasni, which

8 GERM-CELL CYCLE IN ANIMALS

are not supposed to form part of the living sub-
stance ; these are pigment granules, fat globules,
excretory products, vacuoles (y), etc.

It has been found possible to explain many cellular
activities and even the results obtained by experi-
mental animal breeding by studies of the physics
and chemistry of protoplasm. An exhaustive ac-
count of the subject is impossible and even unneces-
sary here, but the importance assigned to the physico-
chemical explanation of life phenomena requires a
brief statement. Kossel has separated the cellular
constituents into two main groups. (1) Primary
constituents are those necessary for life ; these are
water, certain minerals, proteins, nucleoproteins,
phosphatides (lecithin), cholesterin, and perhaps
others. (2) Secondary constituents are not essen-
tially necessary and do not occur in every cell ;
they are usually stored up reserve material or meta-
bolic products representing principally what we have
termed metaplasm.

Water which constitutes about two-thirds of the
animal is necessary for the solution of various bodies,
the dissociation of chemical compounds, the exchange
of materials, the removal of metabolic products,
etc. Mineral substances are present in all animal
tissues, and different tissues are characterized by
the presence of different minerals. The principal
ones are potassium, sodium, calcium, magnesium,
iron, phosphoric acid, sulphuric acid, and chlorine.
The other constituents are of a colloidal nature,
and its richness in colloids is one of the chief charac-

INTRODUCTION 9

teristics of protoplasm. To understand the activi-
ties of protoplasm we must therefore know sometliiiig
of the physics and chemistry of colloids.

Colloids (from colla = glue) do not diffuse, or
diffuse very slowly, through animal membranes ; in
this respect they differ from crystalloids, which
diffuse comparatively rapidly through animal mem-
branes. Wolfgang Ostwald recognized two sorts
of colloids : (1) suspension colloids, which are mix-
tures of solid and liquid phases, are non-viscous,
and easily coagulated by salts, e.g. a mixture of
finely divided metal and water ; and (2) emulsion
colloids, which are composed of two liquid phases,
are viscous, and coagulated by salts with difficulty.
Protoplasm is rich in emulsion colloids ; these may
exist as liquid sols, or more solid gels. In either
case they consist of fine colloidal particles. Accord-
ing to another classification colloids may be separated
into reversible and irreversible ; the former may
change from the sol to the gel state and back again,
but the latter are unable to do this. Protoplasm is
a reversible colloid, and many cellular structures
appear to originate through the gelation of licjuid
colloids. Since protoplasm is a sol or gel due to
water, it is a hydrosol or hydrogel, and because
of its water content is said to be hydrophylic. Il
' contains crystalloids and its chemical reactions take
place in a dilute solution of electrolytes ; these are
substances which dissociate, at least in part, into
their constituent ions when in solution, and the ions
are electrically charged. For example, NaCl disso-

10 GERM-CELL CYCLE IN ANIMALS

dates into electro-positive Na ions (cations) and
electro-negative CI ions (anions). Colloidal par-
ticles are likewise electrically charged, those of acid
colloids usually negatively and those of alkaline
colloids positively. The union and separation of
particles and their consequent rearrangement cause
gelation, liquefaction, etc. ; it is thus evident that
many physiological activities may be due to the
electrical charges of ions instead of the chemical
nature of the particles themselves. Cellular struc-
tures therefore depend upon the tendency of col-
loidal particles to form aggregates (gelation, coagula-
tion), and more or less upon the electrically charged
nature of the particles.

The most characteristic chemical constituents of
protoplasm are the proteins. The most common
proteins in the body show on the average the follow-
ing percentage of elements : —

Carbon 50 -55 %

Hydrogen 6.5- 7.3%

Nitrogen 15 -17.6%

Oxygen 19 -24 %

Sulphur 3- 2.4%

Proteins may be separated into three groups : (1)
simple proteins, such as protamines, albumins, and
globulins; (2) conjugated proteins, the glucopro-
teins, nucleoproteins, and chromoproteins ; and
(3) the products of protein hydrolysis, infraproteins,
proteoses, peptones, and polypeptides. These have
been studied both by microchemical and macro-
chemical methods. In the former method reagents
are applied to the microscopic objects and the

INTRODUCTION 11

changes in color, etc., indicate its constitution ; e.g.,
iron and phosphorus may be detected in this way.
Parts showing affinity for acid stains Hke eosin are
said to be acidophile or oxyphile ; those showing
affinity for basic dyes, Hke methylene blue, are
called basophile. The chromatin is basophile,
whereas the linin and cytoplasm are oxyphile. In
macrochemistry large quantities of the substances
are collected and examined by ordinary laboratory
methods.

Because of the importance that has been assigned
to the chromatin, this substance is particularly
interesting. Chromatin consists of nuclein, which is
a conjugated protein containing nucleic acid, the
latter being an organic acid, rich in phosphorus ;
it is hence called nucleoprotein. Nucleoproteins
are found chiefly in the nucleus but also occur in
the cytoplasm. They may differ from one another
in their protein content as well as in the character
of their nucleic acid constituent. When treated
with dilute acids nuclein is obtained, and when this
is further subjugated to caustic alkali it decomposes
into protein and nucleic acid. The nucleic acids
w^hich have been principally studied are those de-
rived from the thymus gland, and from the si)erma-
tozoa of salmon, herring, and other fish ; they are
probably all the same. Levene (1910) recognizes
three sorts of nucleic acid, of which the most complex
is termed thymonucleic acid. This consists of

two purine bases, guanine and adenine ;

two pyrimidine bases, thymine and cytosine ;

12 GERM-CELL CYCLE IN ANIMALS

a hextose (carbohydrate) ; and

phosphoric acid.
Its formula, according to Schmiedeberg, is C40H56
N14O16 . 2 P2O5, and according to Steudel, C43H57
N15O12 . 2 P2O5. Considerable progress has been
made, especially by Emil Fischer and his students,
in the synthesis of protein-like bodies. Many com-
plex polypeptides have been built up which resemble
peptones in many of their reactions and when in-
jected into living organisms appear to be utilized
in metabolism in much the same way as are native
proteins.

We are still, however, very far from an adequate
understanding of the nature of chromatin. Delia
Valle (1912), for example, after an exhaustive study
of the physico-chemical properties of chromatin
both in the resting nucleus and in the dividing cell,
has concluded that this substance resembles that
of fluid crystals. "Consequently all of the pheno-
mena presented by the chromosomes ; their mode of
origin, differences in size, state of aggregation, form,
structure, colorability, optical characteristics, varia-
tions in form, longitudinal division and the phenom-
ena which follow this mode of scattering, demon-
strate that the chromosomes are crystalloids."

Two other primary constituents of protoplasm may
be mentioned briefly. The phosphatide, lecithin,
belongs with cholesterin to a group of compounds
called lipoids. It consists of glycerophosphoric acid
plus certain fatty acid radicles, such as stearic acid,
oleic acid, etc., and a nitrogenous base (cholin). It

INTRODUCTION 13

probably plays some part in cell metabolism, may
furnish material for building up nucleins, and ac-
cording to Faure-Fremiet is concerned in the forma-
tion of mitochondria. Cholesterin is considered a
waste product of cell life, although it is known to in-
hibit haemolysis produced by certain bodies and is
thus a protective against toxins, and may have
other functions. We should look forward with great
interest to the results of investigations that are now
being carried on by biochemists, since we depend
upon them for an explanation of many of the phe-
nomena of life, cellular differentiation, and heredity.
We even hope that they may be able to create
compounds in the laboratory that we may consider
living organisms. However, the task does not
seem to be so simple to the biochemist, who should
know, as it does to the biologist. Nevertheless,
as Jacques Loeb has said, we should "either succeed
in producing living matter artificially, or find the
reasons why this should be impossible."

Cell Division. Cells may increase in number by
direct (amitotic) or indirect (mitotic or karyokinetic)
division. There is no doubt that mitosis occurs,
but not all investigators are convinced that cells
ever divide amitotically. Direct division was once
considered the only method of cell nudtiplication. It
was described as a simple division of the nucleus
into two parts (Fig. 2), preceded by a division of the
nucleolus into two, and succeeded by a constriction
of the entire cell ; the result was two daughter cells
each with one nucleus containing one-half of the

GERM-CELL CYCLE IN ANIMALS

nucleolus . As we shall see later (Chapter V) , amitosis
has been described in cells of the germ-cell cycle,
and must therefore be reckoned with in any discus-
sion of the phys-
ical basis of
heredity.

Mitosis or ka-
ryokinesis in-
volves a rather
complicated
series of pro-
cesses which
cannot be fully
discussed here
but will be out-
lined very briefly
with the aid of
Fig. 3.

(a) During
the prophase
the chromatin
granules which
are scattered
through the nucleus in the resting cell (A) become
arranged in the form of a long thread or spireme (B) .
At the same time the centrosomes move apart (A, c;
B, a), and a spindle arises between them (C). While
this is going on, the nuclear membrane generally
disintegrates and the spireme segments into a num-
ber of bodies called chromosomes (C) ; these take a
position at the equator of the spindle, halfway be-

Fig. 2. — Amitosis. A. Division of blood-cells
in the embryo chick, illustrating Remak's
scheme, a—e = successive stages of division.
{From Wilson, 1900.) B. Amitotic nuclear
division in the follicle cells of a cricket's egg.
{From Dahlgren and Kepner, 1908.)

INTRODUCTION

tween the centrosomes (Z), ep). The stage shown in
Fig. 3, J9, is known as the amphiaster ; at this time

Fig. 3. — Mitosis. Diagrams illustrating mitotic cell division. {From
Wilson.) A, resting cell; B, prophase showing spireme and nucle-
olus within the nucleus and the formation of spindle and asters (n)',
C, later prophase showing disintegration of nuclear memhrane, and
breaking up of spireme into chromosomes; D, end of prophases,
showing complete spindle and asters with chromosomes in etiuatorial
plate (ep); E, metaphase — each chromosome splits in two; F, ana-
phase — the chromosomes are drawn toward the asters, i'/= inter-
zonal fibers; G, telophase, showing reconstruction of nuclei; //.latex
telophase, showing division of the cell into two.

all of the mechanism concerned in mitosis is present.
There are two asters, each consisting of a centrosome

16 GERM-CELL CYCLE IN ANIMALS

surrounded by a number of radiating astral rays, and
a spindle which lies between them. The chromo-
somes lie in the equatorial plate [ej)).

(b) During the second stage, the metaphase, the
chromosomes split in such a way that each of their
parts contains an equal amount of chromatin {E, ep) .
As we shall see later, this is one of the most significant
events that takes place during mitosis.

(c) During the anaphase (F) the chromosomes
formed by splitting move along the spindle fibers
to the centrosomes. As a result every chromosome
present at the end of the prophase (D) sends half of its
chromatin to either end of the spindle. The mechan-
ism that brings about this migration is as yet some-
what in question. Fibers are usually left between
the separating chromosomes; these are known as
interzonal fibers (F, if).

(d) The telophase (G, H) is a stage of reconstruction
from which the nuclei emerge in a resting condition ;
the chromatin becomes scattered through the nucleus,
which is again enveloped by a definite membrane
(H) ; the centrosome divides and, with the centro-
sphere, takes a position near the nucleus. Finally
the cycle is completed by the constriction of the cell
into two daughter cells.

There are a number of differences between the
sort of mitosis just described and that which occurs
during the maturation of the egg and spermatozoon ;
these and certain other phases of cell division will
be considered in their appropriate places in succeed-
ing chapters.

INTRODUCTION 17

Methods of Reproduction. In the beginning
paragraph of this chapter it was stated, with reserva-
tions, that every individual develops from an v^^.
Before we can discuss the germ-cell cycle intelh-
gently, however, we must consider the exceptions
to this rule, and outline as briefly as possible the
various methods of reproduction which are known
to occur among animals. Reproduction is the forma-
tion of new individuals by division ; this is frequently
preceded by conjugation (in the Protozoa) or fertil-
ization (in both the Protozoa and the Metazoa).

Three principal methods of reproduction occur
in the Protozoa. (1) Binary fission appears to
be the most primitive. The individual divides into
two parts which are similar in size and structure ;
these grow into cells like the original parent. Many
CiLiATA, Flagellata, and Rhizopoda normally
reproduce in this way. (2) Budding occurs when
a small outgrowth or bud separates from the parent
cell. This method occurs among the Suctoria,
Radiolaria, Heliozoa, Ciliata, and Myxospo-
ridia. (3) Sporulation results from the division of
the nucleus of the parent into many daughter nuclei
and a subsequent division of the cell into as many
"spores" as there are nuclei. This process is
characteristic of the Sporozoa and also is found
among the Rhizopoda. Conjugation is of freqnent
occurrence in the Protozoa. Two or more inch*-
viduals may become connected without fusion of
nuclei or cytoplasm, thus forming colonies : a pair of
individuals may unite either temporarily or pvr-

18 GERM-CELL CYCLE IN ANIMALS

manently with fusion of the cytoplasm only; or
both cytoplasm and nuclei of such a pair may fuse
or be interchanged.

Metazoa reproduce either sexually or asexually.
Asexual reproduction is reproduction without the aid
of sex cells. It takes place as a rule by means of
buds or by fission as in many polyps, sponges,
flat- worms, segmented round-worms, and bryozoans.
Even the tunicates, which occupy an advanced posi-
tion in the animal series, form buds. Some of the
sponges produce internal buds called gemmules,
and certain bryozoans form similar bodies known
as statoblasts. Sexual reproduction requires that
the individual develop from a mature egg. As a rule
the egg must be fertilized by the union with it of a
spermatozoon, thus forming a zygote ; but the eggs
.of many animals develop without being fertihzed ;
that is, they are parthenogenetic. In rare cases such
parthenogenetic eggs may be produced, as in the
fly Miastor (see Chapter III), by immature individ-
uals. When this occurs, reproduction is said to be
poedogenetic.

The sex of an animal is judged by the kind of sex
cells it produces, — eggs by the female and sperma-
tozoa by the male, — and when the individuals of a
single species are differentiated as either males or
females, the species is said to be dioecious and the
individuals gonochoristic. In many species there is
but a single sort of individual which produces both
eggs and spermatozoa ; such species are monoecious,
and the individuals are hermaphroditic.

INTRODUCTION W

The Germ Cells. Eggs and spermatozoa differ
from each other both morphologically and physiolog-
ically. Eggs are usually spherical or oval in shape
(Fig. 4), although they may vary greatly from the
typical form and may even be ameboid as in certain
coelenterates. In size they range from that of tlie
mouse, which is only about 0.065 mm. in diameter, to
that of birds, which are several inches long. The
large volume of the latter is due to the presence of
an enormous amount of nutritive material, and the
general statement may be made that the size of an
egg does not depend so much upon the size of the
animal as upon the amount of yolk stored within it.
The egg nucleus, which is frequently very large and
clear, is known as the germinal vesicle ; and its
nucleolus has often been referred to as the germinal
spot. Embedded within the cytoplasm of the ovum
are several bodies besides the yolk globules. A
"yolk nucleus" may be present; mitochondrial
granules or rods may occur ; and special inclusions,
which become associated with the primordial germ
cells and have been named keimbahn-determinants,
have been recorded in many cases. Considerable
evidence has accumulated that the egg substance
is not a homogeneous, isotropic mixture, but is def-
initely organized, and that this organization is
related to the morphology of the embryo which is
to develop from it; hence we speak of the promor-
phology of the egg. Eggs are said to possess polarity,
and even the oogonium as it lies in the ovary is
definitely oriented with respect to its chief axes.

GERM- CELL CYCLE IN ANIMALS

The principal poles are dissimilar ; the end of the egg
containing most of the cytoplasm and nearer which
lie the nucleus and centrosome is known as the
animal pole ; the other end, which is often crowded

cm*

Fig. 4. — Germ cells. Ovarian ovum of a cat just before maturity.
0.771.= cell membrane; mics. = microsomes; ncl = nucleolus; n. m =
nuclear membrane; y/c. aZ. = yolk alveoli. {From Dahlgren and
Kepner.)

with the yolk globules, is called the vegetative pole.
The subject of the organization of the egg will be
referred to more in detail later (Chapter VIII).

The male sex cells or spermatozoa differ very
strikingly from the eggs. They are usually of the

INTRODUCTION

' Apical body or acrosome.

Nucleus.

• End-knob.

• Middle-piece.

• Envelope of the tail.

-Axial filament.

flagellate type (Fig. 4a), consisting of a head, largely
made up of chromatin, a middle piece, and a vibratile
tail. Spermatozoa are comparatively minute, rang-
ing in size from those of Amphioxiis, which are less
than 0.02 mm. long, to those of
the amphibian, Discoglossus,
which reach a length of 2.0 mm.
According to Wilson it would
take from 400,000 to 500,000
sea urchin spermatozoa to equal
in volume the egg of the same
species. It is not surprising,
therefore, to find that the num-
ber of spermatozoa produced by
a single male may be hundreds
of thousands times as great as
the number of eggs developed
in a female. Eggs are, as a rule,
incapable of locomotion, but
spermatozoa are active, swim-
ming about by means of their
tails until they reach the passive
eggs which they are to fertilize.
Since generally only one sperm-
atozoon fuses with an egg, it is
obvious that most of them never perform the function
for which they are specialized ; but apparently an
enormous number are formed to make the ferliHza-
tion of the eggs more certain.

The experiments of Loeb and Bancroft (1912) on
spermatozoa have shown that when the living

. End-piece.

Fig. -ia. — Diagram of a
flagellate spermatozoon.
{From WUson, 1900.)

22 GERM-CELL CYCLE IN ANIMALS

spermatozoa of the fowl are placed in a hanging drop
of white of egg or in yolk they undergo a transfor-
mation into nuclei. The possibility that a sperma-
tozoon may give rise to an embryo without the help
of an egg is recognized, but this has not yet been
accomplished.

The Life Cycles of Animals. The life cycle
of an animal has considerable influence upon the
course of the germ-cell cycle. In all animals that
are produced by the sexual method the beginning
stage in the life cycle is a mature egg, either fertilized
or unfertilized according to the species. Animals
which develop asexually, on the other hand, begin
their cycle with the first recognizable evidence of
budding or fission. As a rule budding or fission are
sooner or later interrupted by the formation of sex
cells, hence the life cycle of such animals may be
considered to extend from the mature egg to that
stage in the life history of the species when mature
eggs are again produced. Such a life cycle consists
really of two or more simple life cycles represented
by individuals differing from one another in both
structure and method of reproduction. As examples
of some of the principal types of life cycles we may
select certain insects and coelenterates.

A very simple life cycle is that of the wingless
insects of the order After a. The young, when they
hatch from the egg, are similar in form, structure,
and habits to the fully grown individual and undergo
no perceptible changes, except increase in size,
until they become sexually mature adults. In

INTRODUCTION 23

certain other groups of insects, such as the grass-
hoppers, the newly hatched young resemble the
adult in many ways, differing i)rincipally in the
absence of wings. The young Rocky Mountain
locust (Melanoplus spretus), for example, changes
its exoskeleton (molts) five times before the adult
condition is attained. After each molt there are
slight changes in color, structure, and size, the most
notable difference being the gradual acquirement of
wings. In still other orders of insects a larva
hatches from the egg ; this larva, on reaching its full
growth, changes in shape and structure, becoming a
quiescent pupa, from which after a rather definite
interval an adult emerges.

A combination of two simple life cycles to form one
complex cycle occurs in certain hydroids. The
eggs of these species produce free-swimming em-
bryos which become fixed to some object and de-
velop into polyps. These polyps form other polyps
like themselves by budding, but finally give rise to
buds which become jelly-fishes or medusae. In-
stead of remaining attached to the parent colony
the medusae, as a rule, separate from it and swim
about in the water ; they later give rise to eggs which,
after being fertilized, develop as before into polyps.
There are thus in this species two life cycles com-
bined, that extending from the egg to the time when
the colony forms medusa-buds, and that beginning
with the medusa-bud and ending with the mature
egg. Such an alternation of an asexual and a sexual
generation is known as metagenesis.

24 GERM-CELL CYCLE IN ANIMALS

There is another sort of alternation which nor-
mally occurs in many species, and that is the alterna-
tion of individuals developing from parthenogenetic
eggs with those from fertilized eggs. In the aphids,
or plant lice, for example, the race in the northern
part of the United States passes the winter in the
shape of fertilized eggs. All of the individuals which
hatch from these eggs in the spring are females called
stem-mothers. The stem-mothers produce broods of
females from parthenogenetic eggs, and these in
turn give rise to other broods of females in the same
manner. Thus throughout the summer, generation
after generation of parthenogenetic females appear ;
but as autumn approaches females develop whose
eggs must be fertilized, and males are also pro-
duced. The eggs of these females are fertilized by
spermatozoa from the males, and the zygotes thus
formed survive the winter, producing stem-mothers
the following spring.

CHAPTER II

GENERAL ACCOUNT OF THE GERM-CELL CYCLE

IN ANIMALS

It will be impossible to present in this chapter even
a general account of all the variations in the germ-
cell cycle that are known to occur in animals. It
will be necessary, therefore, to restrict ourselves to
the series of events that occurs in the majority of
animals, mentioning as many of the more notable
variations and exceptions as possible without causing
confusion. It also seems advisable to consider
the germ-cell cycles in the Protozoa and the Meta-
ZOA separately.

Protozoa. Weismann, in his classical essays
on the germ-plasm, argues in favor of the view that
the Protozoa are potential germ cells, and, since new
individuals arise by division of the parent cell into
two or more parts, that natural death does not occur.
The Protozoa are consequently also potentially
immortal. The Metazoa, on the other hand,
possess a large amount of somatic substance which
always dies a natural death. It has often been
pointed out that a Protozoon, although consisting
of but a single cell, performs most of the physiological
activities characteristic of the larger, complex
Metazoa, and that certain parts of the Protozoon

D. H. HILL LIBRARY

NJr»rt!» ri^rolina State College

GERM-CELL CYCLE IN ANIMALS

are recognizably concerned with the performance
of certain definite functions. The fundamental
difference, then, between the one-celled and the
many-celled animals is that the differentiated struc-
tures in the former are not separated from one
another by cell walls as in multicellular organisms.

Whether all Protozoa possess a body which can be
considered as specialized and set aside for reproduc-
tion purposes,
as the germ-
plasm theory
requires, is a
p question upon
which author-
ities differ. In
certain cases
it seems pos-
sible to distin-
guish between
germinal and
somatic proto-
plasm without any difficulty. The life history of the
fresh water rhizopod, Arcella vulgaris (Fig. 5), will
serve to illustrate this (Hertwig, 1899 ; Elpatiewsky,
1907; Swarczewsky, 1908; Calkins, 1911). The
single nucleus of the young Arcella divides to form
two primary nuclei (N) ; chromatin from these mi-
grates out and forms a layer near the periphery (Ch)
— the " chromidial net " of Hertwig. This chromatin
substance in the mature individual produces hundreds
of secondary nuclei (n), each of which is cut off, with

Fig. 5.— Reproduction in Arcella vulgaris. A. For-
mation of secondary nuclei. Ch = ehroniidia;
n = secondary nuclei; iV = primary nucleus.
{From Hertwig, 1899.) B. Two gametes. {From
Elpatiewsky, 1907.)

ACCOUNT OF THE GERM-CELL CYCLE 27

a small amount of the surrounding cylopiasni,
from the others, thus becoming a swarm spore.
The swarm spores escape from the mouth of the
parent cell ; whereas the two primary nuclei and a
portion of the cytoplasm not used up in the forma-
tion of the swarmers die. The swarmers are not
all alike, being of two sizes ; the larger, which may be
called macrogametes, and which correspond to the
eggs of the Metazoa, fuse with the smaller micro-
gametes. The zygotes which result develop into
normal Arcelloe. The swarmers may be supposed
to represent the germinal protoplasm, of which, as in
metazoan germ cells, the chromatin content may be
considered the essential portion. The conditions
during reproduction in other Protozoa may also be
explained in this way, so that germinal and somatic
protoplasm can be distinguished as in the Metazoa.
The discovery of the chromidia in Protozoa
led to the formulation of the hypothesis of binu-
clearity. Believers in this hypothesis maintain
that each cell contains both a somatic and propaga-
tory nuclear material which, as a rule, are united
into one amphinucleus. The somatic nuclear ma-
terial controls vegetative functions; the propaga-
tive portion serves only for the propagation of new
individuals. Separation occurs rarely except in
certain Protozoa, where, as in Paramecium, the
propagative substance is represented by the micronu-
cleus, the somatic by the macronucleus. Since the
chromatin is the essential substance concerned in
the binuclearity hypothesis, the term dichroma-

28 GERM-CELL CYCLE IN ANIMALS

ticity has been suggested as more appropriate, and
the two kinds of chromatin involved have been called
idiochromatin, which is reproductive in function,
and trophochromatin, which is vegetative in function.
The hypothesis has not gained many adherents and
is considered of doubtful value by eminent proto-
zoologists (Dobell, 1908).

Metazoa. If we consider the mature egg, either
fertilized or parthenogenetic, as the starting point
of the germ-cell cycle in the Metazoa, we may
recognize seven or eight distinct periods as follows :

1. The segregation of the primordial germ cells;
i.e., the formation of one or more primordial germ
cells during the segmentation of the egg ;

2. Early multiplication of the primordial germ
cells ;

3. A long period of "rest" characterized by cessa-
tion of cell division, either active or passive change
of position, separation of the germ cells into two
groups which become the definitive germ glands,
accompanied by the general growth of the embryo
until the larval stage is almost attained ;

4. Multiplication by mitosis of the primitive
oogonia or spermatogonia to form a definite number
(Miastor and perhaps others) or indefinite number
(so far as we know) of oogonia or spermatogonia ;

5. In some cases the differentiation of oogonia
into nurse cells and ultimate oogonia, and the
spermatogonia into Sertoli cells and ultimate sper-
matogonia ;

6. The growth of the ultimate oogonia and sper-

ACCOUNT OF THE GERM-CELL CYCLE !29

matogonia to form primary oocytes and j)riniary
spermatocytes ;

7. Maturation ;

8. Fertilization (if not parthenogenetic).

1. The Segregation of the Primordial Germ
Cells. This phase of the germ-cell cycle is espe-
cially emphasized in this book (see Chapters III to VI)
and need be referred to only casually here. The
mature eggs of animals are organized both mor-
phologically and physiologically ; that is, differenti-
ations have alread}^ taken place in their protoplasmic
contents before they are ready to begin develop-
ment. This organization determines what sort of
divisions the *^%g will undergo during the cleavage
stages. During cleavage certain parts of the cell
contents become separated from other parts and
thus the differentiated substances of the ^^g are
localized in definite parts of the embryo. The
contents of the cleavage cells likewise become
differentiated as development proceeds, until finally
the cells produced form two or three more or less
definite germ layers. In some cases the ^gg always
divides in the same way, and the history or *'cell
lineage" of the cells can be followed accurately,
and the parts of the larva to which they give rise
can be established. This is known as determinate
cleavage in contrast to the indeterminate type in
which there appears to be no relation between the
cleavage cells and the structure of the ^gg or larva.

The degree of organization of the i^gg no doubt ac-
counts for the differences in cleavage; those of the

30 GERM-CELL CYCLE IN ANIMALS

determinate type being more fully organized than
those of the indeterminate type.

The period v:hen the primordial germ cells are es-
tablished is probably due in part to the state of
organization of the egg when development begins,
and it is not strange, therefore, that the primordial
germ cell may be completely segregated in certain
eggs as early as the four-cell stage ; whereas in
others germ cells have not been discovered until a
late larval condition has been reached. An ever
increasing number of species of animals is being
added to those in wdiich an early segregation of the
germ cells has already been recorded. Neverthe-
less, there are certain zoologists who still question
the general occurrence of an early segregation of
the germ cells, but more careful investigations will
probably establish the fact of early segregation in
species in which this has not yet been demonstrated.

2. Early Multiplication of the Primordial
Germ Cells. The number of germ cells present
at the time of their first appearance in the embryo
varies in different species. There may be one, as
in the majority of cases, for example the fly,
Miastor (Fig. 17), the nematode, Ascaris (Fig. 51),
the crustacean, Cyclops (Fig. 48), and the arrow
worm, Sagitta (Fig. 54) ; or a number, as in chrysome-
lid beetles (Fig. 36), certain parasitic Hymenoptera
(Fig. 44), and vertebrates (Fig. 6). As a rule the
primordial germ cell or cells increase in number by
mitosis soon after they are segregated, and then
cease to divide for a considerable interval. For

ACCOUNT OF THE GERM- CELL CYCLE 31

example, in Miastor the single primordial ^^erm cell
produces eight ; in the beetle CaUu/rapha mulfi-
punctata the original sixteen undergo two divisions
resulting in sixty -four ; and in the chick Swift (1914)
has counted as many as eighty-two at this stage.

We shall see later that the primordial germ cells
are often characterized by the presence of certain
cytoplasmic inclusions (the keimbahn-determinants)
which are absent from the other cells of the embryo.
These inclusions appear to be equally divided be-
tween the daughter cells so that each of the eight or
sixty-four, as the case may be, is provided with an
equal amount of the keimbahn-determinants.

3. Period of "Rest" and Migration. By rest
here is really meant cessation of division. During
this period the germ cells either actively migrate
or are passively carried by surrounding tissues to
the position the germ glands occupy in the larva.
In species possessing two germ glands the germ cells
separate to form two groups, with, at least in some
cases, an equal number in each group. Thus in
Miastor the number in each group is four (Fig. 'I'i)
and in Calligrapha, thirty-two (Fig. 37). There is
evidence that an active migration of germ cells
occurs both in vertebrates and invertebrates. Figure
6 shows the positions of the germ cells in four s])ecies
of vertebrates during their change of ])ositi()n.
That the germ cells at this time are actively nngrat-
ing by ameboid movements is the general opinion
of investigators, since frequently these cells are
ameboid in shape and the distance between the i)lace

GERM-CELL CYCLE IN ANIMALS

of origin and the germinal ridge is too great to be
traversed in any other way.

Professor B. M. Allen, who has made extensive
studies of the germ cells of many species of verte-

LepMosteus

LepldosUus

Fig. 6. — Diagrams showing the paths of migration in A, a turtle,
Chrysemys marginata ; B, a frog, Rana pipiens ; C, a fish, Lepidos-
teus osseus, and D, the dog-fish, Amia calva. (From Allen, 1911.)
Arc/i = archenteron; /n^ = intestine; La^ Mes = lateral plate of
mesoderm; Mes = mesentery; Meson = mesonephros; M2/0 = myo-
tome; A^oto = notochord; P. card = post cardinal vein; S. C = sex-
cells; S.Gl = sex gland; V it. End = vitelline endoderm; W.D =
Wolffian duct.

brates, makes the following statement regarding this
phase of the germ-cell cycle :

"The sex-cells are migratory to a high degree.
The path and time of their migration may vary
greatly within a given group of animals, as illus-

ACCOUNT OF THE GERM-CELL CYCLE 33

trated by the case of Amia and Lepidosteus. While
in the forms that I have studied they are first to be
observed in the entoderm, I am quite open to convic-
tion that in other forms they may migrate from tliis
layer into the potential mesoderm before the two
layers are separated, as shown by Wheeler in Petro-
myzon.'^

Swift (1914) has recently obtained evidence which
seems to prove that not only do the germ cells of
the chick migrate by ameboid movements but they
enter the blood vessels and are distributed by the
blood stream to all parts of the embryo and vascular

area.

The migration of the germ cells has been noted in
many invertebrates and has been fully described
in chrysomelid beetles (Hegner, 1909a). In these
insects the primordial germ cells are segregated at
the posterior end of the egg at the time the blasto-
derm is formed (Fig. 36, C). The blastoderm is
never completed just beneath them, but a canal,
called the pole-cell canal, remains. Through this at
a later embryonic stage the germ cells migrate by
means of ameboid movements.

"As soon as the germ cells of CalUgrapha have
passed through the pole-cell canal, they lose their
pronounced pseudopodia-like processes and bocuine
nearly spherical (Fig. 37, E) ; nevertheless, they
undergo a decided change in position. They move
away from the inner end of the pole-cell canal, and
creep along between the yolk and the germ-band.
Thus two groups are formed near the developing

34 GERM-CELL CYCLE IN ANIMALS

coelomic sacs ; each group probably contains an
equal number of cells. The smallest number I
have counted in one group at this time is thirty;
the largest number, thirty-four. As there is some
difficulty in obtaining an accurate count, it seems
probable that the sixty-four germ cells are equally
divided and that each germ gland receives thirty-two.
Some of the germ cells migrate not only laterally
along the germ gland but also back toward the
posterior end of the egg, where we find them forming
narrow strands in the last abdominal segments.
From this stage on, the germ cells are not very active ;
they move closer to one another to form the compact
germ glands. I was unable to determine whether
the later movements of the germ cells are due to
an active migration or to the tensions created by
the growth of the surrounding tissues; the latter
seems the more probable" (Hegner, 1909a, p. 280).

It is thus evident that during the blastoderm stage
the germ cells of this beetle are actually outside of the
egg. How well this illustrates the theory of primary
cellular differentiation, that is, the differentiation of
germ cells from somatic cells, since the two sorts
are here completely separated, the former constitut-
mg a group in contact with but not connected with
the somatic cells. Later, as the germinal con-
tinuity hypothesis demands, the germ cells migrate
into the embryo, there to be nourished, transported,
and protected by the body until they are ready to
separate from the somatic cells, and thus to give rise
to a new generation.

ACCOUNT OF THE GERM-CELL CYC LE 35

4. Period of Multiplication. Soon after tlio
germ cells aggregate to form more or less rounded
groups lying in the position of the definitive germ
glands mitotic division is resumed. At about this
time also, the sex of the individual can often be
determined by the shape of the germ-gland. Then
both the testes and the ovaries acquire envelopes
of the follicular cells, and frequently testicular cysts
and ovarian tubes or chambers develop. The ques-
tion of the origin of the follicular cells is still un-
settled, but the evidence in most cases seems to
favor the view that they are mesodermal.

The multiplication of the germ cells by mitosis
continues rapidly from this time on. In only one
case, so far as I am aware, do we know the actual
number of germ cells produced by the primordial
germ cell ; this is in Miastor, where typically sixty-
four oogonia are formed (Fig. 26). As the germ
cells multiply they become smaller in size and the
substances present in the primordial germ cell
become divided among a large number of progeny.
Thus at the beginning of the growth period each
germ gland contains many oogonia or spermatogonia,
and each of these contains a small fraction of the
material present in the primordial germ cell, plus
whatever substances may have been assimilated
during the period of multiplication.

5. The Origin of Nurse Cells and Sertoli
Cells. Germ cells receive nourishment during the
growth period in many ways, e.g., from nurse cells,
follicle cells, or directly from the blood. The origin

36 GERM-CELL CYCLE IN ANIMALS

of the nurse cells and follicle cells is important since
in a few cases the germ cells themselves are known
to give rise to them. There is thus a second differ-
entiation whereby somatic cells (follicle cells or
nurse cells) become differentiated from germ cells
(oogonia or spermatogonia). In some species, such
as Miastor, we can prove without question that both
the nurse cells and follicle cells are of mesodermal
origin, and that the germ cells give rise only to germ
cells. On the other hand, there are instances in
both vertebrates and invertebrates of a common
origin of germ cells and somatic cells from oogonia
and spermatogonia. Perhaps the most striking
examples are the differentiation of the nurse cells
and ultimate oogonia in the water beetle, Dytiscus,
and the differentiation of the Sertoli cells and ulti-
mate spermatogonia in man. (See Chapter V.)
Haecker (1912) distinguishes' between a somato-ger-
minative period and a true germinative period ; the
former is that during which the primordial germ cells
are established and the latter that of the differentia-
tion of nurse cells and ova.

6. The Growth Period. The last divisions
of the oogonia and spermatogonia are followed by
the growth of these cells. The extent of this growth
depends, in the case of the female, upon whether
or not the mature egg is to be supplied with an
abundance of nutritive material. Nurse cells, fol-
licle cells, and circulating fluids may all assist in the
enlargement of the oogonia. If the eggs are small,
sufficient nutriment is supplied by surrounding

ACCOUNT OF THE GERM-CELL C YCLE 37

liquids and no special nurse cells are required ; but
larger eggs either become surrounded by follicle cells
which nourish them and with which they are often
intimately connected by protoplasmic bridges, or
special nurse cells are provided. In the primitive
type of ovary, such as exists in most coelenterates,
any of the cells surrounding the oogonium may
function as nurse cells and even neighboring oogonia
are engulfed by the oogonium that is successful in the
struggle for development. A more definite mechan-
ism exists in higher organisms, where one or more
cells become differentiated for the special purpose of
supplying nutriment consisting of either their own
substance or of material elaborated by them and
then transferred to the egg. The egg of the annelid,
Ophryotrocha, for example, is accompanied by a single
nurse cell ; that of Myzostoma is provided with two,
one at either end ; and the eggs of certain insects
are more or less intimately connected with groups of
cells in definite nurse chambers (Fig. 46).

The growth of an oogonium may be well illus-
trated by that of the potato beetle.

The general arrangement of the cells in the ovary
of an adult beetle is shown in Fig. 7. The terminal
chamber of the ovarian tubule contains three kinds
of cells: (1) nurse cells (/i.c), (2) young oocytes
{y.o) and growing oocytes, and (3) epithelial cells.
The nurse cells and oocytes are both derived from the
oogonia ; the epithelial cells are of mesodermal origin.

The positions of the stages to be described are
indicated in the diagram (Fig. 7) and the nuclear

GERM-CELL CYCLE IN ANIMALS

ooc

Fig. 7. — Leptinotarsa de-
cemlineata. Diagram of
an ovarian tubule showing
various stages in the de-
velopment of the oocyte.
The capital letters refer to
the positions f cells shown
in Fig. 8. cy = cytoplasm;
es = egg string; n.c = nurse
chamber; doc = oocyte; y.o
= young oocyte.

and cytoplasmic structures
are shown in Fig. 8. Two
oocytes and a neighboring
epithehal cell from position
A in Fig. 7 are shown in Fig.

The nuclei of the oocytes
are large and contain a dis-
tinct spireme ; the cytoplasm
is small in amount and ap-
parently homogeneous. After
a short period of growth the
oocytes form a linear series
in the ovarian tubule and
become connected with the
spaces between the nurse cells
by means of egg strings (Fig.
7, e.s) through which the nu-
tritive streams flow into the
oocytes. One of the young-
est of these oocytes is repre-
sented in Fig. 8, B (position
B in Fig. 7). The nucleus is
no larger than in those of the
earlier stage ; its chromatin
forms a reticulum, and a dis-
tinct nucleolus is present.
The cytoplasm, on the other
hand, has trebled in amount
and within it are embedded
a number of spherical bodies

ACCOUNT OF THE GERM-CELL CYCLE 30

:. J'i

'.■■>

•c^

;.:.,W-_ -r;* ::.•.•,:.•.^vv.■..:..:•...

/ep-|:

-TIG. 8. Leptinotarsa decemlineafn a^tt q. . ,,

oocyte from positions rcHcatod il I^'. 7^'' "^ ''^^' ^'"^^'^'^ "^ ^''"
division of nurse cells ./ - I .^ ' ''~'' = anutotic nuc-l.ar

iur^e cells, ch = chorion; f.ep = follicular epithelium.

40 GERM-CELL CYCLE IN ANIMALS

which stain with crystal violet after Benda's method,
and appear to be mitochondrial in nature. At a
slightly later stage (Fig. 8, C ; position C in Fig. 7)
the nucleus is larger and contains several small
spherical chromatic bodies besides the nucleolus.
The cytoplasm has increased more rapidly in volume
and a corresponding increase in the number of mito-
chondrial granules has also taken place. Further
growth results in an increase in the volume of both
nucleus and cytoplasm (Fig. 8, D ; position D in
Fig. 7), and a slight increase in the number of mito-
chondria. Whether these bodies developed de novo
or by division of the preexisting granules could not
be determined.

In succeeding stages growth is very rapid. The
cytoplasm (Fig. 8, E\ position E in Fig. 7) still
remains homogeneous except for the mitochondria,
which increase slightly in size and become situated
as a rule near the periphery. The nucleus at this
time contains a large number of chromatin granules
and a diffuse reticulum. Part of an older oocyte
is shown in Fig. 8, F (position F in Fig. 7) ; the cyto-
plasm has assumed a reticular appearance; the
mitochondrial granules are present in greater num-
bers, and the nucleus is larger, oval in shape, and
contains a distinct reticulum with manv chromatin
bodies of various sizes. A still older oocyte (Fig.
8, G ; position G in Fig. 7) is interesting particularly
because of the rapid increase in the mitochondria and
the localization of these near the periphery. From
this stage on the character of the contents changes

ACCOUNT OF THE GERM-CELL CYCLE 41

until, as shown in Fig. 7, the central part of the
oocyte consists of homogeneous cytoplasm (cz/), and
the outer region of the cytoplasm is crowded with
granules and spherical bodies of various sizes.
Apparently the mitochondria lying near the periphery
(Fig. 8, H) increase in size, gradually losing their
affinity for the crystal violet stain and swelling up
until they constitute the large yolk globules so
numerous in the mature egg. All stages in the
evolution of these bodies are illustrated at this time
as represented in Fig. 8, H. Li the meantime
material is brought into the egg through the egg
string from the nurse cells, thus probably adding
several sorts of granules to the contents of the oocyte.

The growth period in the male germ-cell cycle is
not so striking as in the female, since many sperma-
tozoa of small size are produced, whereas only
comparatively few large eggs develop. An increase
in the size of the ultimate spermatogonia may occur,
however, but the multiplication and growth periods
are not nearly so distinct as in the case of the oogonia.
In testes which are composed of cysts of spermato-
gonia there is evidence in some cases that all of the
germ cells in a single cyst are descendants of a single
spermatogonium. The proof for this seems certain
in the potato beetle, where I have been al)k> to
follow the formation of the cysts by means of an
uninterrupted series of stages (Hegner, 1914a).

7. Maturation. Maturation or the ripening
of the eggs and spermatozoa comprises a series of
events which results in a reduction in the number

GERM-CELL CYCLE IN ANIMALS

of chromosomes and the amount of chromatin in
the germ cells. Typically, both male and female
germ cells divide twice during the process of matura-

PRIMORDIAL
GERM-CELL

MULTIPLICATION
• • \ f • • ^ r PERIOD

SPFRMATOGONIA^--"' \ • • / V • •

• v <!_V V^V Uv

GROWTH
PERIOD
PRIMARY / t 1

SPERMATOCYTE ' *

SECONDARY . , , ^,

SPERMATOCYTES

MATURATION
_ PERIOD

SPERMATIDS (7>\ Cr>\ (i

SPERMATOZOA ^

PRIMORDIAL ,

GERM-CELL \% * ,

MULTIPLICATION
• •> f * • '^ ^ PERIOD

OOGONIA ^'

'••\ ' ^ *\ [* *\ { * *

f J V • •/ \« •/ \ • •>

GROWTH
Pf'l^'ARY / \ ^ pERioo

OOCYTE I • •

SECONDARY

OOCYTES

(OVARIAN EGG "" ~ ~ ~\ * * } ^ V MATURATION

AND POLARBODY) \ 7 /\ > PERIOD

MATURE EGG /^ ^ \^ d. \_^

AND /^ , , ] P) rt ft

POLAR BODIES \ 7 ^^ V_y ^-^

Fig. 9. — Diagrams illustrating (above) the stages of spermatogenesis
and (below) of oogenesis. The primordial germ cell is represented
as possessing four chromosomes.

ACCOUNT OF THE GERM-CELL CYCLE 43

tion, and as shown in Fig. 9 these divisions result
in the production of four functional spennalozoa
in the male, and one functional egg and three polar
bodies (abortive eggs) in the female. This increase
in the number of cells is not, however, the most im-
portant phase of the maturation process, since a
large part of our knowledge of the physical l^asis of
heredity has been derived from studies of the be-
havior of the chromatin at this time. This subject
will be dealt with more fully in Chapter IX, and
for the present only a brief account of events need
be given.

The first thing to be noted is that the mitoses
leading to the division of the germ cells during mat-
uration differ from those of ordinary cell multiplica-
tion. The germ cells, when they are ready for the
maturation divisions, are known as primary oocytes
and primary spermatocytes. The nuclei of these
cells possess the complete or diploid number of
chromosomes, characteristic of somatic cells ; but
after maturation the eggs and spermatozoa con-
tain only one-half of the original diploid number,
or the haploid number. These mitoses are conse-
quently called reducing or meiotic. The details of
these mitoses differ in male and female germ cells
and in different species of animals.

During and at the close of the growth period in the
male the chromatin granules form a spireme which
condenses at one side of the nucleus, a condition
known as synizesis. After a time the s})irenie
again spreads throughout the nucleus, but is now

44 GERM-CELL CYCLE IN ANIMALS

divided into segments, the chromosomes, which are
only haploid in number. The reduction from the
diploid to the haploid number is brought about by
the union of the chromosomes in pairs, a condition
called synapsis. Each of the haploid chromosomes
thus consists of two of the diploid chromosomes
and is said to be bivalent. That one of the chromo-
somes of each pair is of maternal origin, i.e., is a
descendant of a chromosome present in the egg at
the time of fertilization, and the other of pater-
nal origin, i.e., a descendant of one brought into
the egg by the spermatozoon, seems to be well
established. The final act of fertilization, therefore,
occurs at this point in the germ-cell cycle — an
act of much greater significance than that of the
union of the egg and spermatozoon. Furthermore,
there is considerable evidence that the chromo-
somes differ one from another and that in synapsis
corresponding (homologous) chromosomes unite.
The importance of such a union from a theoretical
standpoint will be discussed later.

The nuclei now prepare for the two maturation
mitoses. In manv nematodes, annelids, and arthro-
pods these are characterized by the formation of
tetrads. Divisions of this sort may be illustrated
as in Fig. 10. The diploid number of chromosomes
is for convenience supposed to be four, as in the sper-
matogonium A. During the spermatogonial divi-
sions these divide as in B, so that each daughter cell
receives the diploid number, four. After synapsis,
however, each of the haploid chromosomes of the

ACCOUNT OF THE GEKM-CELL CY( LE 45

Fig. 10. — Diagrams showing the (.^sential facts of ri'tluction in the
male. The somatic number of chromosomes is supposeil to bo four.
A, B, division of the spermatogonia, showing the full number (four)
of chromosomes. C, primary spermatocyte i^rcparing for division:
the chromatin forms two tetrads. D, E, F, first division to form
. two secondary spermatocytes, each of which receives two dyads.
G, H, division of the two secondary spermatocytes to form four
spermatids. Each of the latter receives two .><ingl(' chromnsonies and
a centrosome which passes into the middle piece of the spermatozoon.
(After Wilson.)

46 GERM-CELL CYCLE IN ANIMALS

primary spermatocyte is seen to be divided into
four parts,, thus forming in this case two tetrads (C).
During the division of the primary spermatocyte,
as shown in /), E, and F, half of each tetrad, or two
dyads, passes to each daughter cell. The division
of the daughter cells, which are known as secondary
spermatocytes {G H), results in the separation of the
two parts of each dyad so that each of the four
spermatids {H) receives one member of each original
tetrad or two monads. Thus the chromosomes
(monads) of the spermatids {H) are already formed
in the primary spermatocytes (C) by two divisions ;
whereas the nuclear and cell divisions do not occur
until later. The spermatids {H), which proceed
to metamorphose into spermatozoa, possess, there-
fore, only two chromosomes, i.e., one-half of the
number present in the spermatogonia {A) and so-
matic cells.

Tetrad formation does not occur in most animals ;
but usually the members of the bivalent chromosomes
become separated on the first maturation spindle,
the pairs appearing U-, V-, or ring-shaped, as in
Fig. 6^. Each secondary spermatocyte receives
one-half of each haploid, bivalent chromosome. The
second maturation mitosis then ensues, during which
each daughter cell is provided with one-half of each
chromosome as in ordinary mitotic division. Be-
cause of the peculiar behavior of the chromosomes
the first division is often called the heterotype,
whereas the second is known as the homotype divi-
sion. The final results are the same whether tetrads

ACCOUNT OF THE GERM-CELL CYCLE 47

are formed or not, each spermatid containing the
haploid number of chromosomes.

The maturation of the egg differs in no very im-
portant respects from the process as it has been
described in the male cells. Tetrads may or may
not be formed according to the species, and the
mature egg and polar bodies each contain the haploid
number of chromosomes. Two phases of the matura-
tion of the egg may be referred to here : (1) when
the nucleus of the primary oocyte prepares for divi-
sion a considerable amount of chromatin separates
from the chromosomes and is lost in the cytoplasm.
The size of the chromosomes is thus diminished, but
no entire chromosomes are lost. (2) The cellular
divisions are very unequal, the polar bodies being
very small as compared with the rest of the egg.
The chromatin content of the polar bodies, however,
is equal to that of the much larger egg. In the male
all of the four spermatids are functional, but in the
female only the egg survives, the polar bodies i\e-
generating. As a rule two polar bodies are produced,
but in certain cases of parthenogenesis (rotifers,
Cladocera, Ostracoda, and aphids) only one is
formed. Rarely the first polar body divides into two.

8. Fertilization. Eggs that develop ])arthen()-
genetically are ready to begin a new germ-cell cycle
as soon as they become mature; but the eggs of
the majority of species must be fertilized before^
they are able to develop. Fertilization mny be de-
fined as the fusion of an egg with a spermatozoiin and
the resulting processes of rearrangement of the egg

48 GERM-CELL CYCLE IN ANIMALS

contents which result in the formation of a uninuclear
cell, the zygote. As a rule one spermatozoon only
enters the egg (monospermy) ; but in a few species
(certain insects, selachians, tailed amphibians, reptiles,
and birds) many spermatozoa may normally fuse
with the egg (physiological polyspermy). The sper-
matozoon, which consists usually of three rather dis-
tinct parts, the head, the middle piece, and tail,
may become entirely embedded within the egg sub-
stance, or the tail may be left outside, or, in excep-
tional cases, only the head succeeds in entering.

The union of the egg and spermatozoon may occur
before, during, or after the polar body formation
(Fig. 11). If the spermatozoon enters before the
maturation of the egg is completed (A), its head
transforms into a nucleus equal in size to that of the
egg (C) ; the middle piece dissolves, giving rise to
a centrosome which inaugurates the formation of a
spindle with asters (B) ; and the tailpiece ap-
parently takes no active part in the fertilization
processes. The middle piece also does not seem to
be necessary for the formation of the centrosomes
and asters. The nucleus of the spermatozoon and
that of the mature egg approach each other and
come into contact between the asters (C). Then the
nuclear walls dissolve; a spireme which segments
into the haploid number of chromosomes is produced
by each nucleus, and the first cleavage spindle of
the developing egg results. This spindle bears the
haploid number of chromosomes from the spermato-
zoon and a like number from the egg nucleus

ACCOUNT OF THE GERM-CELL CYCLE 41)

and thus the diploid or somatic number of diromo-
somes is regained.

When the spermatozoon enters an egg which has
completed polar-body formation, the head does not

Fig. 11. — Diagrams of two principal tj-pcs of fertilization. I. Polar
bodies formed after the entrance of the spermatozoa (annelids,
mollusks, flat-worms). II. Polar bodies formed before entrance
(echinoderms).

A, sperm-nucleus and centrosome at $ ; first polar body forming
at 9 • B, polar bodies formed; approach of the nuclei. C, union
of the nuclei. D, approach of the nuclei. E, union of the nuclei.
F, cleavage-nucleus. {After Wilson.)

have time to transform into a nucleus as large as
the egg nucleus, but nevertheless fuses with the latter
(Fig. 11, D, E, F). Although the two nuclei are very
unequal in size, they possess an equal amount of
chromatin and furnish an equal number of chromo-
somes to the first cleavage spindle.

50 GERM-CELL CYCLE IN ANIMALS

As already indicated, perhaps the most essential
phase in the fertilization process does not occur until
the homologous maternal and paternal chromosomes
unite during synapsis, when the germ cells of the
new individual become mature. The immediate
results of fertilization are : (1) the inauguration of
the development of the egg, (2) the increase of the
chromosomes from the haploid to the diploid (so-
matic) number, and (3) the union of hereditary
substances from, as a rule, two individuals.

This completes the last stage in the germ-cell
cycle of animals. Many extremely important and
interesting phases of the subject have had to be
omitted from the account. Certain of these will
be more fully discussed in succeeding chapters, es-
pecially those concerned with the early history of
the germ cells during embryological development,
but for the details of the nutrition, growth, matura-
tion, and fertilization of the germ cells, the reader
must be referred to other sources (Wilson, 1900;
Jenkinson, 1913; Kellicott, 1913).

CHAPTER III

THE GERM-CELL CYCLE IN THE PiEDOGENETIC

FLY, MIASTOR

Thus far in only one genus of animals has the
history of the germ cells from one generation to the
next been followed in detail through the entire
cycle. This is a genus of flies, Miastor, of the family
Cecidomyidae. One species, Miastor metraloas, oc-
curs in Europe and has there been studied especially
by Leuckart (1865), Metschnikoff (1865, 1866), and
Kahle (1908), and the only other species that has
been investigated is M. americana (Hegner, \^V2,
1914a).

Psedogenesis in Miastor was discovered by Wagner
in 1862, and was confirmed by Meinert in 1864.
In 1865 the first investigations of its embryological
development were published by Leuckart and Metsch-
nikoff. These were the earliest accounts of the
keimbahn in any animals. Only a glance at Metsch-
nikoff's report is necessary to convince one of the
favorableness of Miastor as material for germ-cell
studies. The primordial germ cell is shown to be
established at a very early period in the cleavage of
the egg, and the descendants of the primordial germ
cell are quite easily distinguishable from other cells
in the body even in in toto preparations. In spite of

52 GERM-CELL CYCLE IN ANIMALS

the work of the above named investigators there were
many who were not convinced that psedogenesis
occurs in the genus, and the larvse which were
known to develop within the bodies of other larvae
were considered by these skeptics as parasites. How-
ever, the results of Kahle's (1908) studies, which have
been decisively confirmed (Hegner, 1912, 1914a),
have finally settled the question in favor of psedogen-
esis.

Previous to 1910 no specimens of the genus
Miastor had been recognized in this country, but
on Oct. 5 of that year, Dr. E. P. Felt found them in
great abundance, living in the partially decayed
inner bark and in the sapwood of a chestnut rail.
With material supplied by Dr. Felt, the writer
has been able to follow the entire keimbahn in these
insects. P^dogenetic reproduction normally oc-
curs during the spring, summer, and autumn, multi-
plication being arrested during the cold winter
months. This method of reproduction is interrupted
in midsummer by the appearance of male and female
adults.

The larva of Miastor possesses two ovaries, one on
either side of the body in the tenth or eleventh
segment. Each ovary (Fig. 12) consists of typically
thirty-two oocytes (odc.n) ; these are inclosed in a
cellular envelope (en) . Associated with each oocyte
is a group of mesoderm cells which function as nurse
cells (n.c.) and together with the oocyte are sur-
rounded by a follicular epithelium (f.ep). The
nurse cells furnish nutrition to the growing oocytes,

THE P.EDOGENETIC FLY, MIASTOR 53

gradually becoming reduced as the oocytes increase in
size. Finally the oocyte (and accompanying nurse
cells), still surrounded by the follicular epithelium,

Fig, 12. — Miastor americana. Longitudinal section through an ovary.

en = envelop; /.fp = follicular epithelium; n.c = nurse chanihcr;

n.c.n = nurse-cell nucleus; o.m = mesoderm; ouc.ti = o(')cyte nucleus.
Fig. 13. — Miastor americana. Longitudinal section through a nearly

full-grown oocyte, g.v = germinal vesicle; n.c = nurse chamlMT;

pPl = pole-plasm.

54 GERM-CELL CYCLE IN ANIMALS

becomes separated from the rest of the ovary and is
forced by the movements of the larva into some other
part of its body. Here it continues its growth and
development at the expense of the tissues of the
mother-larva. Not all of the oocytes (thirty-two
in each ovary) complete their development, since
usually only from five to seventeen young are
produced by a single mother-larva. Those oocytes
that do not perish pass through the stages described
in the following paragraphs.

Figure 13 represents the condition of an oocyte just
before the initiation of the maturation processes.
The nucleus, or germinal vesicle (g.v.), is eccentrically
placed and nearer the anterior than the posterior
end of the cell. The nurse chamber has greatly
decreased in volume.

The contents of the oocyte are not homogeneous,
but several distinct regions can be distinguished.
Near the nurse chamber is a body of cytoplasm
evidently elaborated by the nurse cells, and at the
posterior end is an accumulation which we may call
the pole-plasm (pPl) and which is of particular
interest since it is intimately associated with the
formation of the primordial germ cell.

The maturation division occurs soon after the
stage just described has been attained. The ger-
minal vesicle, which lies near the periphery of the
oocyte, breaks down, and the chromatin contained
within it becomes aggregated into about twenty
chromosomes. As a result of the maturation division
(Fig. 14) a polar body (p.b) and the female pronucleus

THE P^DOGENETIC FLY, MIASTOR 55

(f.n) are produced. The nucleus of the pohir body
divides by mitosis and the two nuclei thus formed

Fig. 14. — Miastor americana. Longitudinal section tlirougli mature
egg. c = cytoplasm; /.?i = female nucleus; n.c = nurse chamber;
p.b — polar bodies; pP^ = pole-plasm.

remain within the egg substance near the periphery
for a considerable period (Fig. 14), but finally

CMp

56 GERM-CELL CYCLE IN ANIMALS

disintegrate and disappear, apparently without
performing any function. As in most other animals,
these polar bodies may be considered abortive eggs.
The female pronucleus moves into the central an-
terior part of
the e^g where
it becomes em-
bedded in the
cytoplasmic
mass near the
nurse chamber.
It may now be
designated as
the cleavage
nucleus, since
the eggs of
Mia^^or develop
without ferti-
lization and
hence no male
pronucleus is
ppl present to unite
with it. The

jTjQ 15 — Miastor mctraloas. Three of the four clcavagC dl-

division figures (I, III, IV) of the four- to eight- yjgJQns take
cell stage represented. cMp = chromosome

middle plate ; n.c = nurse chamber : p.b = polar place by Hll-

body ; pPl = pole-plasm. {From Kahle, 1908.) ^^^^^ ^^^ ^^

in most of the Arthropoda, the early cleavage nuclei
are not separated by cell walls, but simply move
apart after each successive division. The egg during
this period is thus a syncytium within which the
limits of the cells are difficult to define.

THE P^DOGENETIC FLY, MIASTOR 57

The nuclei present at the four-cell stage occupy
rather definite positions and may be numbered for
convenience by the Roman numerals I, II, III,
and IV, as indicated in Fig. 15. The division from
the four- to the eight-cell stage is a very important
one, since it is at this time that the primordial

r.»>.c;-...-.:..;:h

A^-^-iji'/itcV*'"''-''

'i'l-

Fig. 16. — Miastor metraloas. Stages in the chromatin-diminution

process. {From Kahle, 1908.)

germ cell is established. Each of the four nuclei
divides by mitosis, but nuclei I, II, and III undergo
a chromatin-diminution process during which a
large part of their chromatin remains in the cyto-
plasm when the daughter nuclei reform. The details
of such a process are indicated in Fig. 16. Nucleus
IV, on the other hand, divides as usual (Fig. 15) and
each daughter nucleus receives one-half of its chroma-
tin. One of these daughter nuclei becomes embedded
in that peculiar mass of cytoplasm at the posterior

M p

GERM-CELL CYCLE IN ANIMALS

end which we have called the pole-plasm, and ap-
parently all of the pole-plasm, together with this

CM p

cR-

Fig. 17. — Miastor americana. Longitudinal section of egg with one
germ cell (p.g.c.) and nuclei undergoing chromatin-diminution pro-
cess, c = cytoplasm ; c Af p = chromosome middle plate; cR =
chromatin remains.

nucleus, is then cut off from the egg (Fig. 17). This
cell, as has been conclusively proven by studies of

THE PiEDOGENETIC FLY, MIASTOR 59

later stages, is the primordial germ cell. At this
time, then, the egg consists of one primordial germ
cell provided with a nucleus with an undiminished
amount of chromatin, and a syncytium containing
seven nuclei of which the sister nucleus of the primor-
dial germ cell contains a complete supply of chroma-
tin, whereas the other six nuclei have lost part of
this chromatin material. Reference to the diagram
on page 65 will assist in making more clear this stage
and the stages yet to be described.

The next developmental process is the mitotic
division of the seven nuclei in the syncytium thus
producing a fifteen-cell stage (Fig. 17). The sister
nucleus of that of the primordial germ cell now under-
goes a chromatin-diminution process and the other
six nuclei in the syncytium pass through a second
chromatin-diminution process. As a result every
nucleus in the egg has lost a part of its chromatin
except that of the primordial germ cell which still
contains a complete amount. The further history
of the somatic nuclei does not differ essentially
from that of the somatic nuclei in other insects.
They increase in number by mitosis, migrate to
the periphery, and there are cut off by cell walls
forming a single layer of cells over the entire surface
except where interrupted at the posterior end by
the primordial germ cells. Next, a thickening of
the cells occurs on the ventral surface, thus forming
the ventral plate. From this plate most of the
embryo arises ; it lengthens until the anterior or
cephalic end almost reaches the anterior end of the

60 GERM-CELL CYCLE IN ANIMALS

egg, and until the posterior or tail end has been
pushed around for a considerable distance on the
dorsal surface. A broadening and a shortening
of this germ-band then takes place so that the pos-
terior end of the embryo coincides with the posterior
end of the egg and the edges of the embryo grow
laterally around the egg until they meet in the
median dorsal line. Meanwhile various changes
have taken place within the embryo, among which
is the formation of the germ glands or ovaries.

Returning now to a consideration of the germ cells,
we shall see that it is possible to trace the descendants
of the primordial germ cell with comparative ease.
This cell divides by mitosis, forming two oogonia
approximately equal in size (Fig. 18). These two
then produce four oogonia of the second order
(Fig. 19), and these in turn increase by mitosis,
forming eight oogonia of the third order (Fig. 20).
When this stage is reached a period sets in during
which the oogonia do not divide, but are apparently
passively carried about by the somatic tissues as
shown in Fig. 21, where they occupy a position
near the end of the tail fold.

One of the most satisfactory conditions in the
keimbahn of Miastor is the comparatively large
size and peculiar structure of the primordial germ
cells leaving in the mind of the observer no doubt
as to the identity of the cells concerned. Through-
out the entire embryonic development of this insect
the germ cells are considerably larger than any of
the somatic cells. The nuclei are correspondingly

THE P^DOGENETIC FLY, MIASTOR 61

large and are characterized by the possession of a
number of spherical chromatin granules which
are evenly scattered about in the nuclear sap.

Fig. 18. — Miastor nmericana. Longitudinal section through an egg
with two oogonia (oogi). 6 c = blastoderm nucleus; c/i== chro-
matin remains.

Fig. 19. — Miastor americana. Longitudinal section through an egg
with four oogonia {oog^.

GERM-CELL CYCLE IN ANIMALS

Even under the lower powers of the compound micro-
scope the germ cells stand out with great distinct-
ness and could not possibly be confused with any
other cells in the embryo.

c R

Fig. 20. — Miastor americana. Longitudinal section through an egg
with eight oogonia {ooqz). cR — chromatin remains.

Fig. 21. — Miastor americana. Sagittal section through embryo show-
ing oogonia {oog-i) near end of tail fold.

THE P.EDOGENETIC FLY, MIASTOR 63

During the shortening and broadening of the germ
band the group of eight oogonia of the third order
becomes separated into two rows of four each — one
row on either side of the body in the region of the
eleventh segment (Fig. 22). Each group of four
oogonia then becomes surrounded by a layer of
mesoderm cells and forms a more or less spherical
body which may now be called an ovary (Fig. 23).
Soon after this occurs, the oogonia begin to divide
again (Fig. 23, a) and by successive mitoses there
are formed oogonia of the fourth, fifth (Fig. 24), and
sixth orders. This completes the number of oogonia,
which is typically thirty-two in each ovary, and
provides us with the only case thus far on record
where the number of oogonial divisions during the
multiplication period in the history of the germ
cells is known (Fig. 26).

There are then six of these oogonial divisions
between the formation of the single primordial germ
cell and the production of the complete number of
oogonia in the two ovaries. Some of the oogonia of
the fifth order may be prevented from dividing, in
which case of course there are less than thirty-two
germ cells in each ovary. And not all of the oogonia
in the ovary succeed in developing into oocytes and
larvae, since a struggle for supremacy takes place
among the germ cells resulting in the survival of only
a few offspring, as may be determined by the fact,
already referred to, that one larva gives rise as a
rule to onlv from five to seventeen daughter larvae.
Each oogonium that succeeds in developing becomes

GERM-CELL CYCLE IN ANIMALS

00 c

Fig. 22. — Miastor americana.

of embryo showing oogonia
Fig. 23. — Miastor americana.

one dividing by mitosis (o).
Fig. 24. — Miastor americana.

{o6(ji). m = mesoderm.
Fig. 25. — Miastor americana.

(n.c).

Frontal section through posterior end
Koogs) formmg two rows of four each.
Ovary containing sixteen oogonia (odgi),

m = mesoderm.

Ovary containing thirty-two oogonia

Young oocyte {ooc) with nurse cells

Cfr,-

PP'

I^pl ...

st.c

PP' ^^

PP'

..oh

-Pk

,pt.

■ eithf

.P5«~...

ti-Ott

063*...,
00^

OX©

.©X©Y©,

0IO,

90f.

00/-.
Co/

i(fa o_03

onia. omi

tt«i,V

■tOTBditt.

.^f4ljV

_-Ja?-Oo

oooonio. 0 Vi I

UtA.)

^^(bO_oo^oni_flL tO_>nV"tt jAjl.

p.^.

Fig, 26. — Miastor americana. Diagram illustrating origin and history
of germ cells from one generation to the next. d.n = cleavage
nucleus ; ex.chr = extruded chromatin ; oog = oogonia ; p.h = polar
body; p. y. c = primordial germ cell; p.o= primary oocyte; p.pl =
polar plasm, st.c = stem cell. (^*'^)

66 GERM- CELL CYCLE IN ANIMALS

provided with a group of about twenty-four meso-
derm cells which form a syncytium at the anterior
end and may be called nurse cells (Fig. 25, n.c), since
they furnish food material to the oocyte. Another
group of mesoderm cells forms a cellular layer about
the oocyte and nurse cells, and thus constitutes a
follicular epithelium. At this stage the oocytes
break away from the ovary and become distributed
in various parts of the body of the mother-larva.

Several facts regarding the germ-cell cycle of
Miastor deserve special emphasis : (1) There is
no stage in the entire keimbahn when the germ cells
cannot be distinguished without the least difficulty ;
(2) the number of oogonial divisions has been defi-
nitely established, and so it is no longer necessary to
make the general statement that the germ cells
pass through 7i divisions during the period of multi-
plication, since here 7i is undoubtedly six; (3) the
descendants of the primordial germ cell are only
germ cells, i.e., the primordial germ cell does not
give rise to both oogonia and nurse cells as seems to
be the case in most other insects; (4) chromatin-
diminution processes take place during the mitotic
divisions of the nuclei from the four- to the eight-
cell stage and form the eight- to the fifteen-cell
stage of such a nature that all of the cells in the
embryo finally are deprived of part of their chromatin
with the exception of the primordial germ cell which
retains the complete amount of this substance;
(5) the primordial germ cell is estabhshed at the
eight-cell stage and is the first complete cell formed in

THE P^DOGENETIC FLY, MIASTOR 67

embryonic development ; and (6) the contents of
the primordial germ cell consist of the nucleus
with undiminished chromatin and of all of the pole-
plasm and apparently no other part of the egg sub-
stance.

The fact that only the primordial germ cell re-
ceives a complete amount of chromatin is of particu-
lar interest, since a similar condition has long been
known in the case of Ascaris as we shall see later.
It may also be noted in this place that the cyto-
plasmic substance in the primordial germ cell may
be recognized as the pole-plasm in the growing
oocyte. Attempts have been made to determine the
origin of this pole-plasm, but so far without success.
It may be distinguished from the rest of the Qgg con-
tents by its position at the posterior end and because
of its affinity for certain dyes. It appears shortly
before the maturation division is initiated, but no
transition stages have been discovered — it has been
either present or entirely absent in the preparations
thus far studied. If we consider the history of this
substance from the formation of the primordial
germ cell to the growth period of the oocytes pro-
duced by this primordial germ cell, we may conclude
that at the time the multiplication period ends the
pole-plasm has become equally distributed among the
sixty-four oogonia. Then ensues the growth period
during which the pole-plasm cannot be distinguished.
Later, however, just before maturation, pole-j^lasm
substance reappears which is equal in amount to
that contained in the primordial germ cell of the

68 GERM-CELL CYCLE IN ANIMALS

preceding generation or to that contained in all of
the sixty-four oogonia which descended from that
primordial germ cell. That is, the pole-plasm of
the oocyte under discussion has in some way increased
until its mass is sixty-four times as great as that of
the oogonium before the growth period began. How
this increase has taken place can only be conjectured.
The pole-plasm in the oogonium may have produced
new material of its own kind either by the division
of its constituent particles or by the influence of its
presence. In any case a localization of this substance
occurs at the posterior end of the egg just before
maturation. Therefore, although we can follow the
germ cells in Miastor throughout their entire cycle
without difficulty, there are certain problems, such
as the history of the pole-plasm during the growth
period of the oocytes, which still remain unsolved.

CHAPTER IV

THE SEGREGATION OF THE GERM CELLS IN
PORIFERA, CCELENTERATA, AND VERTEBRATA

The history of the germ cells has not been seriously
investigated in a number of groups of animals, but, as
will be demonstrated in Chapters V and VI, there
are many species belonging to widely separated
groups in the animal series in which the germ-cell
cycle is almost as well known as in Miastor. On
the other hand, the three phyla to be discussed in
this chapter have been carefully studied for many
years, but an early segregation of germ cells has not
yet been established in them to the satisfaction of a
majority of investigators. It seems strange because
of the uncertainty of the morphological continuity
of the germ cells in these animals that one of these
groups, the Cgelenterata, should have furnished
the material upon which Weismann based his elabo-
ration of the germ-plasm theory.

1. PORIFERA

Sponges reproduce asexually by budding and by
the formation of gemmules, and sexually by means of
ova and spermatozoa. Budding occurs in almost all
sponges. In most cases the buds remain attached
to the parent (continuous budding) ; but in some

70 GERM-CELL CYCLE IN ANIMALS

species the buds become free (discontinuous bud-
ding) .

Gemmules are groups of cells (statocytes) which
occur at certain times of the year in the bodies of
fresh- water sponges and in many marine species.
These gemmules acquire a resistant covering and
serve to preserve the race during the winter in the
north or the dry season in the south. The peculiar
" budding" observed in Tetkya by Deso (1879, 1880)
may be a sort of gemmule formation (see p. 76).

The eggs and spermatozoa are situated in the
middle layer (so-called mesoderm) and in most
cases seem to become ripe at different times in the
same sponge. Fertilization is apparently similar
to this process in other Metazoa. The fertilized
ovum is holoblastic; the free-swimming ciliated
larva becomes fixed, and then metamorphoses into
a young sponge.

The body wall of the sponge consists of two distinct
layers, an outer dermal layer and an inner gastral
layer, and an intermediate jelly-like stratum con-
taining ameboid wandering cells. The various sorts
of cells in these lavers are indicated in the table on
page 71 (from Minchin, 1900, p. 62).

The reproductive cells lie in the jelly-Kke middle
layer, but all of the cells in this layer are not repro-
ductive.

The origin of the archeocytes from which the re-
productive cells arise can easily be pointed out in
the comparatively simple development of Clathrina
hlanca (Minchin, 1900). In this species a ciliated

PORIFERA, CCELENTERATA, VERTEBRATA 71

Table of the Various Classes of Cells in Sponges

Dermal Layer

I. Epithelial stratum

II. Porocytes

III. Skeletogenous
stratum

Gastral Layer IV. Gastral epithelium

Archaeocytes
(primordial
cells)

V. Amebocytes (wan-
dering cells)

3.
4.

^ 6.

9.
10.
11.
12.
13.
U.

VI. Tokocytes (repro- -j , ^
ductive cells)

Pinacocytes

(epithelial cells)
Myocytes

(contractile
cells)
Gland cells
Spongoblasts
Pore cells
Scleroblasts
CoUencytes

(stellate cells)
Desmac3"tes

(fiber cells)
Cvstencvtes

(bladder cells)
Choanocytes

(collar cells)
Phagocytes

(ingestive cells)
Trophocytes

(nutritive cells)
Thesocytes

(storage cells)
Statocytes

(gemmule cells)
Gonocytes

(sexual cells)

blastula-like larva is formed (Fig. 27, A). At the
posterior pole two blastomeres (posterior granu-
lar cells, p.g.c.) remain undifferentiated ; they are
much larger than the other cells, are granular, and
possess vesicular nuclei. The larva becomes fixed
by the anterior pole, and during the metamorphosis
that then takes place, the two posterior granular
cells, the archeocytes, multiply rapidly, forming a
large number of minute cells which resemble certain
leucocytes. These are known as amebocytes. By

GERM-CELL CYCLE IN ANIMALS

the fourth day the amebocytes become separated
into wandering cells or their derivatives and repro-
ductive cells or tokocytes as indicated in the table.
The primordial archeocytes do not always occur
in the Clathrinidse as in Clathrina hlanca. In some

Fig. 27. — A. Clathrina hlanca. Blastula stage showing posterior gran-
ular cells (p.g.c). (From Minchin, 1900.) B. Oogonium of a
sponge containing inclusions in the cytoplasm. {From Jorgensen,
1909.) C. Two oogonia in the ectoderm of Hydra fusca, each with
a cytoplasmic inclusion. {From Downing, 1909.)

species there is only one; in others four or more
appear ; and sometimes they are entirely absent.
This last condition results from the formation of
amebocytes before the fixation of the larva. In
many other sponges the archeocytes migrate in at
the posterior pole and partially or entirely fill up
the segmentation cavity. Comparatively little is
known about the embrvologv of Hexactinellida and

PORIFERA, CCELENTERATA, VERTEBRATA 73

Demospongiae, and few observations have been
made upon their archeocytes. These archeocytes
are of the greatest importance since they give rise
to the amebocy tes and tokocy tes (reproductive cells) .
According to Weltner (1907) both amebocy tes and
tokocytes are only physiological states of one and
the same kind of cell. Many authors have em-
phasized the importance of the amebocytes, such as
Gorich (1904), who maintains that this class of cells
gives rise not only to the gonocytes, statocytes,
and trophocytes, but also to certain pinacocytes.
Weltner (1907) goes further than this when he states
from studies upon the fresh-water sponge that the
sponge could not exist without amebocytes.

The earlier investigators almost invariably con-
sidered the germ cells as mesodermal in origin.
Lieberklihn (1856) discovered the eggs in Spongilla
and later (1859) in Sycandra raphanus. Sponge
eggs were also observed by Kolliker (1864). Haeckel
(1872) thought that the eggs were derived from the
flagellated cells of the gastral epithelium. Schulze
(1875), on the contrary, maintains that they lie
deep in the so-called mesoderm; and Fiedler (1888)
concludes that in Spongilla only certain cells of the
middle layer may become germ cells.

Maas (1893) distinguished two sorts of cells in the
middle layer ; one characterized by uniform, fine-
granuled cytoplasm and an oval nucleus containing
a very fine net-work of chromatin ; the other filled
with coarse-granuled cytoplasm and a spherical nu-
cleus containing a deeply staining nucleolus and

74 GERM-CELL CYCLE IN ANIMALS

chromatin aggregated into large masses. Only
from the latter do the sex cells arise. These two
kinds of cells could be distinguished in larval stages
and the early separation of germ cells from somatic
cells was pointed out. Maas, however, does not
insist that there is here a demonstrated continuity
of germ cells, since the cells which become sex-cells
are separated from the egg by a long series of genera-
tions.

The recent investigations of Jorgensen (1910)
on Sycon raphanus and S. setosa have added consider-
ably to our knowledge of the origin, structure, and
early history of the germ cells of sponges. Jorgensen
does not agree with Maas (1893) regarding the early
segregation of the germ cells from somatic cells,
but finds no particular difference between so-called
mesoderm cells and wandering or egg cells. It is
worthy of note, however, that the youngest recog-
nizable oogonia were found to contain several distinct
bodies in their cytoplasm (Fig. 27, B).

The method of formation of the gemmules has
engaged the attention of many investigators, but
several important points concerning it are still in
doubt. Gemmule formation is of particular interest
since the cells (amebocytes), which by most authori-
ties are said to give rise to the germ cells, are also
considered the cells which form the reproductive
portion of the gemmules. At least four views have
been held concerning the origin of the gemmule
cells : (1) Carter (1849) believed that the gemmule
is derived from a single cell, the "ovi-bearing cell";

PORIFERA, CCELENTERATA, VERTEBRATA 75

(2) Goette (1886) maintains that the gemmule con-
sists of cells from several germ layers ; (3) Carter
believed at one time that the gemmule was made
up of only one kind of cell ; and (4) several
authors (Marshall, 1884 ; Wierzejski, 1886 ; Zykoff,
1892; Weltner, 1892) believe that a number of
cells belonging to several classes are concerned in
the origin of the gemmule.

Evans (1900) has described in detail the formation
and structure of the gemmules of Ephydatia hlemhin-
gia. In this species the first sign of the formation of a
gemmule is the presence of "single cells or groups
of cells scattered about chiefly in the dermal mem-
brane ; the strands of tissue which support the dermal
membrane ; and in the tissues situated immediately
below the subdermal cavity" (p. 89). No mitotic
figures were discovered in these cells and conse-
quently the reproductive part of the gemmule is
probably not derived from one mother-cell. These
cells wander "through the dermal membrane, and
strands of tissue which support the membrane, and
become aggregated in groups situated either deep
in the tissues of the sponge or even in the strands of
tissue above mentioned."

Whether the reproductive cells of the gemmule
arise from a single cell by proliferation or represent
an aggregation without a common origin is still
unsettled, but the latter view is held by most in-
vestigators. If they do arise from a single cell, as
H. V. Wilson (1902) admits is a possibility, the
gemmule formation may be considered a kind of

76 GERM-CELL CYCLE IN ANIMALS

parthenogenesis. If, on the other hand, the re-
productive cells of the gemmule are of multiple
origin, they may either be looked upon as true germ
cells which form a group physiologically equivalent
to the morula stage in the development of an egg,
or as a collection of regenerative cells capable of
producing a new individual.

In this connection should be mentioned the bud-
ding of Tethya (Deso, 1879-1880) which develops
from a group of amebocytes (Maas, 1910) and the
gemmules of Tedania and Esperella (Wilson, 1902)
and of hexactinellids (Ijima) which become ciliated
larvae. Wilson has shown " that silicious sponges,
when kept in confinement under proper conditions
degenerate in such a manner that while the bulk
of the sponge dies, the cells in certain regions become
aggregated to form lumps of undifferentiated tissue.
Such lumps or plasmodial masses, which may be
exceedingly abundant, are often of a rounded shape
resembling gemmules, more especially the simpler
gemmules of marine sponges (Chalina, e.g.), and
were shown to possess in at least one form (Stylo-
tella) full regenerative power. When isolated they
grow and differentiate, producing perfect sponges "
(1907, p. 295). These "lumps of undifferentiated
tissue" have also been noted by F. E. Schulze
(1904) and recognized as probably reproductive;
they have been named by this author, " sorites," and
have been called by several authors "artificial
gemmules." The process involved in their forma-
tion is termed "regressive differentiation." The

PORIFERA, CCELENTERATA, VERTEBRATA 77

undifferentiated tissue of which they are composed,
undoubtedly consists largely, if not entirely, of
amebocytes (Weltner, 1907). These amebocytes
are, however, of heterogeneous origin (Maas, 1910),
since some of them represent transformed pore
cells, whereas the rest are wandering cells.

Even more interesting than these reproductive
bodies are the artificial plasmodia produced by Wil-
son (1907, 1911) in Microciona, Lissodendoryx, and
Stylotella and by Muller (1911) in the Spongillidae.
The method and results from a study of Microciona
as stated by Wilson (1911) are briefly as follows.
Branched specimens are cut up and strained into a
dish of water through fine bolting cloth. The
cells, which are dissociated in this way, "settle
down on the bottom of the dish like a fine sediment."
Three classes of cells are present : (1) *' the most con-
spicuous and abundant" are unspecialized granular
"ameboid cells of the sponge parenchyma (amoebo-
cytes) " ; (2) "a great abundance of partially
transformed collar cells"; and (3) "more or less
spheroidal cells ranging from the size of the granular
cells down to much smaller ones."

"Fusion of the granular cells begins imme-
diately and in a few minutes' time most of these
have united to form conglomerate masses which at
the surface display both blunt and elongated pseu-
dopodia. These masses (plasmodia) soon begin to
incorporate the neighboring collar and hyaline cells."
"The small conglomerate masses . . . early begin to
fuse with one another," and if the tissue is strewn

78 GERM-CELL CYCLE IN ANIMALS

sparsely over a slide, in the course of a week it will
be found that the slide is covered with a thin in-
crusting sponge provided with pores, oscula, canals,
and flagellated chambers." Many, at the end of
two months, had " developed reproductive bodies
(eggs or asexual embryos?) ..." Whether these
reproductive bodies arose from eggs or masses
of cells was not determined. " When the plasmodia
have metamorphosed and the canals and chambers
have developed, the skeleton makes its appearance."

Experiments with Lissodendoryx and Stylotella
were not quite so successful, but plasmodial masses
were formed in every case. Further experiments
proved that " when the dissociated cells of these
two species [Microciona and Lissodendoryx] are
intermingled, they do not fuse with one another,
but fusion goes on between the cells and cell masses
of one and the same species." A similar result was
obtained by intermingling dissociated cells of Micro-
ciona and Stylotella.

Discussion and Summary. The foregoing ac-
count of the origin of the germ cells in sponges
shows conclusively that these cells arise in the so-
called mesoderm from wandering cells (amebocytes)
and that amebocytes are descended from archaeo-
cytes which may be distinguished in certain cases
very early in embryological development (Fig. 27, A,
p.g.c). Oogonia and spermatogonia have not been
recognized by most investigators except in the adult,
but Maas (1893) has observed them in the planula.
Jorgensen (1910), who has made the most careful

PORIFERA, CCELENTERATA, VERTEBRATA 79

study of the development of the oogonia, states
that the youngest recognizable oogonia lie in the
mesoderm, and his figure (Fig. 27, B) shows that
they may be distinguished from neighboring cells by
certain characteristics, among which is the presence
of a darkly staining inclusion. In the adult sponge
the amebocytes from which the oogonia and sperma-
togonia arise occur in the middle layer of all regions of
the body, but, as pointed out by Korschelt and
Heider (1903), the oogonia and spermatogonia may
develop in only certain definite regions {Plakina
monoloplia), or in groups (Aphysilla violacea) which
contain a more or less definite number of cells and
occupy a similar position in each individual (Eu-
spongia). Such an aggregation is the most primitive
form of ovary.

Some of the amebocytes of the sponge are un-
doubtedly germ cells (tokocytes) and are able to
develop into oogonia or spermatogonia, or to form
aggregations (gemmules, " artificial gemmules," '* so-
rites," etc.) which can "regenerate" an entire sponge,
but whether the amebocytes that produce oogonia
and spermatogonia are the same as the reproductive
cells of the gemmules, the regenerative cells of the
" artificial gemmules," and amebocytes which form
the buds in Tethya is still uncertain. It seems
probable that they are all alike potentially but
develop differently because of the effects of different
environmental factors. The distribution of ame-
bocytes with reproductive powers throughout the
entire sponge-body accounts for the great regenera-

80 GERM-CELL CYCLE IN ANIMALS

tive ability of these animals and must also account
for the development of plasmodia formed by dis-
sociated cells (Wilson, 1911; Muller, 1911) into
adult sponges with all specific characteristics in-
cluding reproductive bodies.

It therefore seems possible that there may exist
in the sponges a continuity of the germ-plasm and
that the germ-cell material is distributed among
thousands of cells (tokocytes, see Table, p. 71)
which are derived from archseocytes, and that under
proper conditions these tokocytes may produce
oogonia or spermatogonia, or may aggregate to
form gemmules or regenerative bodies. This wide
distribution of the germ cells is what might be
expected in such lowly organized animals. Figure
28 shows the probable history of the germ cells in
the PoRiFERA from one generation to the next.

2. CCELENTERATA

The origin of the germ cells in the Ccelenterata
has been a much debated subject among zoologists
for three-quarters of a century. As early as 1843 van
Beneden undertook to determine the germ layer
from which the germ cells arise and concluded that
the ova originate in the entoderm and that the
spermatozoa come from the ectoderm. F. E.
Schulze (1871) claims that in Cordylophora both
the ova and spermatozoa are of ectodermal origin.
Kleinenberg (1872), working on Hydra, announced
that the germ cells are interstitial in origin and,
since the interstitial cells arise from the ectoderm,

PORIFERA, CGELENTERATA, VERTEBRATA 81

Statocytes
{Gemmule cells)

Archeocytes

Fig. 28.

Spermatozoon

■ Diagram illustrating the probable history of the germ cells

in sponges from one generation to the next.
G

82 GERM-CELL CYCLE IN ANIMALS

are therefore also ectodermal. Van Beneden (1874),
from investigations on Hydractinia, Clava, and Cam-
PANULARiD^, confirms his earlier results and again
maintains that the ova arise in the entoderm. The
brothers Hertwig (1878) decided that the germ cells
of Hydromedus^ arise from the ectoderm and those
of the ScYPHOMEDUS^ and Anthozoa from the
entoderm. In a second paper, Kleinenberg (1881)
reports the ova of Eudendrium as of ectodermal
origin. Varenne (1882) maintains that both the
ova and the spermatozoa of half a dozen species
examined arise from entoderm cells of the young
blastostyle before the appearance of medusa buds.
The results of Weismann's extended studies were
published in a monograph (1883), and later (1884) a
brief general account appeared.

From this time until the present day almost every
year has witnessed one or more contributions to the
subject of the origin of the germ cells in ccelenterates,
and a perusal of this mass of literature shows that
the problem is not yet solved.

Hydra. The fresh-water polyp. Hydra, has been
employed for germ-cell investigations more often
than any other coelenterate, and a number of de-
tailed papers have appeared mthin the past ten
years upon this genus. Among the earlier workers
who actually saw the ^gg should be mentioned
Trembley (1744), Rosel V. Rosenhoff (1755), Ehren-
berg (1836) and Leydig (1848). The processes
involved in oogeneses were not clearly determined,
however, until Kleinenberg's classic investigations

PORIFERA, CCELENTERATA, VERTEBRATA 83

in 1872, upon which most of the accounts in our
zoological textbooks are still based. Kleinenberg's
researches were followed by those of Korotneff
(1883), Nussbaum (1887), Schneider (1890), and
Brauer (1891). Investigations of the germ cells of
Hydra then almost ceased until 1904, when another
period of activity in this field began and papers
quickly followed one another (Guenther, 1904 ;
Downing, 1905; Hadzi, 1906; Hertwig, R., 1906;
Tannreuther, 1908, 1909; Downing, 1909; and
Wager, 1909). The following account is based
chiefly upon the researches of Downing (1905,
1908, 1909), Tannreuther (1908, 1909), and Wager
(1909).

The origin of the male germ cells has been carefully
investigated by Downing (1905) and Tannreuther
(1909). Previous to Downing's researches all in-
vestigators, beginning with Kleinenberg (1872),
considered the sex cells as interstitial in origin.
Downing, however, believes that germ cells and in-
terstitial cells may be distinct. The sex cells,
according to this investigator, are distinguished
"by their very large nuclei, extremely granular,
and often by the presence of a Nebenkern " (Fig.
27, C). "The characters of the sex cells . . .
seem constant, and my conclusion would be that at
some stage of the embryonic development certain
cells are stamped with these characters and that they
and their progeny form the sex cells distinct through-
out the life of the individual . . . the germ-plasm is
then continuous in Hydra " (p. 413). This tentative

84 GERM-CELL CYCLE IN ANIMALS

opinion is expressed with more certainty in a later
paper (Downing, 1909), since the "distinctive charac-
ter of the germ cell is more marked in the ovary than in
thespermary" (p. 311). Tannreuther (1909), on
the other hand, claims that the male germ cells are
interstitial in origin, and " the progenitors of the
spermatozoa have no special characters by which
they can be recognized as germ cells."

The origin of the eggs of Hydra is better known
than that of the male germ cells. The ova have by
most investigators been considered modified intersti-
tial cells. Downing (1908, 1909) disagrees in several
respects with the results of Tannreuther and Wager.
His most important difference is regarding the ques-
tion of the origin of the ova directly from interstitial
cells or from definite propagative cells that are set
aside for reproductive purposes at some stage in the
animal's embryonic development. He believes " that
in the adult Hydra the oogonia (and spermatogonia)
are distinctly differentiated as a self-propagating
tissue" (p. 310). Wager (1909), on the contrary,
claims that it is impossible to prove that eggs do
not arise from ordinary interstitial cells; whereas
Tannreuther (1909) finds that the primitive ova can
be distinguished from interstitial cells "by their
large nucleus, nucleolus, and abundance of chromatin,
even before the growth of the ovary begins" (p. 205),
especially during the breeding season, and admits
that "If these sex cells could be distinguished during
the budding season as well, it would at least suggest
specificity of the germ cells " (p. 205).

PORIFERA, CGELENTERATA, VERTEBRA FA 85

By far the most important question arisin^^ from
a study of the origin of the germ cells of Hydra is
whether these cells arise from ordinary interstitial
cells, as is claimed by most investigators, or whether
they originate from cells that are set aside for re-
productive purposes at some stage of development,
as Downing maintains. If the latter be true, "the
germ-plasm is then continuous in Hydra ^ (Downing,
1905, p. 413).

Wager (1909) thinks the presence of special prop-
agation cells to be "extremely improbable" and
Tannreuther (1909) does not believe the known
facts warrant the view that there is continuity
of the germ-plasm in Hydra. This is, of course, a
matter that may never be decided definitely, and at
least not until some method of distinguishing
the primordial germ cells, if these be present, from
ordinary interstitial and other cells, has been found.
Furthermore, if the germ-plasm is continuous, primor-
dial germ cells must be present in buds, in adults at
all times of the year, and in pieces of tissue that are
capable of regenerating sexually reproductive adults.
That such primordial germ cells exist seems to me to
be quite possible.

Hydrozoa. Many Hydrozoa besides Hydra have
furnished material for germ-cell studies. Thus
Weismann (1883) reported upon about forty species
belonging to a number of different families. The
results of the researches of the various investigators
do not agree in many instances. In order to indicate
the variety of the opinions expressed, the data re-

86 GERM-CELL CYCLE IN ANIMALS

garding the germ cells in the following genera is
considered below : (1) Eiidendrium, {%) Hijdractinia,
(3) Pennaria, and (4) Clava,

EuDENDRiUM. Fivc spccies of this genus have
been investigated. In E. racemosum, according to
Weismann (1883), the ova arise in the ectoderm and
the male germ cells originate either from entoderm
cells or from ectoderm cells that later migrate into the
entoderm. Ischikawa (1887) asserts that the germ
cells arise in the ectoderm and migrate into the en-
toderm, and Hargitt (1904a) found ova in both
the ectoderm and entoderm, but, since those in the
entoderm were always the smaller, he concludes that
they may have wandered into that layer from the
ectoderm, though such a migration was not ob-
served.

In E. capiUare Hargitt found ova in the entoderm
except in one case where they occurred in the ecto-
derm. This author also reports the female germ
cells of E. tenue and E. racemosum from the entoderm
only. The ova of the Eudendrid^ when first dis-
tinguishable "are slightly larger than the ordinary
cells of the surrounding tissue, and differ also in
shape, being generally ovoid or spherical and with
comparatively conspicuous nuclei. . . . Growth at
this period would seem to take place in situ, through
the direct nutritive activity of the surrounding tissue
cells. ... As growth continues, the ova become
more or less amoeboid, migrating toward the gono-
phore region, where they seem to aggregate in con-
siderable numbers, the presence of which may act as a

PORIFERA, CCELENTERATA, VERTEBRATA 87

stimulus from which results the formation of the
gonophore" (Hargitt, 1904 a, pp. 261-262).

Hydractinia has been investigated by van
Beneden (1874), Weismann (1883), Bunting (1894),
and Small wood (1909). Weismann considered the
ectoderm of the blastostyle to be the probable place
of origin of the germ cells in this genus. Bunting
(1894) was unable to trace the ova to this layer,
although she found them to be quite abundant in
the entoderm of the blastostyle, even before the gono-
phore appeared. According to this author the ova
apparently arise in the entoderm of the blasto-
style, and "reach maturity on the outside wall of
the spadix, lying between the endoderm and the
inner layer of the bell nucleus. The spermatozoa
arise from the inner layer of the bell nucleus ; we
see that they are, therefore, ectodermal in origin "
(p. 228).

These results are not confirmed by the researches
of Small wood (1909), who finds that the eggs arise
in the entoderm in any region of the polyp, at the
base, the side of the polyp, or in the gonophore.
They may be distinguished from other entoderm
cells by the larger size of the nucleus.

In Pennaria cavolini the germ cells arise in the
ectoderm, according to Weismann (1883), and this
conclusion is confirmed for the ova by Hargitt
(19046). In P. tiarella the germ cells are likewise of
ectodermal origin (Smallwood, 1899, Hargitt, for
the ova, 19046). The eggs of this species arise
in the ectoderm of the manubrium and grow by

88 GERM-CELL CYCLE IN ANIMALS

engulfing other primitive ova. Only six or eight,
rarely more, of the eggs survive.

In Clava, according to van Beneden (1874), the
ova arise in the entoderm. Weismann (1883) was
not able to determine whether they originated in
the entoderm or migrated into that layer from the
ectoderm, but he was certain that the male germ cells
were ectodermal. This conclusion regarding the
male germ cells was confirmed by Thallowitz (1885).
Harm (1902) was able to trace the primitive germ
cells back to a very early stage, and could distinguish
them in even young hydranths. The oocytes dif-
fered from the remaining ectoderm cells in the pos-
session of a larger amount of cytoplasm, a larger
nucleus with a big nucleolus, and an ameboid shape.

Hargitt (1906), working on Clava leptostyla, comes
to conclusions different from those of Harm on C.
squamata. He says "that eggs probably never arise
in the ectoderm but always in the entoderm of the
peduncle of thegonophore, or in that of the polyp very
near the base of the gonophore. . . . Clava, like
other Hydroids, has its breeding season, during which
the germ cells are extremely abundant, and at other
times these cells are either entirely absent or very
scarce" (p. 208). Concerning the early origin
of germ cells Hargitt says, '" it may not be im-
possible that ' Urkeimzellen ' should perhaps exist in
undifferentiated stages, still the probability is so
extremely remote as to render doubtful to a degree
any but the most thoroughly substantial claims "
(p. 209).

PORIFERA, (XELENTERATA, VERTEBRA TA 89

One more Hydrozoon may be mentioned —
GonothyroBa loveni — since Wulfert (lOO'^) traced
the germ cells of this species back to the planula
stage where they arise from the interstitial cells of
the ectoderm and later undergo characteristic
migrations.

Our knowledge of the origin of the germ cells
in other coelenterates is very fragmentary and even
less decisive than that of the Hydrozoa. For this
reason a consideration of the subject is omitted here.

Discussion. As in the Porifera we are here
confronted with the question whether or not there is
continuity of the germ-plasm in the Ccelenterata.
There is sufficient evidence for the belief that the
cells which develop into germ cells are not derived
from the ectoderm or the entoderm but belong to a
special sort of propagative cells which, are scattered
about among the other cells throughout the body
and which give rise to ova or spermatozoa under
certain environmental conditions differing in the
different species. This conclusion is based partly
upon the results of Downing (1905, 1908, 1909),
who still holds, as stated in his published papers,
that there is continuity of the germ-plasm in Hydra ;
and upon the fact that germ cells have been recog-
nized in the young hydrant hs of Clava (Harm, 190'-2)
and in the planula of Gonothyroea (Wulfert, 190^2).
It seems certain that more careful studies of the
early stages of coelenterates with special regard
to the origin of the germ cells and with the use of
many and varied stains would result in the discovery

GERM-CELL CYCLE IN ANIMALS

Fig. 29. — Diagram to illustrate the phylogenetic
shifting back of the origins of the germ cells in
medusoids and hydroids. A composite picture.
A, branch of a polyp-colony; P, poljiJ-head
with mouth (m) and tentacles ; »S^, stalk of the
polyp ; M, medusoid-bud with the bell (Gl) ;
T, marginal tentacle; m, mouth; Mst, ma-
nubrium ; GphK, a gonoph ore-bud ; GH, gas-
tric cavity ; ekt, ectoderm ; ent, endoderm ;
st, supporting lamella. The germ cells (kz)
arise in the medusoid in the ectoderm of the
manubrium — first phyletic stage — where they
also attain maturity. In the gonophore-bud
(GphK) they arise in the ectoderm (kz^), or
further down in the stalk of the polj^) at kz'^
— third phyletic stage — or in the ectoderm of
the branch from which the polj'p has arisen,
at kz'" — fourth phyletic stage of the shunting
of the originative area of the germ cells. In
the last two cases the germ cells migrate until
they reach their primitive place of origination
in the medusoid, or in the corresponding layer
of the medusoid gonophore, as may be more
clearly seen in Fig. 30. (After Weismann, 1904-)

of these cells in
younger em-
bryos than yet
recorded, and
might even dis-
close charac-
teristics which
would enable
us to trace the
keimbahn in
some species
back into the
early cleavage
stages.

In discussing
the germ cells
of coelenterates,
it is necessary
to refer to the
work of Weis-
mann who has
added so much
to our knowl-
edge of this
subject. Weis-
mann's position
may best be
presented in his
own words {The
Evolution The-
ory, Vol. I, pp.
413-415, 1904).

PORIFERA, CCELENTERATA, VERTEBRATA 91

"In the hydroid polyps and their medusoids the
germ-cells always arise in the ectoderm ; in species
which produce sexual medusoids by budding, the
germ cells arise in the ectoderm of the manubrium
of these medusoids (Fig. 29, M, kz). But in many
species these sexual stages have degenerated in the
course of phylogeny into so-called gonophores,
that is, to medusoids which still exhibit more or less
complete bells, but neither mouth (m) nor marginal
tentacles {T)^ and which no longer break away
from the colony to swim freely about, to feed in-
dependently, and to produce and ripen germ-cells.
The degeneration of the 'gonophores' often goes
even farther ; in many the medusoid bell is repre-
sented only by a thin layer of cells, and in some even
this token of descent from medusoid ancestry is
absent, and they are mere single-layered closed
brood-sacs (Fig. 30, Gph).

"The adherence of the sexual animal to the hydroid
colony has, however, made a more rapid ripening of
the germ-cells possible, and nature has taken advan-
tage of this possibility in all cases known to me, for
the germ-cells no longer arise in the manubrium of
the mature degenerate medusoid, that is, of the
gonophore, but earlier, before the bud which becomes
a gonophore possesses a manubrium. The birth-
place of the germ-cells is thus shifted back from the
manubrium of the medusoid to the young gono})hore-
bud (Fig. 29, M, kz). The same thing occurs in
species in which the medusoids are liberated, but live
only for a short time, for instance, in the genus

GERM-CELL CYCLE IN ANIMALS

Fig. 30. — Diagram to illustrate the migra-
tion of the germ cells in hydromedusae
from their remotely shunted place of origin
to their primitive place of origin in the
gonophore, in which they attain to ma-
turity. The state of affairs in Eudendrium
is taken as the basis of the diagram, mu,
mouth ; ma, gut-cavity ; t, tentacle ; Sta,
stem; A, a branch of the polyp-colony;
SP, lateral polyp; Gph, a medusoid-bud
completely degenerated into a mere gono-
phore ; Ei, ovum ; GH, gastric cavity ;
st, supporting lamella. The originative
area of the germ cells lies in the stem of
the principal pob-p at kz"", whence the
germ cells first migrate into the endo-
derm of the branch (A) at kz'", creeping
within which they reach kz" in the lat-
eral polyp (blastostyle), finally reaching
the gonophore {kz) and passing again
into the ectoderm. (^After Weismann,
1904.)

Podocoryne. Al-
though perfect
medusoids are
formed, these
have their germ-
cells fully devel-
oped at the time
of their liberation
from the hydroid
colony. But in
species in which
the medusoid-
buds have really
degenerated and
are no longer lib-
erated, the birth-
place of the germ-
cells is shifted
even farther back^
and in the first
place into the
stalk (SU hz") of
the polyp from
the gonophore-
buds. This is the
case in the genus
Hydractinia. In
the further course
of the process the
birthplace of the
germ -cells has

PORIFERA, CCELENTERATA, VERTEBRATA 93

shifted as far back as to the branch from which
the polyp has grown out (Fig. 29, A, kz'") ; and
finally, in the cases in which the medusoid has
degenerated to a mere brood-sac (Fig. 30, Gph),
even to the generation of polyps immediately
before, that is, into the polyp-stem from which the
branch arises that bears the polyps producing
the gonophore-bud (Fig. 30, kz'"). Then we find
the birthplace of the germ-cells still further back
(Fig. 30, kz""), for the egg and sperm cells arise
in the stem of the principal polyps (the main stem
of the colony). The advantage of this arrangement
is easily seen, for the principal polyp is present
earlier than those of the secondary branches, and
these again earlier than the polyp which bears the
sexual buds, and this, finally, earlier than the sexual
bud which it bears. Thus this shunting backwards
of the birthplace of the germ-cells means an earlier
origin of the primordium (Anlage) of the germ-cells,
and consequently an earlier maturing of these.

" But none of these germ-cells come to maturity in
the birthplace to which they have been shifted,
for they migrate independently from it to the place
at which they primitively arose, namely, into the
manubrium of the medusoid, which is still present
even when great degeneration has occurred, or even —
in the most extreme cases of degeneration — into the
ectoderm of the brood-sac. This is the case in the
genus Eudendrium, of which Fig. 30 gives a diagram-
matic representation.

"The most interesting feature of this migration of

94 GERM-CELL CYCLE IN ANIMALS

the germ-cells is that the cells invariably arise in
the ectoderm {kz""), then pierce through the sup-
porting lamella {st) into the endoderm {kz'^^), and
then creep along it to their maturing-place. Once
there, they break through again to the outer layer of
cells, the ectoderm {kz), and come to maturity {Ei).
That they make their way through the endoderm is
probably to be explained by the fact that they are
there in direct proximity to the food-stream which
flows through the colony {GH = gastric cavity),
and they are thus more richly nourished there than
in the ectoderm. But, although this is the case,
they never arise in the endoderm ; in no single
case is the birthplace of the germ-cells to be found
in the endoderm, but always in the ectoderm, no
matter how far back it may have been shunted.
Even when the germ-cells migrate through the en-
doderm, their first recognizable appearance is in-
variably in the ectoderm, as, for instance, in Podo-
coryne and Hydr actinia. The course of affairs is
thus exactly what it would necessarily be if our
supposition were correct, that only definite cell-
generations — in this case the ectoderm-cells —
contain the complete germ-plasm. If the endoderm-
cells also contained germ-plasm it would be hard
to understand why the germ-cells never arise from
them, since their situation offers much better con-
ditions for their further development than that of
the ectoderm-cells. It would also be hard to under-
stand why such a circuitous route was chosen as that
exhibited by the migration of the young germ-cells

PORIFERA, CCELENTERATA, VERTEBRATA 05

into the endoderm. Something must he hickini.^ in
the endoderm that is necessary to make a cell into a
germ-cell : that something is the germ-plasm."

Several important contributions have appeared
within recent years which seem to deprive Weis-
mann's contentions of much of their importance.
For example, Goette (1907) has found that the germ
cells of many Hydromedus^ niay arise in the en-
toderm or in the ectoderm, and that in Clava multi-
cornis the germ cells are transformed half-entoderm
cells. After a long series of studies on coelenterate
development C. W. Hargitt (1911) has attacked
Weismann's position in the following words : " That
there is any such region as may be designated a
'Keimzone' or 'Keimstatte' may be at once dis-
missed as absolutely without warrant as a general
proposition. Furthermore, that the germ cells have
their origin in the ectoderm alone in hydromedusse
may be similarly denied and dismissed as unworthy
of further inquiry or doubt. And still further, I am
thoroughly convinced that the still more recent
controversy as to the hypothesis of the 'germ-plasm,'
if not as clearly a delusion as the preceding, is yet
without the slightest support from the ontogeny of
the group under review.

"It is a matter of easy demonstration that in many
species of hydroids the egg may be followed in every
detail from its origin as an ectoderm or an entoderm
or interstitial cell through its gradual differentiation
and growth to maturation, as a distinct individual
cell, without the slightest tendency to multiplication."

96 GERM-CELL CYCLE IN ANIMALS

"It is passing strange that he should ignore the
body of facts concerned in regeneration, and among
them the reproductive organs. And it is still more
strange that in support of this he should cite in
detail the Hydrozoa as illustrating and supporting
the hypothesis, ignoring the well-known facts that
among these are abounding evidences which afford
insuperable objections to just these assumptions.
The present author has, in many cases, shown that
gonads may be as readily regenerated by hydroids
and medusae as any other organs ; and that not for
once or twice, but repeatedly in the same specimen,
and that de novo and in situ; not the slightest evi-
dence being distinguishable that any migration
through preexisting * germ-tracks' occurred. The
assumption that in these animals the gonads have
* been shifted backwards in the course of phylogenetic
evolution, that is, have been moved nearer to the
starting point of development' seems so at variance
with known facts as to be difficult to appreciate or
respect."

Professor Hargitt finally concludes with the fol-
lowing sentence : "I believe the foregoing facts
must suffice to show that, both as to origin, differen-
tiation, and growth, the germ-cells of the Hydrozoa,
so far from sustaining the doctrine of the germ-
plasm, afford the strongest and most direct evidence
to the contrary."

G. T. Hargitt (1913) has also discovered facts
regarding the history of the germ cells in coelenter-
ates which are decidedly opposed to Weismann's

PORIFERA, CCELENTERATA, VERTEBRATA 97

views. He finds that "The egg cells of Campanularia
flexuosa arise in the entoderm of the pedicel of the
gonophore, by the transformation of a single
epithelial cell, or from the basal half of a divided
cell, the distal half of which remains an epithelial
cell and retains its epithelial functions. Therefore
the egg cells have come from differentiated body-
cells (so-called) and there is no differentiation of
the germ-plasm in the sense that germ-cells are
early differentiated and set aside and do not partici-
pate in the body functions. Any cell of the ento-
derm of Campanularia flexuosa may become an egg
cell if it is in the position of the developing gono-
phore " (p. 411).

In spite of these attacks upon the germ-plasm
theory as applied to ccelenterates, the possibility
and even probability of such a condition seems to
the writer to exist, and he is inclined to accept
Downing's position in the matter. Weismann's
views must, however, be modified, since the germ
cells are not ectoderm cells, as he claims, nor do
they belong to any germ layer. They are, according
to the view adopted here, set aside as a separate
class of cells at some stage during early development,
are scattered about among the cells of the ectoderm
or entoderm, depending upon the species, or lie in
the mesoglea. We know that external conditions
may stimulate reproductive activity in certain
ccelenterates (Frischholz, 1909) and consequently
the development of germ cells, and we must conclude
that these germ cells are present at all times in a

98 GERM-CELL CYCLE IN ANIMALS

more or less dormant condition, just as they are in
more complex animals. Furthermore, the germ cells
must be widely scattered, as has been shown by Harm
(1902) in the young hydranths of Clava, by Wulfert
(1902) in the planula of Gonothyrcea, and by Small-
wood (1909) in the polyp of Hydractinia. This wide
distribution of primitive germ cells accounts for the
reproductive powers of regenerated pieces of hy-
droids.

3. Vertebrata

Efforts have been made by many investigators to
trace the keimbahn in vertebrates, but thus far no
method has yet been devised which will enable us
to distinguish germ cells from other cells in the early
embryonic stages. That we shall be able to recognize
germ cells in still earlier stages of development than
has yet been accomplished seems certain, and the
recent contributions of Rubaschkin (1910), Tschasch-
kin (1910), von Berenberg-Gossler (1912a) and
Swift (1914) have already made considerable ad-
vances by the use of some of the more modern cyto-
logical methods. Three principal theories have been
advanced regarding the origin of the germ cells in
vertebrates, and these will be briefly stated before
the histories of the germ cells in special cases are
discussed.

The germinal epithelium theory was advanced by
Waldeyer in 1870. At that time nothing was known
regarding the migration of germ cells during the
embryonic development of vertebrates, and it is

PORIFERA, CCELENTERATA, VERTEBRATA 99

not strange that he should have come to the con-
clusion that the primordial ova arise from the
epithelial cells of the genital ridge among which they
were observed. Although this theory was accepted
by most embryologists, it has gradually been aban-
doned until now it has very few supporters.

The gonotome theory resulted from the studies of
Ruckert (1888) and Van Wijhe (1889). The germ
cells appeared to these investigators to arise in a
part of the segmental mesoblast of the embryo to
which the latter applied the term 'gonotome.'
From the gonotome they become embedded in the
peritoneum. Thus the same cells are recognized as
germ cells by the adherents of both theories, but a
difference exists regarding their origin.

The theory of early segregation has become the
most prevalent view of the origin of the germ cells
of vertebrates, although there are many who still
hold one of the other hypotheses. According to
this theory the germ cells are set aside during the
early embryonic stages before definite germ layers
are formed, and they later arrive at the germinal
ridge either by their own migration or by changes in
the position of the tissues during development.
The germinal epithelium theories have little if any
evidence in their favor, since no one has actually ob-
served a transformation of peritoneal or mesoblast
cells into germ cells. On the other hand, there is an
abundance of proof that these cells migrate from
some distance into the position of the sex glands.

According to Dustin (1907), Firket (1914) and

100 GERM-CELL CYCLE IN ANIMALS

several others there are two methods of origin, and
primary and secondary sex cells are produced. The
former are probably derived from the blastomeres;
whereas the secondary sex cells are entirely inde-
pendent and arise from the coelomic epithelium.

The first statement of the theory of early segre-
gation was made by Nussbaum (1880), who studied
the history of the germ cells in the trout. Following
Nussbaum, Eigenmann (1892, 1896) contributed to
the support of the theory by his investigations on
the viviparous teleost, Cymatogasier. This proved
to be excellent material for such studies and led
Eigenmann to the conclusion that the germ cells
are set aside in this fish during the early cleavage
stages of the egg, probably at the thirty-two cell
stage. In other cases it has been impossible to
trace the germ cells back to such an early embryonic
condition, but nevertheless the evidence has been
almost uniformly in favor of early segregation.
Some of those who have advocated such an early
origin of germ cells are Wheeler (1900) in the lamprey,
Beard (1900, 1902) in Raja and Pristiurus, Nussbaum
(1901) in the chick. Woods (1902) in Acanthais,
Allen (1906, 1907, 1909, 1911) in Chrysemys, Rana,
Amia, and Lepidosteus, Rubaschkin (1907, 1909,
1910, 1912) in the chick, cat, rabbit, and guinea-pig,
Kuschakewitsch (1908) in Ra7ia, Jarvis (1908)
in Phrynosoma, Tschaschkin (1910) in the chick,
von Berenberg-Gossler (1912) in the chick, Schapitz
(1912) in Amhlystoma, Fuss (1912) in the pig and
man, and Swift (1914) in the chick. This is by no

PORIFERA, COELENTERATA, VERTEBRATA 101

means a complete list but indicates the range of
forms studied and the current interest in this subject.

Some of the characteristics by means of which
germ cells can be distinguished in vertebrate embryos
are as follows : (1) the presence of yolk, (2) an
ameboid shape, (3) large size, and (4) slight staining
capacity. By sectioning embryos of various ages
the changes in position of the germ cells can be fol-
lowed with considerable accuracy. Most investi-
gators agree that the movement of the germ cells from
the tissues where first observed to the genital ridge
is caused by ameboid activities of the cells themselves
and by changes in the position of the organs of the
embryo. The paths of migration of four verte-
brates, a turtle, Chrysemys, sl frog, Rana, the gar
pike, Lepidosteus, and the fresh- water dogfish, Amia,
are shown in Fig. 6. For example :

*'In Lepidosteus the sex-cells [Fig. 6, 3, SI] first
seen in the ventral and lateral portions of the gut-
entoderm [Int] migrate to occupy a position in the
dorsal portion of it, from which they pass dorsally
into the loose mesenchyme that forms the substance
of the developing mesentery [l/e^]. As the mesen-
tery becomes more narrow and compact, owing to
the increase in size of the body cavity, the sex cells
migrate to its dorsal portion and laterally to
the sex-gland anlagen (Fig. 6, 4» Sc). Roughly
speaking, one-half of the total number of sex-cells
reach the sex-gland anlagen, the remainder being
distributed between the intestinal entoderm, the
mesodermal layers of the intestine, the mesentery,

102 GERM-CELL CYCLE IN ANIMALS

and the tissues at and dorsal to the root of the
intestine" (Allen, 1911, p. 32).

Of the more recent investigations, facts discov-
ered by Dodds (1910), Rubaschkin (1910, 1912),
Tschaschkin (1910), von Berenberg-Gossler (1912),
and Swift (1914) are especially worthy of mention.
Dodds (1910) found that in the teleost, Lophius,
the germ cells in the embryos cannot be definitely
distinguished previous to the appearance in their
cytoplasm of a body which stains like a plasmosome
(Fig. 31, A). Germ cells are undoubtedly segregated
before this period, but they exhibited no characteris-
tics with the methods employed which rendered them
distinguishable. Dodds believes that this cyto-
plasmic body is extruded plasmosome material,
probably part of one of the two plasmosomes pos-
sessed by many of the cells at this period.

Rubaschkin, in 1910, announced the results ob-
tained with the eggs of the guinea-pig by certain
methods designed to bring into view the chondrio-
somes. He shows that the chondriosomes of the
undifferentiated cells are granular, and that as
differentiation proceeds, these granules unite to
form chains and threads (Fig. 31, B). The sex
cells, however, retain the chondriosomes in their
primitive granular form, and remain in an undiffer-
entiated condition situated in the posterior part of
the embryo among the entoderm cells. Tschaschkin
(1910), in the same year, came to a similar conclusion
from studies made with chick embryos. Rubaschkin
(1912) has also extended his investigations on guinea-

PORIFERA, COELENTERATA, VERTEBRATA 103

pig embryos. The accompanying diagram (Fig. 3^2)
shows the fertilized egg and the early cleavage cells
all alike (in black) ; some of their descendants become
differentiated into the somatic cells of the germ

Fig. 31. — Germ cells of vertebrates. A. From embryo of the teleost,
Lophius, with plasmosome (?) extruded into cytoplasm. (From
Dodds, 1910.) B. One germ cell and four somatic cells from a
guinea-pig embryo. {From Rubaschkin, 1912.) C. Germ cell of
chick showing " Netzapparat." {From von Berenberg-Gossler, 1912.)
D. Primordial germ cell {g) and blood cell (6) in lumen of blood
vessel {D of a nineteen somite chick embryo, a = attraction-sphere.
(From Swift, 1914.)

layers (circles), but others (in black) remain in a
primitive condition and are recognizable as the
primordial germ cells ; these remain at rest for a
considerable period, but finally multiply and become
part of the germinal epithelium (g.ep).

104 GERM-CELL CYCLE IN ANIMALS

Von Berenberg-Gossler (1912) considers the "Netz-
apparat" in the primitive germ cells of the chick of
particular importance (Fig. 31, C), comparing it
with the "wurstformige Korper " described by Hasper

>d<

FiG. 32. — Diagram to show the history of the germ cells in the embryo
of the guinea-pig. g.ep = germinal epithelium. (From Rubasch-
kin, 1912.)

(1911) in Chironomus (p. 108, Fig. 33). The ap-
pearance of this structure in *'Keimbahnzellen"
is thought to be due to the long period during which
these cells do not divide. Duesberg (191*2), however,
after an exhaustive review of the literature on this

PORIFERA, CCELENTERATA, VERTEBRATA 105

structure concludes that it is not a special cell organ
but an artifact. Kulesch (1914), on the contrary,
finds it to be a constant organ in the eggs of the cat,
dog, and guinea-pig.

The evidence of a continuous germ-cell cycle in
the vertebrates is more convincing than in the
sponges and coelent crates, and leads us to predict
that it will not be long before the gap still existing
during which germ cells cannot be recognized will
be filled in to the satisfaction of the majority of
investigators.

CHAPTER V

THE SEGREGATION OF THE GERM CELLS IN

THE ARTHROPODA

1. The Keimbahn in the Insects

The insects have furnished a very large proportion
of the data upon which many of our biological
conceptions are now based, and they are becoming
more and more popular for studies of the physical
basis of heredity, and for purposes of animal breeding.
It was in insects (Miastor) that the early segrega-
tion of the germ cells in animals was first definitely
established. The accessory chromosome was dis-
covered in insects by Henking in 1891, and our
knowledge of the chromosomes, which has increased
so remarkably within the past fifteen years, is due
principally to the study of oogenesis and spermato-
genesis in insects. In this chapter the chromosomes
will only be considered incidentally, a more detailed
account being deferred until later (Chapter IX).
The early history of the germ cells in insect develop-
ment has not been slighted, for there are many
reports based on this subject alone and still more
data hidden away in contributions on general em-
bryology. It will be necessary here to select from
this abundance of material those reports that give
us the clearest pictures of the keimbahnen. As

106

GERM CELLS IN THE ARTHROPODA 107

usual, certain species or groups of species have
proven more favorable than others for germ-cell
studies, especially those belonging to the orders
DiPTERA, CoLEOPTERA, and Hymenoptera.

DiPTERA. Robin, in 1862, described what he
called "globules polaries" at one end of the nearly
transparent eggs of the crane fly, Tipulides culici-
formes, and the following year Weismann (1863) re-
ported the formation of similar cells, the "Pol-
zellen" at the posterior end of the eggs of the midge,
Chironomus nigroviridis, and the blow fly, Calliphora
(Musca) vomitoria. It remained for Leuckart (1865)
and Metchnikoff (1865, 1866), however, to identify
the pole cells (in Miastor) as primordial germ cells ;
their results were confirmed for Chironomus by
Grimm (1870) and Balbiani (1882, 1885).

Pole cells have also been described among the
DiPTERA, in Musca by Kowalevsky (1886), Voeltz-
kow (1889), and Escherich (1900); in Calliphora
by Graber (1889) and Noack (1901) ; in Chironomus
by Ritter (1890) and Hasper (1911) ; in Lucilia by
Escherich (1900) ; in Miastor by Kahle (1908) and
Hegner (1912, 1914a), and in Compsilura by Hegner
(1914a).

Four genera of flies will serve to illustrate the
methods of germ-cell segregation in this order : (1)
Chironomus (Ritter, 1890; Hasper, 1911), (2) Cal-
liphora (Noack, 1901), (3) Miastor (Kahle, 1908;
Hegner, 1912, 1914a), and (4) Compsilura (Hegner,
1914a). Since Miastor has been discussed in detail
in Chapter III it will be only briefly referred to here.

108 GERM-CELL CYCLE IN ANIMALS

We owe the first accurate account of the germ
cells in Chironomus to Ritter (1890), who, by means
of the section method, showed that the "y^lk
granules" described by Weismann (1863) in the
pole cells are derived from a disc-shaped mass of
substance situated near the posterior end of the egg
and termed by him the "Keimwulst." Hasper
(1911) was able to confirm this discovery, to add
other interesting facts, and to correct several of
Ritter 's errors. The "Keimwulst" of Ritter is
called by Hasper the "Keimbahnplasma."

Ritter advanced the idea that the cleavage
nucleus of Chironomus divides within the "Keim-
wulst" and that here the first cleavage division
occurs, one daughter nucleus remaining in the "Keim-
wulst" and becoming the center of the primordial
germ cell, the other giving rise to somatic nuclei.
This is probably the basis for Weismann's (1904)
statement regarding his conception of the germ-
plasm that, "If we could assume that the ovum,
just beginning to develop, divides at its first cleavage
into two cells, one of which gives rise to the whole
body (soma) and the other only to the germ-cells
lying in this body, the matter would be theoretically
simple. ... As yet, however, only one group of
animals is known to behave demonstrably in this
manner, the Diptera among insects. . . ." There
is, however, nothing in the literature to warrant
the above statement, since Ritter's hypothesis
has been disproved by Hasper.

According to Hasper one of the cleavage nuclei

GERM CELLS IN THE ARTHROPODA 109

at the four cell stage becomes separated from the
rest of the egg, together with all of the Keimbahn-
plasma as the primordial germ cell (Fig. 33 B,
p.g.c). The Keimbahnplasma is apparently equally
divided between the daughter cells when the
primordial germ cell divides. Later the nuclei
of the germ cells increase in number without an
accompanying division of the cell, thus producing
binucleated cells (Fig. 33, C). The history of the
pole cells during embryonic development will be
more fully described in the Coleoptera, since in
the beetles the Keimbahn is much more distinct.
The origin and nature of the Keimbahnplasma
was not determined by Hasper, but it was found to
persist in certain cases even until the larval stage
was reached (Fig. 33, D).

In Calliphora Noack (1901) described a dark
granular disc at the posterior end of the egg (Fig. 34)
which he termed the " Dotterplatte " and which,
like the pole-plasm of Miastor and the Keimbahn-
plasma of Chironomus takes part in the formation of
the primordial germ cells. The eggs of the parasitic
fly, Compsilura concinnata, were also found by the
writer (Hegner, 1914a) to possess a granular pole-
disc, thus adding one more species to the list of
DiPTERA in which such a structure exists.

Coleoptera. The origin of the germ cells in
beetles and their subsequent history are well known
only in certain species of the family Chrysomelid^
of the genera Calligrapha and Leptinoiarsa. The
contributions of Wheeler (1889), Lecaillon (1898),

'BO,:^ OJO;^

ng.c.

Fig. 33. — Chironomus. A. Longitudinal section through the posterior
end of a freshly laid egg. B. Longitudinal section through egg
during division of first four cleavage nuclei ; at posterior end the
primordial germ cell is just being formed. C. One of primordial
germ cells containing two nuclei and remains of " Keimbahnplasma."
D. Germ gland of the larva in which remains of " Keimbahnplasma "
still appear. Khpl = " Keimbahnplasma " ; p.g.c. = primordial germ
cell. {Froyn Hasper, 1911.) (HO)

GERM CELLS IN THE AUTHROPODA 111

Hegner (1908, 1909a, 19096, 1911a, 19116, 1914a),
and Wieman (1910a, 19106) will be referred to in
the following paragraphs.

Wheeler (1889) figured several primordial germ
cells in an egg of Leptinotarsa with a segmented germ
band and suspected
their true nature, but
did not discover them
in earlier stages. Le-
caillon (1898) de-
scribed the pole-cells
in several chrysomelid
beetles, but did not
make out any of the
details concerning
their origin, structure,
and migrations.

Within the last
seven years the writer
has devoted a consid-
erable portion of his
time to morphological
and experimental
studies of the eggs of
beetles, particularly
Calligrapha higshjana, C. multipunctata, C. lunata,
and Leptinotarsa decemlineata. The eggs of these
species are peculiarly favorable for study, since they
are definitely oriented in the body of the mother
and various surfaces can be recognized in the newly
laid egg : they can be placed under the most severe

Fig. 3i. — Calliphora. A. Longitudi-
nal section through posterior end of
freshly laid egg, showing " Dotter-
platte (Dpi). B. Longitudinal sec-
tion through posterior end of egg at
time of blastoderm formation, showing
protrusion of primordial germ cells
(p.ff.c). (From Noack, 1901.)

112 GERM-CELL CYCLE IN ANIMALS

experimental conditions without killing them or
stopping their progressive development ; and they
can be killed, fixed, sectioned, and stained with
comparative ease. Furthermore, the eggs of these
beetles possess a well-defined pole-disc, and the
primordial germ cells which arise even before the
blastoderm is formed are easily distinguishable
from the somatic cells and thus can be traced from
the time of their appearance until they become ma-
ture eggs and spermatozoa.

The ova of insects have long been considered
among the most highly organized of all animal
eggs. That they are definitely oriented while still
within the ovary was expressed by Hallez (1886) in
his "Law of the Orientation of Insect Embryos"
as follows : "The cell possesses the same orientation
as the maternal organism that produces it ; it has a
cephalic pole and a caudal pole, a right side and a
left side, a dorsal surface and a ventral surface;
and these different surfaces of the egg-cell coincide
to the corresponding surfaces of the embryo." The
orientation of an ovarian egg is indicated in Fig. 35,
and here also is shown the position and surfaces of
the egg at the time of deposition. When the egg is
laid the beetle clings to the under surface of a leaf,
and with a drop of viscid substance from the acces-
sory glands of the reproductive organs, fastens the
egg by its posterior end (p) to the leaf ; then with the
tip of the abdomen the egg is pushed back through
the arc indicated by the dotted line. It is a simple
matter to determine the various surfaces of eggs

GERM CELLS IN THE ARTHROPOD A 113

laid in this manner. Gravity apparently has no
influence upon the development, since eggs in a state
of nature occupy all positions with respect to
this factor without becoming altered in any way.
Only one case has come to the writer's attention
of an influence of gravity in insect development —
the eggs of the water beetle, Hydrophiliu- atterimus.

, p r d X a

Fig. 35. — A diagramatic drawing of Calligrapha bigsbyana clinging to
the under side of a willow leaf and showing the orientation of the
egg in the ovarian tubule and after deposition, a = anterior ; d =
dorsal ; p = posterior ; r — right side ; x = place where egg was
marked with India ink as means of orientation after removal from
leaf.

according to Megusar (1906), develop abnormally if
the cocoon in which they are laid is inverted.

The events that precede the establishment of the
primordial germ cells in chrysomelid beetles may be
described briefly as follows : The egg, when laid
(Fig. 36, A), consists of a large central mass of yolk
globules (y), among which are very fine strands of
cytoplasm ; a thin peripheral layer of cytoplasm, the
''keimhautblastem" of Weismann (JxJihl), a delicate
vitelline membrane (v.m.), a chitinous shell, the
chorion, and a nucleus consisting of the egg nucleus

•ga

...khbl

-:.■.... .y.

-••v.m.

-...fH

'•:.'^-''.."-'v.-- •■••.'• •■'.•■■•■'.•■.■ ^..''v'"-"V.-*Vv:!.?v.'V"v).";;;>' ■:..";•••.■'•. •..•.■-■ •;.l : '•"..• V]-* •;•;.■".•.•■ ': '•. .■*■::
••:.•::..• •.■:■..".■..• .•.■;•;■■■;..■ . ■* . ■.-■.•.. >.•:..•■• .•"■•'•■•vvv/.;|"::; .• : . ; •.•.•..•■■.•..•"... •■• •■ •.■••.'•;".■.••-"••=

Fig. 3,.-Calli,rapha. A. Long^udinal section ^^^-^^ ^J^^^^^^
biasbyana four hours after deposition. B. Longitudinal sec
through an egg of C. biosbyana 14 hours after deposition C.Jjo
germ cells just protruding from posterior end of egg of C.rnulH
fuZata D The pole-disc in an egg of C. mulhpunctata g.c.d -
pole dtc- gn. = germ nuclei fusing; khbl = keimhautblastem ; P -
posterior 'end of egg; pM.n. = preblastodermic nuclei; v.m. = vrtel-
Une membrane ; vt. = vitellophags ; y. = yolk. ^

GERM CELLS IN THE ARTIIROPODA 115

and a sperm nucleus combined (g.n). Frequently
the two polar bodies have not yet been produced
when the egg is laid and thus many stages may be
encountered in the newly laid eggs. Polyspermy is
a normal condition in insects and several sperma-
tozoa are often observed among the yolk globules.
The keimhautblastem is not homogeneous through-
out, for at the posterior end there is embedded in it a
disc-shaped mass of darkly staining granules which I
have called the pole-disc (g.c.d.) and which resembles
the pole-plasm of Miastor, the "Keimwulst" or
" Keimbahnplasma " of Chironomus and the " Dotter-
platte" of Calliphora,

The cleavage nucleus divides by mitosis; the
daughter nuclei separate slightly, and divide; and
this process is continued until nuclei, each surrounded
by a small mass of cytoplasm, are scattered more or
less regularly throughout the egg. Then a division
of the nuclei into tw^o groups occurs; those of one
group migrate to the periphery, fuse with the periph-
eral layer of cytoplasm, and are cut off by cell walls,
thus forming the blastoderm; whereas the other
nuclei, the vitellophags, remain behind among the
yolk globules which it is their function to dissolve.
The blastoderm consists of a single layer of cells,
except at the posterior end where its formation has
been interrupted by the process resulting in the
establishment of the primordial germ cells.

The primordial germ cells are formed in the fol-
lowing manner. The cleavage nuclei at the posterior
end of the egg that encounter the pole-disc granules

116 GERM-CELL CYCLE IN ANIMALS

behave differently from those at other points, since
they do not remain to form part of the blastoderm
but continue to migrate until they have become
entirely separated from the rest of the egg. During
this process each of the sixteen nuclei that act in this
way becomes surrounded by a halo of granules —
part of the pole-disc. Then cell walls appear and
sixteen primordial germ cells result. These form a
group at the posterior end, each member of which
divides twice, thus producing sixty-four germ cells
in all. During these divisions, which are mitotic,
the pole-disc granules appear to be equally distrib-
uted between the daughter cells (Fig. 37, B).
A rest period then occurs, as far as cellular multipli-
cation is concerned, during which a ventral plate,
which later grows into the germ band, develops on
the ventral surface of the egg. As in Miastor the
germ-band pushes around on the dorsal surface
and the group of sixty-four germ cells is carried
along with it. In the meantime the germ cells
begin to migrate from the amniotic cavity in which
they lie through a sort of canal at the bottom of a
groove in the germ-band and thus make their way
inside of the embryo (Fig. 37, F). That the germ
cells actually migrate and are not simply forced
about by the surrounding tissues seems certain since
they are ameboid in shape and pseudopodia extend
out in the direction of their movement (Fig. 37, F) .

After penetrating into the embryo the germ cells
become separated into two groups. It was difficult
to count the number in each group, but many

GERM CELLS IN THE ARTHROPOD A 117

•Pdg.

V.2.-.-\

-.'■•

#^--khbI

- 0

-0:

^.~^^...

SP^

- — go.d.

•---n

PgC

&a.

•R

Fig. 37. — Calligrapha. A. A germ cell of C. maltijnmctala shortly after
being cut off from the egg. B. Division of a primordial germ cell.
C. Longitudinal section through egg of C. biysbyana at blastoderm
stage ; the posterior end was killed with a hot needle just after
deposition. D. Longitudinal section through uninjured egg at
same stage. E. Two ectoderm cells (e), two mesoderm cells (m),
and two germ cells (g.c.) from an egg three days old. F. Germ
cell during migration into the embryo (three days old). G.II.I. Longi-
■ tudinal sections through eggs centrifuged for one hour, two hours,
and four hours respectively, bl = blastoderm ; g.c.d. = granules of
pole-disc ; A: = killed portion of egg ; khhl. = keinihaut hi astern ; p. =
posterior; 7>f;r = primordial germ cells; v — vitcllophags ; v.z. =
vesicular zone ; j. ~ yolk.

118 GERM-CELL CYCLE IN ANIMALS

attempts seem to justify the conclusion that the
division is equal or approximately equal, that is,
each group contains about thirty-two germ cells.
These groups acquire a covering of mesoderm cells,
are carried by the somatic tissues to a position
near the dorsal surface on either side of the body in
the last two abdominal segments, and thus become
germ glands situated in their definite positions.
Some time before the larval stage is reached, the
sex of the embryo can be determined by the shape
of the germ glands ; those of the male become dumb-
bell shape, whereas the female organs retain the earlier
pear shape and begin to acquire terminal filaments.
It is interesting to note that much time and
effort have been wasted by those who have attempted
to influence the sex of caterpillars by over-feeding
or starving. Kellogg (1907), for example, "dis-
covered," after an unsuccessful attempt to change the
sex of silk worms by this means, that these cater-
pillars already possess germ glands which are dif-
ferentiated as male or female. If he, and others
who have undertaken similar experiments, had
examined the literature on the origin of the germ
cells in insects, they would have found that as long
ago as 1815, Herold published results of investiga-
tions on Papilio brassica and other species of Lepi-
DOPTERA which proved that the sex of the larva is
already determined before it hatches from the egg.
A similar condition was reported in Bomhyx pini by
Suckow (1828), in Zeuzera oesculi hy Vessels (1867),
and in Pieris brassica by Brandt (1878).

GERM CELLS IN THE ARTHROPODzV 119

There now ensues a period of activity during
which a large number of ovarian tubules develop
in the female and testicular follicles appear in the
male. A number of much debated problems exist
regarding the cellular elements within the ovaries
and testes of insects — problems which are of con-
siderable importance in any discussion of the germ-
cell cycle. Put in the form of questions, two of these
are with respect to the ovary: (1) Do the nurse
cells originate from the oogonia, thus becoming
abortive eggs, or are they of mesodermal parentage ?
(2) Does amitotic nuclear division occur in nurse
cells and oogonia ?

The answers to these questions differ according to
the species of insects studied, and, as usual, the ob-
servations and interpretations of different investi-
gators do not always agree. They can be answered
with certainty in the case of Miastor. All of the
oogonia in this form are direct descendants of the
primordial germ cell ; the nurse cells are of meso-
dermal origin; and amitotic division occurs neither
in the nurse cells nor in the oogonia. The situation
is quite different in chrysomelid beetles. The nurse
cells in the ovaries of the potato beetle all seem to be
of germ-cell origin. That the nurse cells which are
derived from oogonia are abortive eggs is the general
opinion of zoologists. Convincing evidence for this
view has recently been provided by De Winter
(1913) from studies of the apterous insect, Podura
aquatica. In this species the proportion of eggs and
nurse cells which develop from the oocytes is about

120 GERM-CELL CYCLE IN ANIMALS

one to ten. The oocytes that become eggs are those
that chance to He at the periphery of the ovary and
hence are in a position to derive abundant nutrition
from the blood. The oocytes that fail to become
eggs are not, according to De Winter, " vitello-
genes" but true abortive eggs, representing a more
primitive stage than the nurse cells of other insects
which have acquired, secondarily, a nutritive func-
tion.

On the other hand, Govaerts (1913) argues strongly
in favor of the view that the oogonia divide differen-
tially, the daughter cells becoming true germ cells
(the ultimate oogonia) and true somatic cells (the
nurse cells). He bases his position upon the condi-
tions existing in the ovaries of certain beetles of the
genera Carabus and Cicitidela, and upon the dis-
coveries of Giardina (1901), Debaisieux (1909),
and Gunthert (1910) in Dytiscus marginalis. Giar-
dina established for Dytiscus the fact that the mito-
ses which result in the formation of nurse cells are
differential, as theoretically postulated by Paulcke
(1900). During the four divisions preceding the
formation of the oocyte a single oogonium gives rise
to one oocyte and fifteen nurse cells (Fig. 38). A
differentiation takes place in the chromatin of the
oogonial nucleus, one half consisting of a condensed
mass, the other half of large granules which corre-
spond to the forty chromosomes of the oogonium
(Fig. 38, A). During mitosis the chromosomes
become arranged as an equatorial plate, and the
chromatic mass forms a ring about it — the "anello

GERM CELLS LN THE ARTHROPODA 121

cromatico" (B). This ring passes intact to one of
the daughter cells (C), whereas the chromosomes are

Fig. 38. — Differentiation of nurse cells and oocytes in Dytiscus inar-
ginalis. A. Oogonium with chromatin of nucleus separating into
two parts. B. Metaphase of oogonial mitosis; the " anello croma-
tico" is situated at the lower end of spindle. C. Two-cell stage;
the lower cell with nucleus containing two sorts of chromatin.
D. Four-cell stage; "anello cromatico" in one cell. E. Eight-cell
stage ; cells ready to divide. F. Sixteen-cell stage ; one large cell
(oocyte) with chromatin from the "anello cromatico," and fifteen
- nurse cells. {A-D, F, from Giardina, 1901; E, from Dchaiseaux, 1909.)

equally divided. During the succeeding mitoses
similar differential divisions occur result in"' in one
oocyte containing the chromatic ring (Fig. 38, F) and

122 GERM-CELL CYCLE IN ANIMALS

fifteen nurse cells lacking this nuclear substance.
Thus as Paulcke's theory demands, the difference
between the nurse cells and the oocytes is the result
of internal and not external causes.

Giardina considered the formation of the chromatic
ring as a sort of synapsis, and later (1902) distin-
guished between a complete synapsis, such as
ordinarily occurs in the germ-cell cycle, and a partial
synapsis as exhibited by Dytiscus. Regarding the
significance of this differential mitosis, he maintains
that this phenomenon is the cause of the differen-
tiation into nurse cells and oocytes, resulting in a
complete amount of chromatin in the keimbahn
cells and perhaps also an unequal distribution of cyto-
plasmic substances. As in the case of Ascaris and
Miastor, it might better be regarded as a means of
depriving the nurse cells of part of their chromatin.
Moreover, Boveri (1904) has compared the chroma-
tin-diminution in Ascaris with Giardina's differ-
ential mitoses. Debaisieux (1909) and Gunthert
(1910) have confirmed Giardina's results, and the
latter studied two other Dytiscid^, Acilius and
Colymhetes, which also exhibit differential mitoses
similar except in certain details. Gunthert found
that the chromatic ring is composed of fine granules
which may split off from the surface of the chromo-
somes (compare with Ascaris and Miastor) and stain
like cytoplasm. He interprets this as " Zerfallspro-
dukte" of the chromosomes. Debaisieux, on the
other hand, claims that this cast-out nuclear material
is nucleolar rather than chromatic in nature.

GERM CELLS IN THE ARTHROPODA 123

It seems highly probable that the "anello croma-
tico" of Giardina consists of chromatin, and Gold-
schmidt (1904) and others do not hesitate to class
it as an example of a " Chromidialapparat." Further-
more it is apparently the result of a chromatin-
diminution, as Boveri (1904) maintains, differing
from the similar process in Ascaris and Miastor in
details but not in the ultimate result. Finally, the
discovery of this peculiar body in Dytiscus adds one
more argument to the hypothesis that the chromatin
content of the germ cells differs from that of the
somatic cells quantitatively, at least in some cases,
and perhaps also qualitatively.

Many are the bodies that have been homologized
with the " anello cromatico" of Dytiscus. Buchner
(1909) claims that the nucleolar-like structure in
the oogonia and young oocytes of Gryllus is homol-
ogous to both accessory chromosomes of the sper-
matogenesis and to this chromatin ring in Dytiscus.
This " accessorische Korper" passes intact into one
half of the oocytes where it disintegrates into granules
of a "tropische Natur." Foot and Strobell (1911)
have also compared it with the chromatin nucleolus
in the oogonia of Protenor with which it has certain
characteristics in common, but no such differential
divisions occur as in Dytiscus.

Govaerts (1913) was unable to find anything
resembling the chromatic ring of Giardina, and con-
cludes that the formation of a chromatic mass dif-
ferentiating the oocytes and the nurse cells is unique
in the Dytiscid^. His investigations demonstrate

124 GERM-CELL CYCLE IN ANIMALS

that this phenomenon does not occur in all insects and
that we must seek some larger cause than the un-
equal distribution of chromatic elements.

If no differential divisions are present, as in
Dytiscus, what is the cause of the formation of
oocytes and nurse cells ? Govaerts decides that since
the ultimate oogonium possesses a definite polarity
marked by the localization of the "residu fusorial,"
and the two kinds of daughter cells arise from op-
posite ends of the mother cell, the cause of the differ-
entiation resides in the polarization of the oogonium.
He does not, however, account for this " polarite pre-
differentielle."

Haecker (1912) has described in Cyclops and
Diaptomus a three-cell stage in the development
of the gonad which is brought about by the delayed
division of one of the germ cells of the two-cell
stage, and concludes that as in Dytiscus there must
be an internal difference in the cells to account
for this condition.

Wieman (19106) has followed the history of the
oogonia in Leptinotarsa signaticollis through the
larval and adult stages, but was unable to find any
evidence that the nuclei inaugurate differentiation
as in Dytiscus. He concludes that "the process
seems to be the result of several distinct cell elements
which operate together as a whole" (p. 148) and that
the semi-fluid matrix which results from the lique-
faction of cells at the base of the terminal chamber
may exert a " specific effect on those germ cells
coming under its influence, enabling them to develop

GERM CELLS IN THE ARTHROPOD A 125

into ova, while the more distant germ cells become
nurse cells" (p. 147). My observations agree
with those of Wieman ; no definite relations nor
nuclear evidence were discovered during the differ-
entiation of the oogonia into oocytes and nurse cells.
The data available do not suggest any method of
differentiation not already proposed, and still leave
the question whether the nurse cells should be
regarded as abortive germ cells or true somatic
cells one of personal opinion.

A study of cyst formation in the testis of the potato
beetle has revealed what seems to be a series of events
in the male germ-cell cycle parallel to that in the
females of Dytiscus, Carabus, and Cicindela, during
which the nurse cells are produced. There are in
Leptinotarsa two pairs of testes, one on either side of
the body. Each testis consists of a large number of
follicles radiating out from near the center. Figure
39 is a diagram of a longitudinal section made
from the testis of an old larva. At the lower end
is attached the sperm duct (s.d) which is con-
nected with a cavity (c) within the testis. Just
above this cavity is a region containing degenerating
cells ; above this region is a mass of spermatogonia
(sg) not yet within cysts ; and this mass is capped
by a small group of epithelial cells (t.c). The major
part of the testis is composed of radiating follicles
containing cysts of spermatogonia, spermatocytes,
or spermatozoa (cy) .

In that region of the testis surrounding and under-

126 GERM-CELL CYCLE IN ANIMALS

lying the terminal cap (Fig. 39, t. c) there are a large
number of spermatogonia not yet contained in cysts.
All stages in cyst formation may be observed here
not only in larval testes but also in those of pupse
and adults. The youngest spermatogonia are those
lying near the terminal cap. Figure 40, A shows a

few cells of the
terminal cap {t.c),
some of the neigh-
boring spermato-
gonia {sjpg) , and
several of the epi-
thelial cells {ep)
that are scattered
about among the
spermatogonia.
V vsts fj re Tormed

Fig. 39. — Leptinotarsa decemlincata. Longi- "^

tudinal section through testis of full-grown toward the edge
larva. c = cavity; c?/ = region of cysts; e: x.\^ x.
s.d = sperm duct; s^ = region of spermato- ^'- ^^^ Spermato-
gonia ; sv = region of spermatozoa ; t.c = gonial maSS away
terminal cap. ° . '^

from the terminal
cap, and Fig. 40, A to G represent certain of the
stages observed. The spermatogonia divide ap-
parently exclusively by mitosis. A well-developed
spindle is formed and this persists after the cell wall
has separated the two daughter cells. The spindle
fibers which are at first perfectly distinct (Fig. 40, B)
unite into a compact strand (Fig. 40, C) which
stains dense black in iron hsematoxylin after fixa-
tion in Carnoy's fluid. In many cases it was im-
possible to distinguish an intervening cell wall

Fig 40 _r ,. ^ ^

^- ^^Vi^notarsa decemlineata qfn„ •

in ^amo process n'"R''"'f" »' ^P"n,atoSun,^' r""! T'""'"'"
lial envdopepp, """■''■'•'*'' «P<'™atogo„iarocJl wi,^ '''";'"'

<?• S^^*'» through eysfeomairg'thStr'""'""'^ '''''" '-" •

g Mirty-two spormatogouia. (127)

128 GERM-CELL CYCLE IN ANIMALS

between the daughter nuclei (Fig. 40, D). In
either case, however, the spindle remains persist,
forming a basic staining strand with enlarged ends
connecting the two nuclei. Since at this time and
in all later stages the two or more spermatogonia may
be found surrounded by an envelope of epithelial
cells, it seems certain that, as Wieman (19106)
maintains, the spermatozoa in a single cyst are
derived from a single spermatogonium.

A cyst containing four spermatogonia is repre-
sented in Fig. 40, E. Here again appear the strongly
basic staining spindle remains connecting the nuclei.
These black strands persist until the succeeding
mitotic division occurs as Fig. 40, F, which was
drawn from a section of a cyst containing eight
spermatogonia, shows. Spindle remains are still
evident in later stages, as in Fig. 40, G, which repre-
sents a cyst containing thirty-two spermatogonia, but
were not observed in cysts containing more than
sixty-four cells.

Many investigators have figured spermatogonial
divisions which result in rosette-like groups of cells
similar to that represented in Fig. 40, F. Ap-
parently, however, the spindle remains, if present, did
not possess such a strong affinity for basic stains.
Furthermore, only those of my preparations that
were fixed in Carnoy's fluid and stained in iron
hsemotoxylin exhibited these black strands. Similar
spindle remains have been observed in Dytiscus,
especially by Gunthert (1910), and Carabus (Go-
vaerts, 1913), during the differentiation of nurse

GERM CELLS IN THE ARTHROPODA 129

cells and oocytes from oogonia, and there can be
little doubt but that the process of cyst formation
in the male as described above is similar to the differ-
ential divisions in the female.

Thus the discovery of these distinct spindle re-
mains in the spermatogonial divisions enables us
to homologize one more period in the cycle of the
male germ cells with a corresponding period in the
cycle of the female germ cells.

According to this view the ultimate spermato-
gonium passes through a certain number of divisions
— probably five or six — which correspond to the
differential divisions so clearly exhibited by the
ultimate oogonia of Dytiscus. Just as in the matura-
tion processes, however, where only one female cell
but all of the male cells are functional, so these
earlier divisions result in the female in the pro-
duction of a single oocyte and a number of nurse
cells which may be considered abortive eggs, whereas
in the male every daughter cell is functional. The
limited period of division in the cycle of the male
germ cells in man (Montgomery, 1911 ; von Wini-
warter, 1912) is also similar to those in Dytiscus and
Leptinotarsa. The Sertoli cells are intimately con-
nected with the germ cells in the mammalian testis
and probably perform three functions : (1) they
nourish the spermatocytes ; (2) they provide the
spermatic fluid ; and (3) they exert some chemico-
tactic stimulus which serves to orient the spermato-
zoa into bundles. The origin of the Sertoli cells has
been for many years in doubt. Many investigators

130 GERM-CELL CYCLE IN ANIMALS

claim that they arise from cells other than germ cells ;
these writers have been called by Waldeyer (1906)
" dualists." An equal number of authorities be-
lieve that both Sertoli cells and spermatogonia

Fig. 41. — Stages in the formation of the Sertoli cell in man. A. Sper-
matogonium containing granular inclusion {X) from which " Sertoli
cell determinant " ma^/ arise. B. Antepenultimate spermatogonium
showing rod (R) and idiozome (/). C. Division of rod. D. A Ser-
toli cell containing a divided rod (R) and two rodlets {rz). E. Ser-
toli cell with crystalloid of Charcot and lipoid granules ; at lower
right corner a spermatogonium with crystalloid of Lubarsch. {A—D,
from Montgomery, 1911; E, from von Winiwarter, 1912.)

originate from primordial germ cells ; these are the
" monists."

The researches of Montgomery and von Wini-
warter have decided the question, at least so far as
man is concerned, in favor of the monists. Mont-
gomery's results are diagrammatically shown in
Fig. 42. Of thirty antepenultimate spermatogonia
examined, twenty-three contained each a rod-shaped
structure (Fig. 41, B, R) and it seems probable
that this peculiar body, which is identified by von
Winiwarter with the " cristalloide de Lubarsch "
(Lubarsch, 1896), is present in every cell of this

GERM CELLS IN THE ARTHROPODA 131

generation. This rod is considered by Montgomery
to be of cytoplasmic origin and is termed by him a
" Sertoli cell determinant." During the division

Antepenultimate
Spermatogonium

Penultimate
Spermatogonia

Ultimate _ /
Spermatogonia

Sertoli y"
cell \

Germ Cells Proper

Fig. 42. — Diagram illustrating the differentiation of the Sertoli cell in

man. {From Montgomery, 1911.)

of the antepenultimate spermatogonia the rod passes
undivided into one of the daughter cells ; thus one-
half of the penultimate spermatogonia possess a

132 GERM-CELL CYCLE IN ANIMALS

rod, the other half do not. Of the forty-nine penul-
timate spermatogonia examined, twenty-four ex-
hibited a rod and twenty-five did not. This result
has been confirmed by von Winiwarter. When the
rod-containing penultimate spermatogonia divide,
there is a similar segregation of the rod in one of
the daughter cells, hence only one-fourth of the cells
resulting from the divisions of the antepenultimate
spermatogonia possess a rod. Of one hundred
and forty-two cells of this generation studied by
Montgomery, twenty-five were found with a rod and
one hundred and seventeen without. That this
ratio is less than one to three (1:3) is explained by
the fact that some of the spermatogonia with rods
may already have become Sertoli cells. The further
history of the rod in the Sertoli cell is as follows : A
primary rodlet is produced by a splitting of the rod
(Fig. 41, C) after which the rod either disappears
at once or else persists for a time, in which case it
may split longitudinally as shown in Fig. 41, D.
However, in four-fifths of the cells examined (one
hundred in number) the large rod disappeared
before the growi;h of the Sertoli cell had begun.
Each primary rodlet splits longitudinally into two
approximately equal parts, called secondary rodlets
(Fig. 41, Z), r2), which persist until the end of the
cycle of the Sertoli cell.

Neither Montgomery nor von Winiwarter were
able to determine the origin of the rod. They do
not consider it mitochondrial in nature, although
it may arise from granules lying in the cytoplasm.

GERM CELLS IN THE ARTHROPOD A 133

Montgomery found in one cell a mass of granules
from which the rod may have developed (Fig. 41, yl,
X), and von Winiwarter noted that the rod had a
granular appearance in the earliest stages he ex-
amined. It is also perfectly distinct from the io-
zome (see Fig. 41, B, I) and is apparently not
directly derived from the nucleus. Von Winiwarter
is not so certain as Montgomery regarding the history
of the spermatogonia, the" cristalloide de Lubarsche,"
and the " batonnets accessoires," as he calls the
rodlets. He was unable to decide regarding the
number of spermatogonial divisions and believes it
to be indeterminate. He finds, contrary to Mont-
gomery, the rod persisting in fully developed Sertoli
cells, and considers the fragmentation or fission of
the rod to form the primary rodlets as doubtful.
Further investigations with more favorable material
are very desirable, but notwithstanding certain
differences of opinion between the two writers whose
results have been briefly stated above, it seems cer-
tain that Sertoli cells and germ cells are both derived
from primordial germ cells, and that the Sertoli
cells differ from the ultimate spermatogonia in the
possession of a peculiar rod probably of cytoplasmic
origin. Montgomery considers this a sort of secon-
dary somatic differentiation (the Sertoli cells repre-
senting the soma of the testis) ; the first somatic dif-
ferentiation occurring when the tissue cells become
differentiated from the germ cells in the embryo.

Amitosis. Wilson (1900) defines amitosis as
" mass-division of the nuclear substance without

134 GERM-CELL CYCLE IN ANIMALS

the formation of chromosomes and amphiaster"
(p. 437) and concludes from a review of the literature
up to the year 1900 "that in the vast majority of
cases amitosis is a secondary process which does not
fall in the generative series of cell-divisions" (p. 119).
During the past ten years interest in direct nuclear
division has been maintained principally because of
the claims of certain investigators that germ cells
may multiply in this way and still give rise to func-
tional eggs or spermatozoa.

During amitosis the chromatin remains scattered
within the nucleus and does not form a spireme
nor chromosomes, and therefore its individual ele-
ments, the chromatin granules, do not divide. As
a result of this ma^^-division there can be no accurate
segregation of chromatin granules in the daughter
nuclei as is demanded by the theory that the nucleus,
and particularly the chromatin, contains the de-
terminers of hereditary characteristics. Further-
more, nuclear division without the formation of
chromosomes obviously condemns the hypothesis
of the genetic continuity of the chromosomes, and
hence seriously interferes with current ideas regard-
ing the significance of the accessory chromosomes in
the determination of sex. Among the animals in
whose germ cells amitosis has been reported are cer-
tain Amphibia, coelenterates, cestodes, and insects.

Amphibia. Vom Rath (1891, 1893), Meves (1891,
1895), and McGregor (1899) have recorded amitosis
in the germ cells of Amphibia. Meves claims that
the spermatogonia of Salamandra divide amitotically

GERM CELLS IN THE ARTHROPODA 135

in the autumn but return to the mitotic method in
the spring, later giving rise to functional spermato-
gonia. Vom Rath finds amitosis but contends that
the cells that divide in this way do not become sper-
matozoa but are degenerating, being used as nutritive
material by the other spermatogonia. The amitotic
divisions described by McGregor (1899) in Aiii-
phiurna differ in certain respects from those of
both Meves and vom Rath. In this species the
primary spermatogonia divide by amitosis; their
products later divide by mitosis and produce func-
tional spermatozoa. Our knowledge concerning ami-
tosis in the spermatogonia of Amphibia is therefore
in an unsatisfactory state, although the observations
of Meves and McGregor argue strongly in favor of
this method.

CcELENTERATA. While uo direct nuclear divi-
sions were recorded by Hargitt (1906) in the germ
cells of Clava leptostyla the absence of mitotic figures
in the early cleavage stages of the egg led him to the
conclusion that the "nuclear activity differs greatly
from the oridinary forms of mitosis, and appears
to involve direct or amitotic division" (p. 229).
If this were true, the germ cells which are derived
from these cleavage cells must be descended from
cells which once divided amitotically. This case
of supposed amitosis has been cleared up by the sub-
sequent studies of Beckwith (1909), who collected
material of Clava very early in the morning and found
typical mitotic divisions during the maturation and
early cleavage of the egg and no evidence of amitosis.

136 GERM-CELL CYCLE IN ANIMALS

Cestoda. Child concluded (1904) from a study
of the cestode, Moniezia expansa, that this method
of cell division occurs in the antecedents of both
the eggs and the spermatozoa. This writer has
published a series of papers upon this subject using
Moniezia expansa and Moniezia planissima for his
material (1904, 1906, 1907, 1910, 1911), and his
principal conclusion is that in these species the
division of the cells destined to become eggs and
spermatozoa is predominantly amitotic. Mitotic
division also occurs but comparatively rarely. Cells
which have divided amitotically then divide mitoti-
cally during maturation and form typical ova.

The nature of the nuclear division in the cestodes
was later investigated by Richards (1909, 1911) who
studied the female sex organs of the same species
employed by Child as well as material obtained from
Tcenia serrata. Richards finds that mitosis unques-
tionably occurs in the young germ cells but was
unable to demonstrate amitosis. Richards claims
that amitosis cannot be demonstrated except by the
observation of the process in the living material and
the subsequent study of this material by cytolog-
ical methods. Child (1911) agrees with Richards
that amitosis cannot be demonstrated in fixed
material but nevertheless concludes after an examina-
tion of Richards' preparations *'that direct division
plays an important part in the developmental cycle
of Moniezia, in the germ cells as well as in the soma "
(Child, 1911, p. 295).

Finally Harman (1913) was unable to find any

GERM CELLS IN THE ARTHROPOD A 137

evidence of amitotic divisions in the sex cells of either
Tcenia tenioeformis or Moniezia and concludes that
the conditions that suggest amitosis can just as well
or better be explained by mitosis. Experiments
with living cells of Tcenia were without results,
since the cells did not divide when placed in Ringer's
solution, although they continued to live outside the
body of the host for forty-eight hours. Morse
(1911) likewise failed to observe divisions in living
cells of Calliohothrium and Crossohothrium which
were kept in the plasma of the host. That the
observation of amitosis in living cells is possible
seems certain since Holmes (1913) has recorded an
actual increase in the number of epithelial cells
from the embryos and young tadpoles of several
Amphibia that 'were cultivated in lymph, and has
noted various stages of amitotic nuclear division,
although no convincing evidence was obtained that
this was followed by cell division.

Insecta. In the Hemiptera amitosis was de-
scribed by Preusse (1895) in the ovarian cells of
Nepa cinerea and similar conditions were reported
by Gross (1901) in insects of the same order. Gross,
however, claims that the cells which divide amitoti-
cally do not produce ova but are degenerating or
secretory.

Foot and Strobell (1911) described in ovaries of
the bug, Protenor, the amitotic division of certain
cells which later produce ova. There is, however,
considerable difference of opinion among investi-
gators as to the origin of the ova from the various

138 GERM-CELL CYCLE IN ANIMALS

regions of the insect ovary and, since Payne (1912)
has shown that in Gelastocoris the cells that appar-
ently multiply amitotically do not produce ova, it
seems safe to conclude that in Protenor the ova are
not descended from cells that divide amitotically.

Amitotic division of germ cells followed by mitotic
division has been described by Wieman (19106,
1910c) in the ovaries and testes as well as in the nurse

Fig. 43. — Stages in amitosis in spermatogonium of Leptinotarsa signa-

ticollis. {From Wieman, 1910.)

cells of a chrysomelid beetle, Leptinotarsa signati-
collis. Germ cells in both ovary and testis taken
from full-grown larvae were found in stages of divi-
sion recognized by Wieman as amitotic (Fig. 43).
It was difficult to demonstrate actual division of
the cytoplasm, but that such a division really occurs
was inferred because binucleated cells apparently
gave rise to spermatocytes with single nuclei. Rapid
cell division is assumed by Wieman to account for
amitosis. This is brought about by fluctuations in
the nutritive supply or, in the case of the testis, by
the rapid proliferation of cells during the formation of
cysts.

GERM CELLS IN THE ARTHROPODA 139

I have studied my preparations of chrysomelid
beetles carefully with the aim of detecting amitotic
division and have observed what appears to be direct
nuclear division among the nurse cells, but could not
demonstrate with certainty this kind of division
among the oogonia, or spermatogonia. Three stages
in the direct division of nurse cell nuclei in Leptiiiotarsa
decemlineata are shown in Fig. 8, a-c. Oogonia
and spermatogonia, however, do not exhibit such
clearly defined stages and after examining my prep-
arations and several slides kindly sent me by Doctor
Wieman I am forced to conclude that amitosis has
not been demonstrated. It is true that frequently
dumb-bell shaped nucleoli occur in certain of the
nuclei and frequently two nucleoli are present at
opposite ends. Also two nuclei may be surrounded
by a single cell wall, but no stages were present which
could not be attributed as well or better to mitotic
phenomena.

Conclusion. From the evidence at present
available we must conclude that amitotic division
of the germ cells has not been demonstrated, and
that not until such a process is actually observed
in living cells will any other conclusion be possible.

There are still two questions regarding the germ-
cell cycle in beetles that we shall attempt to answer ;
(1) Does a chromatin-diminution process occur
such as has been described in Miastor and Ascaris?
and (2) Is the segregation of the germ cells controlled
by the nuclei or by the cytoplasm ?

The fact that part of each chromosome is cast out

140 GERM-CELL CYCLE IN ANIMALS

into the cytoplasm in all except the "stem-cell"
during the early cleavage of Ascaris is well known
(see p. 174, Fig. 51). A similar process was described
by Kahle (1908) in Miastor metraloas and confirmed
by me (Hegner, 1912, 1914a) in Miastor americana
(see p. 57, Fig. 16). This chromatin-diminution
process results in the formation of a single primordial
germ cell containing the complete amount of chroma-
tin and a number of somatic cells with a reduced
amount of chromatin. The origin of the germ cells
has been carefully studied in a number of forms which
in other respects resemble Ascaris and Miastor, but
in none of them has such a process been discovered.
Hasper (1911) was unable to establish it for Chirono-
mus which is very similar to Miastor in early develop-
ment, nor has such a phenomenon been found in Sagitta
(Elpatiewsky, 1909, 1910 ; Stevens, 19106; Buchner,
1910a, 19106) and the copepods (Haecker, 1897;
Amma, 1911) and Cladocera (Kuhn, 1911, 1913)
which undergo total cleavage and are in certain
other respects similar to Ascaris.

The nuclear divisions in the eggs of chrysomelid
beetles have been examined by the writer with con-
siderable care, but nothing resembling a diminution
process was found. Furthermore, there are no
evidences of chromatin bodies in the cytoplasm or
yolk as in Ascaris (Fig. 51) and Miastor (Fig. 18, cR),
where the cast-out chromatin does not disintegrate
immediately, but can be distinguished for a consider-
able period during early embryonic development.
It seems necessarv to conclude therefore that in

GERM CELLS IN THE ARTHROPODA 141

chrysomelid eggs both germ cells and somatic cells
possess the full amount of chromatin or else the
elimination of this substance takes place in some
other way.

The Differentiation of the Nuclei of the
Blastoderm cells, Primordial Germ Cells, and
ViTELLOPHAGS. The couclusiou that no chromatin-
diminution process occurs during the early cleav-
age divisions in the eggs of chrysomelid beetles
necessitates the search for some other method of
differentiation among the cleavage nuclei. The
insect egg is particularly advantageous for testing
Roux's hypothesis of qualitative nuclear division,
since we. have here the production of an enormous
number of nuclei before any cell walls are formed,
and an egg that is remarkably definitely organized,
as indicated by my experiments (Hegner, 19096,
1911a), before the blastoderm is formed.

I have been unable to find any differences in the
nuclei before they fuse with the keimhautblastem,
but as soon as this does occur, a gradual change takes
place, and at the time when the blastoderm is com-
pleted three sorts of nuclei are distinguishable:
(1) The nuclei of the primordial germ cells (Fig. 36,
C) are larger than the others and contain compara-
tively few spherical chromatin granules evenly dis-
tributed. The cytoplasm of these cells is distin-
guishable from that of all other cells because of the
presence of granules from pole-disc. (2) The nuclei
of the blastoderm cells are small and completely
filled with large spherical chromatin granules.

142 GERM-CELL CYCLE IN ANIMALS

(3) The nuclei of the vitellophags resemble the
early cleavage nuclei ; they are midway between
the other two kinds in size, and their chromatin is in
a more diffuse condition.

Whether these three kinds of nuclei were all
potentially alike before their differentiation is an
important question. Visibly they are all similar
until they become localized in definite regions of
the egg, and associated with particular cytoplasmic
elements. One cannot help but conclude that they
were all potentially alike and that their differentia-
tion was brought about through the influence of
the cytoplasm in which they happened to become
embedded. The writer has shown (Hegner, 1911a)
that if the posterior end of a freshly laid egg of
Leptinotarsa decemlineata is killed with a hot needle,
thus preventing the pole-disc granules and surround-
ing cytoplasm from taking part in development, no
primordial germ cells will be produced. A large
series of similar experiments have also proved that
at the time of deposition, " The areas of the peripheral
layer of cytoplasm (Fig. 36 khbl.) are already set
aside for the production of particular parts of the
embryo, and if the areas are killed, the parts of the
embryo to which they were destined to give rise
will not appear. Likewise, areas of the blastoderm
are destined to produce certain particular parts of
the embryo" (Hegner, 1911a, p. 251). What
becomes of the nuclei that are prevented from enter-
ing the injured region of the egg ? No evidence
has been discovered to indicate that they disinte-

GERM CELLS IN THE ARTHROPODA 143

grate, so they probably take part in development
after becoming associated with some other part of
the egg. If these nuclei were qualitatively different
they should produce germ cells and other varieties of
cells in whatever region they chance to reach. It
is evident that they are not potentially different
and that their "prospective potency" and "pro-
spective significance" do not coincide. The cyto-
plasm is, therefore, the controlHng factor at this
stage in the germ-cell cycle, although cytoplasmic
differentiations are for the most part invisible and
probably the result of nuclear activity during earlier

stages.

Hymenoptera. a number of papers have ap-
peared which contain references to the germ glands
of Hymenoptera (Hegner, 1909, pp. 245-248).
The most important of these from the standpoint of
the present discussion are: (1) Silvestri (1906, 1908)
and Hegner (19145) on some parasitic species, and
(2) Petrunkewitsch (1901, 1903), Nachtsheim (1913),
and others on the honey-bee.

In an endeavor to test the " Dzierzon theory,"
that the eggs which produce drone bees are normally
unfertilized, Petrunkewitsch (1901-1903) discovered
some usual maturation divisions. In " drone eggs"
the first polar body passes through an equatorial
division, each of its daughter nuclei containing one-
half of the somatic number of chromosomes. The
inner one of these daughter nuclei fuses willi the
second polar body, which also contains one-half of
the somatic number of chromosomes ; the resultant

144 GERM-CELL CYCLE IN ANIMALS

nucleus with sixteen chromosomes, the " Richtungs-
kopulationskern " passes through three divisions,
giving rise to eight " doppelkernige Zellen." After
the blastoderm is completed, the products of these
eight cells lie in the middle line near the dorsal surface
of the egg, where the formation of the amnion begins ;
the nuclei of these cells are small, and lie embedded
in dark staining cytoplasm. Later they are found
just beneath the dorsal surface near the point of
union of the amnion with the head-fold of the em-
bryonic rudiment. They are next located between
the epithelium of the mid-intestine and the ectoderm ;
from here they migrate into the coelomic cavities,
and finally, at the time of hatching, form a " wellen-
artigen" strand, the germ-gland, extending through
the third, fourth, fifth, and sixth abdominal segments.
The fertilized eggs of the bee were also examined
by Petrunkewitsch, but no *' Richtungskopulations-
kern" was discovered. In these eggs the genital
glands arise from mesoderm cells. Doubt was
immediately cast on these results, although Weismann
(1904, p. 336) vouched for their accuracy. Thus
Wheeler (1904) says, " Even in his first paper there is
no satisfactory evidence to show that the cells re-
garded as derivatives of the polar bodies in the figures
on plate 4 are really such, and not dividing cleavage
cells or possibly vitellophags. . . . When we take
up the second paper we wonder how anybody could
regard the figures there presented as even an adum-
bration of proof that the testes of the drone are de-
veloped from the polar bodies." Dickel (1904)

GERM CELLS IN THE ARTHROPOD A 145

could find no connection between the polar bodies and
the cells Petrunkewitsch claims originate from the
" Richtungskopulationskern," but considers these
'' Dotterzellen." Nachtsheim (1913) agrees with
Dickel, that these are yolk cells and have no relation
to the polar bodies. He also finds these cells in both
fertilized and unfertilized eggs, not as Petrunkewitsch
states only in the latter.

The investigations of Silvestri (1906, 1908) on
parasitic Hymenoptera are of particular interest,
since in both the polyembryonic species and those
whose eggs produce a single individual, the keimbahn-
determinant is considered by him to represent a
plasmosome which escapes from the germinal vesicle.
Silvestri (1906, 1908) has described the embryonic
development of both monembryonic and polyem-
bryonic hymenopterous parasites. Of the former
Encyrtus aphidivorus and Oophthora semhlidis were
studied ; in both species the series of events were
found to be similar. The egg at the time of deposi-
tion is elongated and irregularly oval in shape (Fig.
44, A) ; it contains a germinal vesicle (.1) in the
anterior region and a deeply staining body near the
posterior end which is called by Silvestri the " nu-
cleolo" (N) and is stated to be derived from the
nucleolus of the oocyte nucleus. The eggs may
develop parthenogenetically or after fertilization;
the unfertilized eggs produce males, whereas the fer-
tihzed eggs develop into females. In either case
two polar bodies are produced; these disintegrate
later. The cleavage nucleus produces by a series

146 GERM-CELL CYCLE IN ANIMALS

of divisions a number of nuclei which migrate to
the periphery, as is the rule in insect development.
The " nucleolo" remains during this cleavage period
unchanged near the posterior end (Fig. 44, 5) ; then,
when cell walls appear, it becomes distributed among

several of the cells
thus formed. These
multiply less rapidly
than the other em-
bryonic cells and are
the only cells that
give rise to the germ
cells in the adult.
It is thus obvious
that there is here an
early segregation of
germ cells and that
these germ cells dif-
fer from the somatic
cells in the posses-
sion of part of the
{From disintegrated "nu-
cleolo."

The polyembryonic species described by Silvestri
are Copidosoma {Litomastix) truncatellus and Agenias-
pis (Encyrtus) fuscicollis. The eggs of these species
when laid are vase-shaped (Fig. 45), the posterior
end corresponding to the base of the vase. Here
also a germinal vesicle and "nucleolo" are present,
the latter almost always near the posterior end.
Parthenogenetic eggs were found to produce males.

A B O

Fig. 44. — Oophthora. A. Egg with germ-
inal vesicle (A) and "Nucleolo" (A'').

B. Egg containing many cleavage nuclei.

C. Formation of primordial germ cells
(G) at posterior end of an egg.
Silvestri, 1908.)

Fig

45 —Copidosoma (Litomastix) truncatdlu^. A. Oocyte showing
germinal vesicle {g.v) containing a chromatin-nucleolus (c.n) and a
plasmosome KP)- B. Egg a few minutes after deposition showing
first maturation spindle {m.s) and " Nucleolo" (A). C. Egg ;ibout
one hour after deposition, showing three polar bodies (p.6), the
first cleavage nucleus and the "Nucleolo." D. Egg m two-cell
stage, about one and one-half hours old. p.n = Pohir nucleus.
E. Four-cell stage. F. Egg about four and one-half hours old
showing two polar nuclei dividing, two embryonic cells containing
nucleolar substance, and six embryonic cells (dividing) ^ithout
nucleolar substance. {From Silvestri, 1906.) (i-^'^

148 GERM-CELL CYCLE IN ANIMALS

whereas fertilized eggs give rise to females. First
and second polar bodies are formed and the first
divides, thus making three in all. The events of
early cleavage are the same whether the nucleus
consists of the female pronucleus only or of the
female and male pronuclei fused. Unlike the eggs
of monembryonic species, the cleavage nuclei here be-
come separated from one another by cell walls and
the " nucleolo" from the very beginning is segregated
at each division in a single cleavage cell (Fig. 45, D).
This cell divides more slowlv than the others; the
"nucleolo" gradually becomes vacuolated, breaks
down, and finally is evenly scattered throughout
the entire cytoplasm. Just before the sixteen-cell
stage is reached the cell containing the disintegrated
"nucleolo" divides and the two daughter cells are
provided with equal amounts of its substance (Fig.
45, F). Silvestri was only able to trace the cells
containing the remains of the "nucleolo" until
four of these were present. Nevertheless, he con-
cludes that these and these alone give rise to the
germ cells. This conclusion seems well founded when
the history of this "nucleolo" is compared with
that of similar bodies (keimbahn-determinants)
in the eggs of certain other animals.

Two regions develop in the eggs of these polyem-
bryonic Hymenoptera : (1) an anterior or polar re-
gion containing the polar bodies, and (2) the posterior
embryonic region. The latter again becomes differen-
tiated into two regions : (1) an anterior "massa germi-
nigera, " which gives rise to normal larvae, and (2) a

GERM CELLS IN THE ARTHROPOD A 149

posterior " massa monem])rioiiale, " which produces
the so-called asexual larvtie. These lack reproductive,
respiratory, circulatory, and excretory systems. They
are supposed to develop from cell masses which do
not contain descendants of the cell with " nucleolar"
material, and to serve the purpose of tearing apart
the organs of the host, thus making it available as
food for the normal larvae. The " massa monem-
brionale," according to this view, consists entirely of
somatic cells, whereas the " massa germinigera"
possesses both somatic and germ cells. Doubts
have been expressed regarding the development of
the asexual larvae, and Silvestri's results need con-
firmation. There seems to be no doubt that the
"nucleolo" is a keimbahn-determinant in both
monembryonic and polyembryonic Hymenoptera,
but its identification as the nucleolus from the oocyte
nucleus did not seem to the writer to be well estab-
lished. Its history was, therefore, studied by the
writer (Hegner, 19146) during the growth period of
the eggs, with the following results.

My material consisted of a brood of females
belonging to the polyembryonic species Copidosoma
gelechicB. As in most other insects, the two ovaries of
Copidosoma consist of rows of oocytes in various
stages of growth — the oldest and largest near the
posterior end, and the youngest and smallest at the
opposite pole. Before the oogonia enter the growth
period (Fig. 46, A, o) each becomes surrounded by
a follicular epithelium (fe) and is provided with a
group of nurse cells (nc) which likewise are enclosed

f e

Fig. 46. — Copidosoma gelechice. A. Young oocyte (o) surrounded by
an epithelium (f.e) and accompanied by nurse cells (n.c). B. Older
oocyte with nurse string {n.s). C. Oocyte containing spindle on
which are pairs of chromosomes. D-G. Stages in condensation of
this spindle. //. Cross section of spindle in stage shown in C.
I. Cross section of spindle in stage shown in D. J-K. Late stages
in condensation of spindle. (150)

GERM CELLS IN THE ARTHROPOD A 151

by a cellular envelope. Increase in size takes place
synchronously in both the nucleus and the cytoplasm
of the oocyte, and a number of stages in this process
are illustrated in the accompanying figures. In
Fig. 46, B a strand of cytoplasm is shown extending
forward to the nurse chamber, and it is evidently by
means of this pathway that nutritive material is
conveyed to the oocyte. During the growth period
the nurse cells decrease in size until they occupy but
a very small space and the follicular epithelium
becomes very much attenuated (compare Figs. 46, A
and 47, D).

The fully developed oocytes (Fig. 47, D) are more
or less vase-shaped with a broad base (posterior),
a narrower " waist-line," and a slightly thicker distal
(anterior) portion. They are not so long and slender
as those illustrated by Silvestri, but perhaps this
shape is attained later when the eggs are laid.
Within the oocyte are two conspicuous bodies. At
the anterior end is a very large nucleus {n) which
almost completely fills that portion of the egg; it
contains a few scattered rods of chromatin. Near
the posterior end is a smaller but even more con-
spicuous body (Fig. 47, D, k) which stains very deeply
with iron-haematoxylin. This may be vacuolated
and irregular, showing signs of disintegration, as
shown in Fig. 47, or may possess a smooth outline and
be entirely homogeneous. It is undoubtedly of a
very tough nature, since it not infrequently tears out
of the egg substance when struck by the sectioning
knife. This obviously represents the "nucleolo" of

152 GERM-CELL CYCLE IN ANIMALS

Silvestri. Silvestri claims that this " nucleolo" is a
plasmosome which was cast out of the oocyte nucleus
at an early stage in the growth period, but an exami-
nation of my material proves that it really contains
all of the chromatin of the oocyte nucleus. Since it is

Fig. 47. — Copidosoma gelechice. Stages in fusion of two contiguous
oocytes end to end. fe = epithelium ; k = keimbahn-chromatin ;
n = nucleus ; s = spindle breaking down ; u = point of union of
oocytes.

not a nucleolus, at least in the species I have studied,
it can no longer be called a " nucleolo" and therefore
the term ' keimbahn-chromatin ' will be applied to
it.

Figure 46, A was drawn from a longitudinal section
through an oocyte (o) in an early stage of growth. It
is surrounded by follicle cells (fe) and accompanied by

GERM CELLS IN THE ARTHROPOD A 153

a group of nurse cells (n.c) at the anterior end. A
large part of the oocyte is occupied by the nucleus
(?i) within which are a comparatively few irregular
rods of chromatin, forming a group in the center.
This nucleus thus differs quite strikingly from those
of the follicle and nurse cells. In Fig. 46, B is
shown an older oocyte and two of the accompanying
nurse cells (n.c). The nucleus contains many long
slender rods of chromatin which often cross each
other near their extremities.

Soon after this stage is reached the nuclear mem-
brane disappears and a sort of spindle is formed as
illustrated in Fig. 46, C. No asters could be dis-
covered, but the spindle fibers are quite distinct.
The chromatin rods are arranged longitudinally on
the spindle, and in material fixed in Carnoy's solu-
tion and stained in iron-hsematoxylin followed by
eosin, are remarkably distinct. The arrangement
of these rods seems to indicate either that entire
chromosomes are separating after synapsis or that
daughter chromosomes are being pulled apart
after a longitudinal split. I am unfortunately
unable to state definitely what processes do precede
the condition shown here, but it seems probable that
the chromatin of the early oocytes forms a spireme
which breaks up into chromosomes, and that these
chromosomes become united in pairs at or near their
ends, and are there drawn out upon the s]Mndle as
represented in Fig. 46, C. It seems also certain tliat
a definite number of these chromosome-pairs are
present. Only a few cross sections of spindU-s were

154 GERM-CELL CYCLE IN ANIMALS

found in my preparations, but in these the chromo-
somes are widely separated and consequently easily
counted. Apparently there are twelve double rods
in each spindle (Fig. 46, H, I),

Instead of continuing its activity and forming two
daughter nuclei this spindle persists for a long time,
undergoing a gradual contraction and condensation.
Thus in the stage succeeding that just described the
chromatin rods are close together and the entire
spindle has decreased in diameter although not in
length (Fig. 46, D) . Spindles in this condition are not
always parallel to the long axis of the egg but may be
oblique or, more rarely, almost perpendicular to this
axis. Hence several transverse sections were ob-
tained, one of which is illustrated in Fig. 46, 7.
Here also is shown a closer proximity of the chromo-
somes as compared with the cross section of the
younger spindle represented in Fig. 46, H. The
number of chromosomes also appears to be constant,
namely, twelve. During succeeding stages the
spindle continues to shorten and condense. That
shown in Fig. 46, E still exhibits spaces between
the rods and the presence of only a few spindle fibers.
A further contraction is indicated in Fig. 46, F,
where the chromosomes have become so closely
crowded as to form an apparently sohd body in
the shape of a cross. This chromatin body still
continues to contract as shown in Fig. 46, G, J, and K.
At about this time vacuoles begin to appear within it
(Fig. 46, K) and its shape becomes more or less irreg-
ular, most often assuming a nearly spherical condi-

GERM CELLS IN THE ARTHROPOD A 155

tion. This may now be recognized as the " nucleolo "
of Silvestri or the keimbahn-chromatin as we have
decided to call it.

The spindle at first lies nearer the anterior than
the posterior part of the oocyte. As it shortens
and condenses it is more often found below the middle
of the cell, and finally reaches a position near the
posterior end. The conclusion is thus reached that
the *' nucleolo" of Silvestri is not a plasmosome
(metanucleolus) which escapes from the oocyte
nucleus, but consists of all of the chromatin of this
nucleus condensed into a more or less spherical body
during a peculiar process of spindle formation.

The discovery of the origin and nature of the keim-
bahn-chromatin brought forth a new problem,
namely, that of the origin of the egg nucleus. It
was early noted that the oocytes containing this
peculiar spindle were free from any other inclusions
in the cytoplasm. How then do they acquire a
nucleus? Two hypotheses have been considered,
one of which has a considerable body of evidence in
its support. In the first place the nucleus might
arise from chromatin granules which break away from
the chromosomes during the formation or conden-
sation of the spindle. There is, however, no evidence
for this view, since the entire chromatin content of
the oocyte nucleus seems to take part in the forma-
tion of the spindle and later the keimbahn-chromatin.
The second hypothesis was suggested when a number
of cases were discovered of two oocytes lying end
to end without any intervening follicular epithelium.

156 GERM-CELL CYCLE IN ANIMALS

This hypothesis is that pairs of oocytes unite end to
end, the posterior oocyte containing the keimbahn-
chromatin and the anterior furnishing the egg
nucleus. Stages in this process are shown in Fig. 47,
A, B,C, and D.

As the oocytes increase in size and age the f olHcular
epithehum becomes gradually thinner and in several
instances only a delicate strand could be observed
between the ends of adjoining oocytes. In Fig. 47, A
two oocytes are shown without any cellular layer
between them, although the follicular epithelium
extends in a short distance at the point of contact.
The posterior cell is much the larger and older, and
possesses keimbahn-chromatin, but no nucleus.
The other oocyte is younger and smaller and con-
tains what has been interpreted as a disintegrating
spindle (s). The condition illustrated in Fig. 47, B
is similar except that the keimbahn-chromatin in
the posterior oocyte is less regular, having already
begun to break up, and the chromatin rods in the
anterior cell represent a further stage in the trans-
formation of a spindle into a nucleus. Figure 47, C
illustrates what is considered a later stage in the fusion
process. The anterior part, which contains a definite
nucleus, is connected with the posterior position
by a thick strand. The nuclear membrane is not
very distinct in the preparation indicating that the
nucleus is not yet completely formed. The posterior
part is not as large as in the other figures, since the
section was not exactly in the longitudinal axis, but
slightly oblique. The keimbahn-chromatin has been

GERM CELLS IN THE ARTHR0P(3DA lo7

added in the figure from a part of the oocyte three
sections away. A still further stage of fusion is
indicated in Fig. 47, D.

In all these cases and in fully developed eggs
there is a distinct "waist line" which can be ac-
counted for upon the view that two oocytes fuse end
to end as above described, the narrow part corre-
sponding to the region of union. The conclusion
seems warranted, therefore, that every egg when laid
consists of two oocytes which have united end to end,
the posterior or older oocyte being provided with
keimbahn-chromatin derived from the chromatin of
its nucleus, and the anterior supplied with a nucleus
which has arisen from the disintegration of a spindle
similar to that from which the keimbahn-chromatin

originated.

A number of references are present in literature to
what have been termed "uterine," "disappearing,"
or "aborting" spindles. Such a spindle was first
noted by Selenka (1881) in the turbellarian, Tlujsano-
zoori diesingii. Here apparently a completely de-
veloped maturation spindle was observed in the
fully grown eggs after they had entered the uterus;
then, just before the metaphase of mitosis, the spindle
broke down and the nucleus returned to a resting
condition. This same nucleus later gave rise to
polar bodies as in the eggs of other animals. Similar
aborting spindles have been described by Lang (1884)
in several species of polyclads, by Wheeler (1894) in
Planocera inquilina, by Gardiner (1895, 1898) in
Polychoerus caudatus, by Surface (1907) in Planocera,

158 GERM-CELL CYCLE IN ANIMALS

by Patterson (1912) in Graffilla gemellipara, and by
Patterson and Wieman (1912) in Planocera inquilina.
Patterson and Wieman have given the uterine spindle
in Planocera careful study, and have established the
fact that in this species it is simply a maturation
spindle which forms near the center of the egg and
later moves to the periphery, undergoing during
this migration a distinct contraction. They further
suggest that the uterine spindles described in the
eggs of other animals are really one phase in a typical
maturation process.

It has thus been shown that the first maturation
spindle in certain eggs may remain practically in-
active for a considerable period. It should be noted,
however, that in Copidosoma the spindle arises not
in the fully grown egg but in very young oocytes,
and that it appears to lack asters at every period of
its history. While therefore this structure may be a
precocious maturation spindle, it differs markedly
from any other such spindle that I have been able
to find described in cytological literature.

The second view is that the oocyte spindle repre-
sents a special mechanism leading to an accurate
distribution of chromatin in the keimbahn-chromatin
mass. The position of the contracted and condensed
spindle, however, is not definite, since it has been
found to occupy almost any part of the oocyte and
to lie with its long axis parallel to the long axis of
the oocyte, or oblique or even perpendicular to this
axis (Fig. 46, E, G). Furthermore the keimbahn-
chromatin does not seem to be of a definite structure.

GERM CELLS IN THE ARTHROPOD A 159

but soon after it reaches a sphere-like shape it begins
to vacuolate and becomes irregular (Figs. 46, K ; 47).
It also seems probable that in some oocytes the
oocyte spindle gives rise to the keimbahn-chromatin,
whereas in others it becomes disorganized, forming
the nucleus of the egg (Fig. 47, A, B, C). What
causes the difference in the history of the oocyte
spindles ? No definite answer can be given to this
question, but there are two possibilities, (1) external
and (2) internal influences. It seems very improb-
able that any internal mechanism exists which
determines what the history of the oocyte spindle
shall be. On the other hand, the arrangement of
the oocytes in the ovary might cause the spindle of
those most posteriorly situated to become keimbahn-
chromatin and of those next in order to transform
into nuclei. According to this view the oocytes de-
pend upon chance for their final position in the
ovary, and the fate of the spindle is decided by the
environment of the oocyte.

There are numerous cases of cell fusion in both
Protozoa and Metazoa, and germ cells and somatic
cells. For example. Protozoa engulf other cells ;
the fully grown ova of Hydra consist of several
germ cells fused together ; and leucocytes may fuse
with one another. In all such cases the nucleus of
one cell persists, whereas those of the other cells
disintegrate and disappear. Among certain leuco-
cytes of Axolotl, however. Walker (1907) has de-
scribed a sort of fusion which results in the trans-
ference of the chromatin from one cell to another

160 GERM-CELL CYCLE IN ANIMALS

without the disintegration of the migrating chroma-
tin. In plants also Gates (1911) has shown that
chromatin may migrate from one pollen mother-cell
of (Enothera gigcis into a neighboring mother-cell
where it remains visible for some time before be-
coming incorporated with the surrounding cyto-
plasm. Many more cases of cellular fusion might be
mentioned, but in no instance so far as I am aware
has the union of two well-developed oocytes to form
one egg been reported. It is true that in Copido-
soma the chromatin in one (the proximal) oocyte
(the keimbahn-chromatin) finally disintegrates and
disappears in the cytoplasm, and thus the condition
here may be compared with that in the cases men-
tioned above, but the stage of fusion in Copidosoma
is extremely late in the growth period, and the
chromatin material remains visible for a remarkably
long interval of the germ-cell cycle.

According to Silvestri the first cleavage cell of
Copidosoma consists of the egg nucleus surrounded
by only a small portion of the substance in the pos-
terior end of the egg in which is embedded the keim-
bahn-chromatin. If the two materials within the
oocytes do not become intimately fused, it is obvious
therefore that the cells of the embryo which are
descended from the first cleavage cell are derived
from the nucleus of the anterior of the two fused
oocytes and cytoplasm from the posterior oocyte
with the addition of the keimbahn-chromatin.

The history of the germ cells after their segrega-
tion is not known for any polyembryonic animal.

GERM CELLS IN THE ARTHROPOD A 161

Polyembryony has been described in an earthworm.
Lumbricus trapezoides (Kleinenberg, 1879), in cer-
tain Bryozoa (Harmer, 1893; Robertson, 1903),
in the armadillo (Patterson, 1913), and in parasitic
Hymenoptera (Marchal, 1904; Silvestri, 1906,
1908) . In every case cleavage is of the indeterminate
type, and the cell lineage is unknown. Various
theories have been advanced to account for poly-
embryony, such as (1) blastotomy or the early
separation of blastomeres, each giving rise to a
single individual as has been brought about by
Driesch (1892) and others by separating the blas-
tomeres of the eggs of certain animals ; (2) polyovular
follicles may occur in mammals and by some (Rosner,
1901) are considered sufficient to account for poly-
embryony among the members of this class ; and
(3) precocious budding has been suggested to account
for the production of man}^ individuals from a
single egg, most recently by Patterson (1913),
who has shown that in the armadillo the blastoderm
produces two primary buds from each of which two
secondary buds arise, and hence four young develop
from each egg. According to the theory of germinal
continuity each of the buds must be supplied with
germ cells or with germ-plasm which has not yet
been segregated into germ cells. Silvestri \s in-
vestigations seem to indicate that the former is
true for parasitic Hymenoptera, but it is difficult
to see how a definite number of germ cells can be
supplied to each bud during a process of development
which is apparently so indeterminate. If, however, a

16^ GERM-CELL CYCLE IN ANIMALS

definite number is not required, and the germ cells
become generally distributed throughout the cellular
mass before budding begins, the chances are that
every bud will contain one or more germ cells. For
example, if germ cells occur in all parts of the blas-
toderm of the armadillo, as is quite possible, each of
the four embryos must become provided with a por-
tion of them. On the other hand, the germ-plasm
may be rather widely distributed among the cells
and only becomes segregated in germ cells after bud-
ding takes place. Careful studies of the germ-cell
history in polyembryonic species are much needed
and would no doubt produce important results.

The data presented in this chapter are sufficient
to prove that in many insects a complete germ-cell
cycle can be demonstrated. There are many species,
however, in which no early segregation of germ cells
has been discovered even after very careful examina-
tion. It is therefore too early to make any general
statements for the entire class, but we must base our
conclusions regarding the germ-cell cycle upon our
knowledge of those forms in which the keimbahn
actually can be traced. Finally one point should be
emphasized ; in every case the segregation of the
primordial germ cells is intimately associated with a
substance which can be made visible by proper
staining methods. In Miastor this is the pole-
plasm; in Chironomus the "Keimwulst" or **Keim-
bahnplasma" ; in Calliphora the " Dotterplatte " ;
in chrysomelid beetles the pole-disc ; and in parasitic
Hymenoptera, the keimbahn-chromatin. The na-

GERM CELLS IN THE ARTHROPOD A 163

ture and significance of these substances will be dis-
cussed later.

2. The Keimbahn in the Crustacea

The keimbahn in the Crustacea is best known
in certain Cladocera and Copepoda. Of special
interest are the investigations of Grobben (1879),
Weismann and Ischikawa (1889), Haecker (1897),
Amma (1911), Kuhn (1911, 1913), and Fuchs (1913).

Grobben (1879) studied the embryology of Moina
rectirostris and gives a remarkably fine account of
early cleavage stages, considering the early date
when the work was done. He figures stages showing
a foreign body which he considered a polar body,
segregated in one of the early blastomeres, the segre-
gation and characteristics of the primordial germ
cell and the first entoderm cell, and the division and
later history of the germ cells. His results have been,
in the main, confirmed by Ktihn (1911, 1913).

Weismann and Ischikawa (1889) have contributed
an interesting account of the primary cellular differ-
entiation in the fertilized winter eggs of six species
of the Daphnid^, belonging to four genera. The
germinal vesicle in the eggs of these species casts
part of its chromatin contents into the cytoplasm
which there became organized into a "Paranucleus."
This paranucleus then acquired a cell body and in
this condition was termed the "Copulationszelle"
because of its future history. In two of the species
examined this Copulationszelle united with one of
the first two cleavage cells ; in the other four species

164 GERM-CELL CYCLE IN ANLVLVLS

it united with one of the first eight cells. Further-
more, it apparently always fused with a certain
definite cleavage cell. The authors conclude that the
Copulationszelle has some important relation to the
history of the germ cells.

The keimbahn of Cyclops and some closely allied
forms has been very carefully investigated by Haecker

ak-'

Fig. 48. — Cyclops. A. Egg showing " Aussenkornchen " (ak) at one
end of first cleavage spindle. B. Thirty-two-cell stage showing
" Aussenkornchen " (ak) in the primordial germ cell (Kz). Rk =
polar bodies. (From Haecker, 1897.)

(1897), Amma (1911), and Fuchs (1913) with results
which are of particular interest. In Cyclops, accord-
ing to Haecker, "Aussenkornchen" arise at one pole
of the first cleavage spindle (Fig. 48, A, ak) ; these
are derived from disintegrated nucleolar material
and are attracted to one pole of the spindle by a dis-
similar influence of the centrosomes. During the
first four cleavage divisions the granules are segregated
always in one cell (Fig. 48, B, Kz) ; at the end of the
fourth division these "Aussenkornchen" disappear,
but the cell which contained them can be traced by
its delayed mitotic phase and is shown to be the
primordial germ cell.

GERM CELLS IN THE ARTHROPODA 165

The most recent and complete accounts of the
keimbahn in the Copepoda are those of Amma (1911)
and Fuchs (1913). Amma studied the early cleavage
stages of eleven species of Cyclops (Fig. 49, A-G),
three species of Diaptomus (Fig. 49, //), one species
of Canthocamptus, and one species of Ileterocope.
Cyclops fucus var. distinctus is made the basis for
the most detailed study, but short descriptions and
figures are presented of the others. In all of the
sixteen species examined the stem-cell which gives
rise to the primordial germ cell may be recognized,
as Haecker (1897) discovered in Cyclops, first by the
presence of granules which do not occur in the other
cleavage cells, and later by a delayed mitotic divi-
sion. The process is essentially as described by
Haecker.^

^ The following summary of the keimbahn in Cyclops fuscus var. dis'
tindus is given by Amma :

"1. Wahrend der ersten Furchungsteilungen ist eine bestimmte
Folge von Zellen, die Keimbahn, durch das Auftreten von Kornchen,
die sich bei der Teilung jewels um einen Spindelpol der Teilungsfigur
ansammeln gekennzeichnet (Fig. 49, A).

"2. Die Kornchen oder Ectosomen entstehen immer erstmals wahrend
des Stadiums der Diakinese, vermehren sich wahrend der niichst folgen-
den Phasen noch bedeutend und verschmelzen gegen das Ende der
Teilung zu grosseren, unformigen Brocken, welche allmahlich ^^■iih^cnd
des Ruheperiode der Zelle aufgelost werden (Fig. 49, B).

"3. Die neue Kornchenzelle geht stets vom kornchenfiihronden
Produkte der alten Kornchenzelle hervor, was dirckt dadurch hcwiesen
werden kann, dass sich in der neuen Kornchenzelle immer noch unauf-
geloste Uberreste der Ectosomen der alten Kornchenzelle vorHndon ;
alle Komchenzellen stammen somit in direkter Linie von einander ab
(Fig. 49, C).

"4. Vom II — Zellenstadium an bleibt die Kornchenzelle imincr in
der Teilung hinter den andem Furchungszellen zuriick ; es ergibt sich

z u E^

Fig. 49.

Stages in the keimbahn of copepods. A-G. Cyclops fuscus
var. distinctus. H. Diaptomus coeruleiis. I. Cyclops viridis.
A. Ectosomes at end of first cleavage spindle. B. Two-cell stage;
ectosomes dissolving. C. Old and newly formed ectosomes at end
of one of second cleavage spindles. D. Eight-cell stage ; ectosomes
dissolving in stem-cell. E. Sixteen- to twenty-eight-cell stage.
S = cell with, E = cell without, granules. F. One hundred and
twelve-cell stage with two primordial germ cells (u) and three en-
toderm cells (E). G. Two hundred and forty-cell stage, u = pri-
mordial germ cells. H. Appearance of ectosomes before cleavage
spindle forms. /. Increased production of ectosomes due to car-
bonic acid gas. (From Amma, 1911.) (166)

GERM CELLS IN THE ARTHROPOD A 1G7

An important departure from the usual method
of origin of the "Ectosomen" is recorded for Diapto-
mus cceruleus. Amma says concerning the process
in this species that " whereas in other forms the Ecto-
somen first appear during the stage of diakinesis of
the first cleavage spindle, in this species they are
already present before the pronuclei unite" (Fig.
49, H).

The origin and nature of the Ectosomen are con-
sidered by Amma at some length. The hypothesis
that these granules arise by the splitting off of parti-
cles of chromatin from the chromosomes as occurs in
Ascaris is rejected (1) because in one species, Diap-
tomus coeruleus (Fig. 49, H), the Ectosomen appear
before the nuclear membrane breaks down in prepara-
tion for the formation of the first cleavage spindle,
and (2) because the Ectosomen do not stain as deeply
as chromatin but only slightly darker than the cyto-
plasm. The origin of the Aussenkornchen (Ectoso-
men) from the nucleolus, as considered probable by

eine Phasendifferenz, welche in immer starkeren Masse in den hoheren
Furchungsteilungen zunimmt (Fig. 49, C).

"5. Aus dem kornchenfuhrenden Produkte der Kornchenzelle des
vierten Teilungsakts, der Stammzelle S, gehen, nachdem diese sich an
dem fiinften Furchungsschritte nicht beteiligte, gegen Ende des sechsten,
im LX — Zellenstadium, die beiden definitiven Urgeschlechtszellen her-
vor ; bei dieser Teilung der S-Ze\\c erscheinen die Ectosomen in ganzen
Zellraume (Fig. 49, E, F).

"6. In Ausnahmefiillen beginnt die S-Zelle sich etwas friiher zu teilen,
namlich schon wahrend des Ubergangs des XXX — zum LX — Zellen-
stadium.

"7. Die Urgeschlechtszellen verlieren den Verband mit dem Blasto-
derm, sie werden allmahlich in die Tiefe gedriingt (Fig. 49, G)"
(pp. 529-530).

168 GERM-CELL CYCLE IN ANIMALS

Haecker (1897), could not be confirmed. The con-
dition in Diaptomiis coerideus (Fig. 49, H) is also a
serious objection to this theory. The Ectosomen are
different from chromidia, since chromidia arise from
the nucleus and no connection could be discovered
between the Ectosomen and the nuclei. The hy-
pothesis that they may represent chondriosomes is
also rejected.

Amma finally decides ^ that the Ectosomen repre-
sent the ' ' Endprodukte des Kern-Zelle-Stoff wechsels, ' '
in which case a greater amount of Ectosomen would
be present if an egg were allowed to develop in car-
bonic acid gas. The results of a number of experi-

^ " Aus dem ganzen Verlaufe der K6rnchenent^\acklung geht nun so\ael
mit Sicherheit hervor, dass man es bei den Ectosomen mit vergang-
lichen Gebilden zu tun hat, denen keine weiteren Funktionen zukommen,
die im Leben der Zelle nicht weiter verwendet werden. In den Prophasen
der Kernteilung entstehen die Kornchen zunachst als feine Tropfchen im
Zellplasma ; im weiteren Verlauf der Teilung erfahren sie dann noch
eine Zunahme, bis sie ungefahr im Stadium des Dyasters ihre hochste
Entwicklung erreicht haben. Von hier ab beginnt der regressive Prozess
der Kornchen : sie fliessen zu grosseren, unformigen Klumpen zusammen,
welche vom Zellplasma allmahlich voUstandig resorbiert und aufgelost
werden. Bei der nachsten Teilung der Keimbahnzelle erscheinen dann
die Ectosomen wieder von neuem. Um ein einfaches Unsichtborwerden
wahrend der Zellenruhe, wie es. z. B. vom Centrosoma von vielen
Forschern angenommen wird, kann es sich bei den Ectosomen nicht
handeln, denn vielfach konnten ja neben den neuen, frisch entstandenen
Ectosomen noch die Uberreste der Ectosomen der letzen Kornchenzelle
nachgewiesen werden. Es erfolgt also bei jedem neuen Teilungsschritte
tatsachlich eine Neuhildung und Wiederauflosung der Kornchen.

" Gestutzt auf diese Tatsachen, mochte ich nun die Ansicht vertreten,
dass die Ectosomen als Ahscheidiingen, Endprodukte des Kern-Zelle
Stoffwechsels aufzufassen sind, welche zu bestimmten Zeiten im Plasma
der Zelle zur Abscheidung gelangen und wieder aufgelost werden"
(p. 557).

GERM CELLS IN THE ARTHROPODA 169

ments with oxygen and carbonic acid gas indicate
that a greater amount of Ectosomen occur when the
egg is developed in the latter, as shown by Fig. 49, I,
which is from an egg of Cyclops viridis placed one
hour after deposition into carbonic acid gas for one
hour.

When various stains were used it was found that
the Ectosomen became colored much like the
cytoplasm. For example, when stained in methylene
blue followed by eosin the chromosomes were blue
and the Ectosomen and cytoplasm red, and when
stained by the methyl green-f uchsin-orange G method
of Heidenhain the chromosomes were green and
the cytoplasm and Ectosomen red.

Amma also attempts to explain the fact that the
Ectosomen appear at only one end of the first cleav-
age spindle and in only one of the cleavage cells
until the two primordial germ cells are formed.
He rejects the hypothesis Haecker advanced that the
centrosomes possess an unequal influence upon the
Ectosomen and that one centrosome attracts all of
them because it is stronger than the other, and is
inclined to favor the idea that the Ectosomen are the
visible evidence of an organ-forming substance which
is thus distinguished from the rest of the cytoplasm
as *'Kornchenplasma." ^

Fuchs (1913) has confirmed for Cyclops viridis

^ Amma*s statement is, "dass im Zellplasma des noch ungcfurchten
Copepodeneies ein vom iihrigen Eiplasma qualitativ versclnedenes Kbrn-
chenplasma existiert, urlches die organbildende Substanz, die Anlagesub-
stanz fur die Geschlechtsorgane darstellt" (p. 5G4).

170 GERM-CELL CYCLE IN ANIMALS

many of Amma's results and has pointed out the
similarities between the cell lineage of the Copepoda
and Cladocera. Kuhn (1913) has studied the keim-
bahn in the summer egg of a cladoceron, Polyphemus
pediculus, and has confirmed certain parts and cor-

FiG. 50. — Polyphemus pediculus. A. Egg with three nurse cells.
B. Egg at close of maturation, n = " Nahrzellenkern." C. Two-
cell stage ; view of vegetative pole. D. Eight- to sixteen-cell stage.
K ='' keimbahnzelle." E. Sixteen- to thirty-cell stage, e = en-
toderm cell. F. Thirty-two-cell stage from vegetative pole. K =
primordial germ cells ; e = entoderm cells. (From Kiihn, 1911,1913.)

rected other portions of the work done by earlier
investigators — Grobben (1879), Samassa (1893),
and Weismann and Ischikawa (1889). In this
species usually one (but sometimes two or three) of
the nurse cells (Fig. 50) pass into the egg before
cleavage. This cell (or cells) becomes embedded
near the periphery at the vegetative pole (Fig. 50,
B, n). During each of the early cleavage divisions

GERM CELLS IN THE ARTHROPODA 171

this nurse cell is confined to one cell (Fig. 50, C-E)
which gives rise during the third cleavage (8- to IG-
cell stage) to the primordial germ cell, containing
the remains of the nurse cell (Fig. 50, E, K), and to
the primordial entoderm cell which does not receive
any part of the nurse cell (Fig. 50, E, e). The pri-
mordial germ cell and primordial entoderm cell do not
divide as quickly as the other blastomeres during
the succeeding cleavage stages — a fact that aids
in their identification. While the egg is undergoing
cleavage the nurse cell is gradually changing, so that
when the sixteen-cell stage is reached it has become
disintegrated into dark staining granules and frag-
ments of various forms and sizes (Fig. 50, E). Dur-
ing the division of the ''Keimbahnzelle" (from 16-
32-cell stage) these granules and fragments are about
equally distributed between the daughter cells (Fig.
50, F). A similar distribution takes place in suc-
ceeding divisions of the primordial germ cells, and
this is accompanied by a further decrease in the size
of the dark staining granules. A blast ula of "^SG
cells is figured by Kuhn which shows at the vegeta-
tive pole four primordial germ cells lying next to
eight entoderm cells and bordered by twelve meso-
derm cells. During gastrulation this grouj) of twenty-
four cells becomes surrounded by the ectoderm cells,
and the primordial germ cells may then be recognized
as the anlage of the reproductive organs.

Kuhn discusses the origin and significance of the
'*Nahrzellenkern " and comj)ares this body with
similar bodies which have been found in the jjriinor-

172 GERM-CELL CYCLE IN ANIMALS

dial germ cells of other animals, but is unable to ar-
rive at any final conclusion.

In certain Cladocera and Copepoda, as we have
seen, there are visible substances within the cyto-
plasm of the egg which become segregated in, and
render distinguishable, the primordial germ cell . Some
species belonging to these and other groups of Crus-
tacea have been studied in which such a visible sub-
stance pecuKar to the primordial germ cell is absent.

Samassa (1893) not only failed to find the pri-
mordial germ cell during the cleavage stages of
Moina rectirostris, but claims that the germ cells
arise from four mesoderm cells. Kuhn (1908), from
a study of the parthenogenetic generation of Daphnia
j)ulex and Polyphemus pediculus, also derives the germ
cells from the mesoderm. Vollmer (1912) could not
distinguish the germ cells of Daphnia magna and D.
pulex in the developing winter eggs until the blasto-
derm was almost completed and Muller-Cale (1913)
could not find these cells in Cypris incongruens until
the germ layers were fully formed. McClendon
(1906a) has shown that in two parasitic copepods,
Pandarus sinuatus and an unnamed species, the pri-
mordial germ cell is established at the end of the
fifth cleavage (32-cell stage) instead of at the end of
the fourth as Haecker (1897) found in Cyclops. It
is suggested that this delay may be due to the large
amount of yolk present. The stem-cell from which
it arises is, however, not made visibly different from
the rest of the blastoderm by peculiar granules as is
the case in Cyclops.

GERM CELLS IN THE ARTHROPODA 173

Bigelow (1902) has described in Lepa.s cuiafifera
and L. fascicularis certain stages which may Ijring
the forms in which no early segregation of the germ
cells has been discovered into line with the apparently
more determinate species. In Lepas the yolk,
which at first is evenly distributed within the egg,
passes to the vegetative pole and becomes segregated
in one of the first two cleavage cells {cd~). At the
16-cell stage the yolk lies within the single entoblast
cell (d^-^), which occupies a position corresponding to
that of the primordial germ cell in Moina. In this
connection may be mentioned the fact that in many
animals the germ cells are supposed to come from the
entoderm and are characterized by the possession
of much yolk.

CHAPTER VI

THE SEGREGATION OF THE GERM CELLS IN NEM-
ATODES, SAGITTA, AND CERTAIN OTHER MET-
AZOA'

1. The Keimbahn in the Nematoda

The classical example of the keimbahn in animals
is that of Ascaris megalocephala as described by
Boveri (1887, 1892). The first cleavage division of
the egg of Ascaris results in two daughter cells, each
containing two long chromosomes (Fig. 51, A). In
the second division the chromosomes of one cell
divide normally and each daughter cell receives one
half of each (Fig. 51, B, S). The chromosomes of
the other cell behave differently; the thin middle
portion of each breaks up into granules (Fig. 51, A)
which split, half going to each daughter cell, but the
swollen ends (Fig. 51, B, C) are cast off into the cyto-
plasm. In the four-cell stage there are consequently
two cells with the full amount of chromatin and two
with a reduced amount. This inequality in the
amount of chromatin results in different-sized nuclei
(Fig. 51, C) ; those with entire chromosomes (S)
are larger than those that have lost the swollen ends
(C). In the third division one of the two cells w^ith
the two entire chromosomes loses the swollen ends
of each ; the other (Fig. 51, Z), 8) retains its chromo-

174

GERM CELLS IN NEMATODES, SAGITTA 175

somes intact. A similar reduction in the amount of
chromatin takes place in the fourth and fifth divi-
sions and then ceases. The single cell in the 32-cell
stage which contains the full amount of chromatin

Fig. 51. — Ascaris. Stages in early cleavage showing the chromatin-
diminution process in all cells except the stem cell (S). {From
Boveri, 1892.)

has a larger nucleus than the other thirty-one cells
and gives rise to all of the germ cells, whereas the
other cells are for the production of somatic cells
only. The cell lineage of Ascaris is shown in the
accompanying diagram (Fig. 5'i),

176 GERM-CELL CYCLE IN ANIMALS

Meyer (1895) extended the study of chromatin-
diminution to other species of Ascaris. In A. lum-
bricoides no diminution takes place until the four-
cell stage ; then three of the nuclei become deprived
of part of their chromatin. A diminution of this

O O

Q O O O O

o o o o o

Fig. 52. — Ascaris. Diagram showing segregation of primordial germ
cell. E = egg ; Pi, P2, P3 = stem cells ; P4 = primordial germ cell.
Circles represent somatic cells. {From Boveri, 1910.)

sort had been described by Boveri as a variation in
the process observed in A. megalocephala. In A.
rubicunda the differentiation of the cleavage cells
seems to resemble A. megalocephala more than it
does A. lumbricoides. Only late cleavage stages
of A. labiata were obtained by Meyer, but there is

GERM CELLS IN NEMATODES, SAGITTA 177

no doubt that a similar process occurs here. The
general conclusion is reached that the cleavage cells
of all AscARiD^ undergo a chromatin diminution.

Bonne vie (1901), however, while able to confirm
Meyer's results so far as A. lumbricoides is concerned,
could discover no process of diminution in Strongylus
paradoxus and Rhabdonema nigrovenosa.

The elimination of chromatin from all of the
somatic cells of Ascaris and not from the germ cells
led to the conclusion that the germ-plasm must re-
side in the chromatin of the nucleus. The more
recent experimental investigations of Boveri (1910a,
19106), however, indicate that it is not the chromatin
alone that determines the initiation of the diminu-
tion process, but that the cytoplasm plays a very im-
portant role. Dispermic eggs were found to segment
so as to produce three types as follows :

Type I, with one stem cell (P) and three primordial
somatic cells (AB) ;

Type II, with two stem cells and two primordial
somatic cells ; and

Type III with three stem cells and one primordial

somatic cell.

Fig. 53, B shows a cleavage stage of Type II.
Here are represented two stem cells (P) with the com-
plete amount of chromatin, both of which are pro-
paring to divide to form the stem cells (P-z) of the next
generation. From the study of these dispermic
eggs Boveri (1910) concludes ^ that it is ''die unrich-

i"Durch die simultane Vicrteilung eines dispermen Ascaris-Eies
entstehen (vielleicht mit ganz seltenen Ansnahmen) Zellen. welche die
N

178 GERM-CELL CYCLE IN ANIMALS

tigen plasmatischen Qualitdten des sich entwickeln-
den Zellenkomplexes " that cause the injurious re-
sults of dispermy, and that if, of the three types
of dispermic eggs described, the cells could be iso-
lated in pairs, one AB-ceW paired with one Pi-cell,

Fig. 53. — Ascaris. A. Chromatin-diminution in a centrifuged egg.
B. In a dispermic egg. (From Boveri, 1910.)

an embryo, normal except in size, would result
from each pair.

Eggs that were strongly centrifuged cut off at the
beginning of the first cleavage at the heavy pole a

gleiche Wertigkeit besitzen, wie diejenigen, die durch Zweiteilung eines
normal-befruchteten Eies gebildet werden, namlich die Wertigkeit AB
oder Pi. Es konnen drei Zellen die Qualitat AB besitzen oder zwei oder
eine; dem jeweiligen Rest kommt die Qualitat Pi zu. Schon beim
Uebergang vom vierzelligen zum achtzelligen Stadium lasst sich aus der
Teilungsrichtung mit sicherheit diagnostizieren, welche der vier primaren
Blastomeren als AB, welche als Pi aufzufassen sind; und diese Wert-
bestimmung wird durch die weiteren Schicksale der vier Zellfamilien in
jeder Hinsicht bestatigt " (p. 157).

GERM CELLS IN NEMATODES, SAGITTA 179

granular ball (Fig. 53, A, B). This phenomenon was
reported by Hogue (1910) and such eggs were termed
"Balleier." In these eggs the two cells of the four-
cell stage which are adjacent to the "Ball" undergo
the diminution process (Fig. 53, A, AB) ; the re-
maining two are stem cells which give rise to the
germ cells (Fig. 53, A, P). Thus there are two
"Keimbahnen" proceeding side by side in a single
egg and four primordial germ cells are produced in-
stead of two as in normal eggs (see Fig. 51). Miss
Hogue's experiments w^ith centrifugal force led her
to conclude that these must be an "unsichtbare
Polaritat" or " Protoplasmaachse " in the egg of the
Ascaris. Boveri agrees w^ith this and considers
further that the initiation of the diminution process
is not determined by the chromatin but by the
cytoplasm of the egg.^

2. The Keimbahn in Sagitta

Sagitta has proved to be of considerable impor-
tance to those interested in the keimbahn of animals.
Hertwig (1880) figures the four primitive germ cells
in the gastrula and later stages, proving that these
cells are early set aside in embryonic development.
Recently the work of Elpatiewsky (1909, 1910) has

1 He states that, "Was aber aiich hier durch weitere Untersuchungen
noch erreicht werden mag, Eines halto ich fur sicher, dass sich alios, was
uber die Wertigkcit der priiniiron Blastoiiicron bei abnonner Fiirchung
ermittelt worden ist, durch die Annahrae sehr einfacher Plasmadifferenzen
erklaren lasst, wogegen die HyixUhese cinor differenzierenden Wirkung
des Kerns in jeder Form auf imUberwindliche Schwierigkeiten stosst "
(p. 206).

180 GERM-CELL CYCLE IN ANIMALS

given Sagitta a new importance, since this writer has
found within the fertiHzed egg a cytoplasmic inclu-
sion which- is intimately associated with the segre-
gation of the germ cells. The presence of this inclu-
sion has been confirmed by Buchner (1910a, 19106)

'^^^^m^M:^^::^.

C D ^ fr

Fig. 54. — Sagitta. A. First appearance of the " besondere Korper"
(bK) in the egg. B. Egg with germ nuclei fusing. X = " besondere
Korper." C. Thirty-two-cell stage; the primordial germ cell (G)
contains the "besondere Korper" (X). D. Two entoderm cells (E)
and dividing primordial germ cell. E. Two primordial germ cells
showing unequal distribution of "besondere Korper" (X). F. Di-
vision of first two primordial germ cells ; one dividing more rapidly
than the other. (Frorti Elpatiewsky, 1909, 1910.)

and Stevens (19106), and several ideas have been
expressed regarding its origin, fate, and significance.
Elpatiewsky (1909) found in Sagitta, at the time
when the male and female nuclei were lying side by
side in the middle of the egg, a body situated near

GERM CELLS IN NEMATODES, SAGITTA 181

the periphery at the vegetative pole (Fig. 54, B, x).
This body, which he called the *'besondere Korper,"
consists at first of " grobkornigeri " plasma which
stains like chromatin but not so intensely ; later it
condenses into a round homogeneous body with a
sharp contour. During the first five cleavage
divisions the "besondere Korper" is always confined
to a single cell. At the completion of this fifth
cleavage (32-cell stage), the blastomere containing
this cytoplasmic inclusion is recognizable as the first
*'Urgeschlechtszelle" (Fig. 54, C, G) and its larger
sister cell as the first **Urentodermzelle" (Fig. 54,
Cy E). The primordial germ cell is the last to divide
during the sixth cleavage and the "besondere Kor-
per" does not, as before, pass entire into one of the
daughter cells, but breaks up into a number of pieces,
part of which are included in each of the two daughter
cells (Fig. 54, D, X). One of these daughter cells
apparently acquires more of the *'besondere Korper"
than the other. This division appears to Elpatiew-
sky to be differential, separating the primordial
oogonium from the primordial spermatogonium, the
latter being the cell which receives the larger portion
of the "besondere Korper" and which during the
next (seventh) division is sHghtly delayed (Fig. 54,
F). Subsequent to the seventh cleavage the remains
of the "besondere Korper" become pale and grad-
ually disappear, apparently dissolving, and in the
four germ cells resulting from the next division only
occasionally can stained granules from this body be
distinguished.

182 GERM-CELL CYCLE IN ANIMALS

Buchner (1910a, 19106) had no difficulty in find-
ing the "besondere Korper" of Elpatiewsky and in
tracing it during the cleavage stages. He claims that
it originates from the "accessory fertilization cell"
described by Stevens (1904) as degenerating after
the egg breaks away from the oviduct wall, and that
it is chromidial in nature and should therefore be
called *'Keimbahnchromidien." Stevens (1910),
however, has carefully examined abundant material
from Sagitta elegans and S. bipunctata, and no connec-
tion between the "accessory fertilization cell" and
the "besondere Korper" could be traced, the latter
appearing for the first time at the stage when the
egg and sperm nuclei lie side by side in the middle
of the egg, thus confirming Elpatiewsky's conclusions.
She admits the possibiHty of the origin of the "be-
sondere Korper" from granules of the accessory
fertiHzation cell, provided this material loses its stain-
ing capacity for a period, and suggests also that the
granules of chromatin-like material extruded from
the nucleus of the egg during maturation may take
part in its formation. Miss Stevens also beheves
with Elpatiewsky that the "besondere Korper"
divides unequally between the two daughter cells of
the primordial germ cell and that this is a differential
division. She was unable, however, to detect any
constant difference between either the cytoplasm or
the nuclei of oogonia and spermatogonia. It is
worthy of mention that Elpatiewsky (1910) believes
that the "besondere Korper" may originate "aus
dem achromatischen Kernkorper."

GERM CELLS IN NEMATODES, SAGITTA 18S

3. The Keimbahn in Other Metazoa

Certain phenomena have been reported in the
early development of the eggs of many other animals
which have either been compared or can be compared
with conditions such as we have described in the
preceding portions of this book.

The large nucleolus in the germinal vesicle of the
medusa, Mquorea forskalea (Fig. 55, A), according
to Haecker (1892), disappears from the germinal
vesicle about half an hour after the egg is laid, and
a similar body becomes evident near the egg nucleus
which has in the meantime become smaller (Fig. 55,
B). These two bodies are considered by Haecker
to be identical, and the term ''Metanucleolus" has
been applied to them. The metanucleolus is, in each
division up to the sixty-four cell stage, segregated
intact in one cell. Its further history was not
traced, but in the blastula (Fig. 55, D) when the colls
at the posterior pole begin to differentiate, nucleolar-
like bodies appear in some of them which are absent
from the undifferentiated blastula elements. These
may be the descendants of the metanucleolus.

A body similar to the metanucleolus was also dis-
covered by Haecker near the copulating germ nuclei
in the egg of Aurelia aurita, but its history could not
be determined because of the large amount of yolk
present. Haecker identifies the metanucleolus of
Mquorea with the spherical body described hy Metcli-
nikoff (1886) near the egg nucleus of M ifroconia
annoe, and considered by him as a sperm nucleus.

184 GERM-CETJ. CYCLE IN ANIMALS

A similar interpretation is given by Haecker for the
cytoplasmic inclusion ("Spermakern") found by

Boveri (1890) in
Tiara. Similarly
the ''Kleinkern"
which Chun (1891)
discovered in the
egg cells of Ste-
phanophyes su-
perba, and the
bodies described
by Hertwig (1878)
near the matura-
tion spindles of
Mytilus and Sa-
gitta, resemble
very closely the
metanucleolus of
^quorea.

Furthermore,
the metanucleolus
is considered by
Haecker homolo-
gous to the "Par-
acopulationszelle
described by Weis-
mann and Ischi-
kawa in the winter
eggs of certain
Daphnid^, and in both cases it is considered prob-
able that these peculiar bodies are restricted to the
*'Keimbahnzellen" of the embryo.

Fig. 55. — A-D. Stages in formation of
blastula of jEquorea forskalea showing seg-
regation of metanucleolus. {From Haecker,
1892.) E. Oocyte of the cat containing
the "corps enigmatique" (c.e). {From
Vander Stricht, 1911.)

GERM CELLS IN NEMATODES, SAGITTA 185

In the eggs of Myzostoma, Wheeler (1897) found
that the nucleolus of the germinal vesicle does not
dissolve soon after it is cast out into the cytoplasm
during the formation of the first maturation spindle,
but remains visible at least until the eight-cell stage,
at which time it lies in the large posterior macromere,
a cell which "very probably gives rise to the entoderm
of the embryo." Later embryonic stages were not
studied. According to Wheeler "the nucleoli are
relegated to the entoderm cells as the place where
they would be least liable to interfere in the further
course of development and where they may perhaps
be utilized as food material after their disintegra-
tion " (p. 49).

McClendon (19066) has likewise described a body
embedded in the cytoplasm of the egg of Myzostoma
clarki which he derives from the "accessory cells"
which, as Wheeler (1896) has shown, attach them-
selves to either pole of the oocytes. These "acces-
sory" cells are really the "Nahrzellen" of other
authors. The cleavage of the egg was not studied.
Buchner (19106) suggests that this body described
by McClendon and the "nucleolus" of Wheeler are
identical and that through them the keimbahn may
be determined.

Granules of various sorts have been noted in the
eggs of various animals which are segregated in par-
ticular blastomeres and may have some relation to
the keimbahn. For example, among the mollusks,
Blockmann (1881) has described the appearance of
a group of granules in the early cleavage cells of

186 GERM-CELL CYCLE IN ANIMALS

Neritina which finally reach the velar cells. It is
also probable that Fol (1880) observed similar gran-
ules in the 16-cell stage of Planorhis. In the same
category, no doubt, belong the bodies figured by
Fujita (1904) in the 4-cell to the 16-cell stages of
Siphonaria lying at the vegetative pole, and the
"Ectosomen" described and figured by Wierzejski
(1906) in Physa. These granules appear at the vege-
tal pole in the blastomeres of Physa during the
second cleavage ; are at first embedded in the ento-
derm mother cells, but finally become localized in
the ectoderm cells. They periodically appear and
disappear, and may, as suggested by Wierzejski,
represent only "eine besondere Erscheinung des
Stoffwechsels" (p. 536).

Similarly in the rotifer, ^52?Zanc/ma, Jennings (1896)
has traced a "cloud of granules" from the eight-cell
stage until the seventh cleavage, when this mass
forms part of the smaller entodermal cell. In Lejpas
there has also been recorded (Bigelow, 1902) a segre-
gation of granules in one blast omere. Many other
substances granular in form have been described in
the eggs of animals, some of them at least having
migrated there from the somatic tissue. Blockmann
(1887) discovered a number of bacteria-like rods
in the undeveloped eggs of Blatta germanica; these
rods multiplied by division and were considered sym-
biotic bacteria. " Bacterienartige Stabchen" were
also noted by Heymons (1895) in the eggs of Peri-
planata orientalis and Ectobia livida; these sink into
the yolk and disappear. More recently a report of

GERM CELLS IN NEMATODES, SAGITTA 187

Buchner (1912) indicates that these bodies are really
organisms which seem to be symbiotic and not para-
sitic, although it remains to be proved what advan-
tage the host receives from their presence. Of a
similar sort are the Zooxanthellse which Mangan
(1909) has shown enter the developing ovum from
the parental tissues. All of these organisms become
in some way embedded in the germ cells, but so far
as we know never serve to distinguish the keimbahn,
although a more selective distribution within the
developing animal would obviously be greatly to
their advantage.

Vander Stricht (1911) has compared the *'beson-
dere Korper" found by Elpatiewsky (1909, 1910)
in the egg of Sagitta with several bodies, the *' corps
enigmatique," which he discovered in the oocyte of
the cat (Fig. 55 y E). One or two of these *' corps
enigmatique" are present in the young oocyte
originating from a few (one to five) cytoplasmic
safranophile granules which are visible at the begin-
ning of the growth period. They at first lie near the
nucleus, but as the size of the oocyte increases they
become situated near the periphery. Usually three
parts can be recognized in the *' corps enigmatique" :
*' granulation centrole, couche intermediaire et couche
corticale foncee." As the term applied to them indi-
cates, the functions of these bodies were not deter-
mined. The following suggestion is, however, made :
**il est possible que cet element nous montre, des
Forigine, la 'Keimbahn' ainsi que les premieres
cellules genitales constituees." A body stained

188 GERM-CELL CYCLE IN ANIMALS

deeply by nuclear dyes which was found by O. Van
der Stricht (1909) in the bat at the time of the first
cleavage mitosis may be similar to the "corps enig-
matique" of the cat.

In many animals no keimbahn-determinants nor
similar bodies have as yet been discovered. The best
we can do in cases of this sort is to determine from
what cleavage cell or cells the germinal epithelium
probably originates. For example, in Arenicola,
Lillie (1905) has shown that the part of the perito-
neum from which the germ cells arise develops from
teloblast cells which are probably derived (Child,
1900) from cell 4;d, At present, however, no charac-
teristics have been discovered which enable us to
distinguish between the germ cells and the somatic
cells in the early embryonic stages of such animals
(Downing, 1911).

CHAPTER VII

THE GERM CELLS OF HERMAPHRODITIC ANIMALS

Many of the most interesting biological problems
are those connected with the phenomenon of sex.
The term **sex" is applied to the soma or body of an
organism; it indicates the presence of certain mor-
phological and physiological characteristics, which
may be separated into primary and secondary sexual
characters. The primary sexual characters are those
immediately connected with the reproductive organs ;
the secondary sexual characters, such as the beard of
man, the brilliant feathers and beautiful songs of
many male birds, and the antlers of the moose, repre-
sent differences between male and female individuals
not directly concerned with the production of germ
cells. It is customary to speak of male germ cells
and female germ cells; this is not strictly proper,
since in only a few special cases can we predict the
sex of the individual which will develop from an egg.
Moreover, every germ cell must contain the poten-
tiality of both sexes since sooner or later its descend-
ants will give rise, some to male, some to female or
perhaps to hermaphroditic offspring. Thus the egg
is an initial hermaphrodite ; it may or may not be-
come an eventual hermaphrodite according to the sex-
ual condition of the individual to which it gives rise.

189

190 GERM-CELL CYCLE IN ANIMALS

All the species of Metazoa may be separated into
two groups. The individuals in one group of species

Fig. 56. — Diagram of the reproductive organs of the earthworm, dorsal
view. A, B, C, seminal vesicles ; N, nerve-cord ; O, ovary ; OD, ovi-
duct ; R, egg sac ; S, spermatheca ; SF, seminal funnel ; T, testes ;
VD, vas deferens. {From Marshall and Hurst.)

possess only one sort of reproductive organs (male or
female) and produce only one sort of germ cells (eggs
or spermatozoa) ; these species are said to be dice-

GERM CELLS OF HERMAPHRODITES 191

cious or gonochoristic. In the other group both
male and female reproductive organs occur in each
individual ; and such species are called monoecious
or hermaphroditic. The majority of animals are
gonochoristic, but a number of classes and orders
consist almost entirely of hermaphroditic species,
and probably no large group of animals is free from
species which are monoecious. A study of hermaph-
roditism is necessary for the elucidation of many
biological problems ; and some of those dealing
more directly with the germ-cell cycle will be con-
sidered in this chapter.

There are many variations in the morphology of
the reproductive organs in hermaphrodites. In
some, such as the earthworm (Fig. 56), the male
and female organs, consisting of all the parts typically
present in gonochoristic animals, are present and
entirely separate from each other. All gradations
between such a state and an intimate association of
male and female germ cells are known. Perhaps the
most interesting series occurs among the mollusks.
Here the germ gland may consist of two regions, as
in Pecten maximus, one of which gives rise to ova, the
other to spermatozoa ; or certain cysts may contain
only female germ cells and other cysts only male
germ cells, or both sorts of germ cells may occur in a
single cyst.

Hermaphroditism has been shown to be prevalent
among animals that are parasitic or sedentary, or for
some other reason may become isolated from their
fellows. Thus, it is of advantage for a parasite, such

192 GERM-CELL CYCLE IN ANIMALS

as the tapeworm, to be able to form both male and
female germ cells, since it may at any time become the
only one of its species to occupy the alimentary canal
of a host. Hermaphroditism in such a case, however,
is of no benefit if self-fertilization is not possible.
Although there are thousands of hermaphroditic
species of animals there are comparatively few whose
eggs are known to be fertilized by spermatozoa from
the same individual. We must therefore distinguish
between morphological and physiological hermaphro-
ditism and recognize the fact that the former condi-
tion is much more prevalent than the latter. Among
the species in which self-fertilization normally occurs
are certain rhabdocoels, digenetic trematodes, ces-
todes, ascidians, and mollusks. Van Baer, in 1835,
claims to have observed self-copulation in the snail,
Lymncea auricularia; that is, an individual with its
penis inserted in its own female opening. That
species of this genus fertilize their own eggs has
frequently been stated by investigators. Frequently
the spermatozoa of an hermaphrodite are capable
of fertilizing the eggs of the same individual,
but penetrate more readily the eggs of other individ-
uals. Such is the case in the ascidian, Ciona in-
testinalis (Castle, 1896; Morgan, 1905).

Both sorts of germ cells are seldom produced at
the same time by hermaphrodites. Those species
in which spermatozoa mature first are called protan-
dric; this is the usual condition. In a few cases
eggs are formed first and later spermatozoa; in-
dividuals in which this occurs are called protogynic.

GERM CELLS OF HERMAPHRODITES 193

Proterogyny has been described in certain ascidians
(Salpa), pulmonale gasteropods, and corals. That
hermaphrodites are not sexless but really animals
with double sex is well shown by the life history of
a worm, Myzostoma pulminar, which passes through
a short male stage during which spermatozoa are
produced, then a stage when no functional germ cells
are formed, and finally a female stage, characterized
by the development of eggs (Wheeler, 1896). Thus,
in this species, although hermaphroditic, there is no
functional hermaphroditic stage. All variations be-
tween this entire separation of the periods of germ-
cell development and the simultaneous production
of male and female germ cells have been recorded.
Some degree of protandry has been observed among
the sponges, coelenterates, flat worms, segmented
round-worms, mollusks, echinoderms, Crustacea, and
chordates.

Hermaphroditism may occur in only a few families,
genera, or species in a class. This is true, for example,
among the anthropods and vertebrates. Normally
the insects are called dioecious, but among bees, ants,
and butterflies, and more rarely other groups, individ-
uals appear which exhibit male characters on one side
of the body and female characters on the other, or the
anterior part may be male, the posterior female, etc.
(von Siebolt, 1864 ; Schultze, 1903 ; Morgan, 1907,
1913). Such a phenomenon is known as gynan-
dromorphism. Several hypotheses have been pro-
posed to account for this condition. Boveri has
suggested that if the egg nucleus should chance to

194 GERM-CELL CYCLE IN ANIMALS

divide before the sperm nucleus fuses with it, the
latter may unite with one of the daughter nuclei
of the egg nucleus ; this cell with this double nucleus
might then produce female structures, whereas the
other cell with only a single nucleus representing one-
half of the egg nucleus might give rise to male char-
acters. Morgan has proposed another theory which
is based on the fact that more than one spermatozoon
is known to penetrate the eggs of insects. If one
of these supernumerary spermatozoa should chance
to divide, it might result in the formation of male
structures, whereas the cells containing descendants
of the egg nucleus fused with another sperm nucleus
would exhibit female characteristics.

There is some evidence that true hermaphroditism
may exist among insects, at least during their embry-
onic and larval stages. Thus Heymons (1890) has
described in a young larva of the cockroach, Phyllo-
dromia gervianica, what appear to be rudimentary
egg-tubes, and in another larva eggs were found in
the testes which resembled those present in the egg-
tubes of female larvae of the same size (1 mm. in
length). More recently, Schonemund (1912) has
reported the presence of egg-tubes attached to the
anterior end of the testes of stone-fly nymphs {Perla
marginata) .

True hermaphroditism is rare in man and other
mammals, but several cases have been described in
the pig by Sauerbeck (1909) and Pick (1914), and in
man by Simon (1903), Uffreduzzi (1910), Gudernatsch
(1911), and Pick (1914).

GERM CELLS OF HERMAPHRODITES 195

One of the problems connected with hermaphrodit-
ism that has caused a great amount of discussion is
whether the dioecious or the monoecious condition
is the more primitive. The majority of zoologists
are inclined to consider the hermaphroditic condition
more primitive, but a number of careful investigators
have decided in favor of gonochorism. Among these
are Delage (1884), F. Muller (1885), Pelseener (1894),
Montgomery (1895, 1906), and Caullery (1913).

Very little is known regarding the segregation and
early history of the germ cells of hermaphrodites.
The principal results have been obtained from studies
on Sagitta by Elpatiewsky (1909), Stevens (19106),
and Buchner (1910a, 19106), and on Helix by Ancel
(1903), Buresch (1911), and Demoll (1912). Boveri
(1911), Schleip (1911), and Kruger (1912) have made
some interesting discoveries on the chromosome
cycle in nematodes, and likewise Zarnik (1911) on
pteropod mollusks. To this list we may add such
investigations as those of King (1910), Kuschake-
witsch (1910), and Champy (1913), on amphibians.

The segregation of the germ cells in Sagitta was
described and figured in Chapter VI (Fig. 54) . Here
the first division of the primordial germ cell is probably
differential; one daughter cell becomes the ancestor
of all the ova, the other of all the spermatozoa in the
hermaphroditic adult. None of the three investi-
gators who have studied this subject in Sagitta have
been able to discover with certainty any visible differ-
ences between the first two germ cells, but Elpatiew-
sky thinks the peculiar cytoplasmic inclusion, called

196 GERM-CELL CYCLE IN ANIMALS

by him the "besondere Korper," may be unequally
distributed between these cells, and that the one
which procures the larger portion is the progenitor
of the spermatozoa, the other of the ova. The evi-
dence for this view is, however, insufficient.

In Helix both eggs and spermatozoa originate in
every acinus of the ovo-testis ; it is therefore an ex-
cellent species for the study of the differentiation of
the sex cells. According to Ancel (1903) the anlage
of the hermaphroditic gland of Helix pomatia appears
a few hours before the larva hatches ; it consists of a
group of cells situated in the midst of the mesoderm,
from which germ layer it seems to originate. It
soon loses its rounded form and becomes elongate;
then a lumen appears wdthin it, thus changing it into
a vesicle whose wall consists of a single layer of cells
— a true germinal epithelium. Secondary, tertiary,
etc., vesicles bud off from the single original vesicle,
forming the acini of the fully developed gland. Cel-
lular differentiation takes place by the transformation
of the germinal epithelial cells into male, nurse, and
female elements. An indifferent epithelial cell is
shown in Fig. 57, A ; the chromatin granules are con-
densed to form irregular clumps. Some of these
indifferent epithelial cells increase in size and give
rise to indifferent progerminative cells ; the chroma-
tin clumps fuse, forming more or less spherical masses
(Fig. 57, E). From cells of this sort originate both
the oogonia and spermatogonia. The progermina-
tive male cell passes through the stages shown in
Fig. 57, B-D ; part of the chromatin of the progermi-

GERM CELLS OF HERMAPHRODITES 107

native cell loses its affinity for nuclear dyes ; the
chromatin masses become less numerous and more
nearly spherical ; and the entire cell increases in size,
the nucleus growing much more than the cytoplasm.
These progerminative male cells divide mitotically

Fig. 57. — Helix pomatia. Stages in differentiation of male and female
sex cells from indifferent cells. A. Epithelial indifferent cell.
E. Progerminative indifferent cell. B-D. Stages in transformation
of progerminative cell into a spermatogonium. F—G. Stages in
transformation of progerminative cell into an oocyte. (From Ancel,
1903.)

and then pass into the lumen of the acinus, where
they may be recognized as spermatogonia of the
first order.

After the spermatogonia have passed into the
lumen of the acinus the wall is seen to consist of two
groups of cells ; those of one group are central and in
contact with the spermatogonia, the others are periph-

198 GERM-CELL CYCLE IN ANIMALS

eral. The centrally situated cells now increase in
size ; but their nuclei retain the original condition ;
that is, the chromatin is present in irregular clumps.
These are nurse cells. After the nurse cells have
formed, certain of the peripheral cells increase in
volume and pass through an indifferent progermina-
tive stage (Fig. 57, E). Then they transform into
female progerminative cells, as shown in Fig. 57, F, G.
The chromatin clumps break up and become oriented
near the nuclear membrane, where they form a layer
of more or less rounded bodies bearing chromatic
filaments. In the meantime, both nucleus and cyto-
plasm increase in amount, especially the cytoplasm.
This (Fig. 57, G) represents an oocyte, which does not
divide before maturation.

Ancel concludes from these observations that there
are three successive periods of cellular differentiation
in the hermaphroditic gland of Helix: (1) the ap-
pearance of spermatogonia, (2) nurse cells, and
(3) oocytes. Both spermatogonia and oocytes pass
through the indifferent progerminative-cell stage, but
the nurse cells do not ; there are therefore two sorts
of differentiation of the indifferent epithelial cells.
Regarding the cyto-sexual determination, the follow-
ing hypothesis is advanced : A progerminative in-
different cell becomes a male or female element
according to its environment at the time of its trans-
formation ; if it appears before the nurse cells are
formed it becomes a spermatogonium ; if nurse cells
are already present it grows into an oocyte. The
discovery of certain individuals containing only male

GERM CELLS OF HERMAPHRODITES 199

elements is explained by Ancel by supposing the
transformation of the cells into sex cells to cease

Fig. 58. — Helix arhustorum. Stages in the differentiation of male and
female sex cells. A. Nucleus of germinal epithelium. B. Nucleus
of nurse cell. C. Nucleus of indifferent sex cell. D. Spermatogo-
nium of first order. E. Spermatogonium of second order. F. Grow-
ing oocyte. {From Buresch, 1911.)

before nurse cells are formed ; thus all the sex cells
would become spermatogonia.

More recently Buresch (1911) has repeated the

200 GERM-CELL CYCLE IN ANIMALS

work of Ancel, using Helix arbustorum for his materiaL
He confirms many of Ancel's results, objects to others,
and adds certain new observations. The germinal
epithelium is considered by Buresch to be a syncy-
tium containing both in young and old specimens
three sorts of cells, indifferent cells, egg cells, and
nurse cells. Likewise spermatogonia are present
not only in young but also in fully developed her-
maphroditic glands. This is contrary to Ancel's idea
of successive transformation. Buresch' s view is
indicated in Fig. 59. Here the vertical row of circles
represents the nuclei of the syncytial germinal epithe-
lium, some of which, as at m, change to indifferent
germ cells. These may pass into the lumen of the
acinus as spermatogonia of the first order {Sg. I)
and divide to form spermatogonia of the second order
(Sg. II) which grow into spermatocytes (Sc) ; sper-
matozoa are derived from these in the usual manner.
Other indifferent germ cells remain in the wall, as at
Wy and grow into oocytes, and a third class of cells
become nurse cells {71). In Fig. 58, A is shown a
nucleus of the germinal epithelium about 4 microns
by 6 microns in size. During differentiation into an
indifferent germ cell (Fig. 58, C) the chromatin forms
a nucleolus, and both nucleus and nucleolus increase
in size until the former reaches a diameter of about 7
microns. Those indifferent germ cells that are to
produce spermatozoa separate from the epithelium
with a small amount of cytoplasm and fall into the
lumen of the acinus as spermatogonia of the first
order (Fig. 58, D). These divide to form spermato-

GERM CELLS OF HERMAPHRODITES 201

Fig. 59. — Helix arhustorum. Diagram showing row of gorminal cijitho-
lial cells some of which, as at m, become spermatogonia and droj)
into lumen of germ gland; others beconu' nurse cells {n) ; and .still
others oocytes {w). Sgl = spermatogonium of first order; Sgll =
spermatogonium of second order ; Sc = spermatocyte ; St = sperma-
tid ; Sp = spermatozoa. {Fro7H Buresch, 1011.)

202 GERM-CELL CYCLE IN ANIMALS

gonia of the second order (Fig. 58, E). Those in-
different germ cells that are to form oocytes grow
large, remain in the germinal epithelium, and do not
divide. They possess a double nucleolus (Fig. 58, F) .
When a diameter of 36 microns is attained, the
oocyte passes out of the hermaphroditic gland into
the uterus.

The nurse cells, like the oocytes, remain in the wall
and do not divide ; their nuclei grow to be about 15
microns in diameter and the chromatin forms irregu-
lar clumps more or less evenly distributed (Fig. 48, B).
No differences could be discovered in the indifferent
germ cells by means of which the future history of
these cells could be determined. It was noted, how-
ever, that egg cells were never present without a
neighboring nurse cell, and the conclusion was
reached that a favorable position with regard to a
nurse cell determines whether an indifferent germ
cell shall develop into a spermatogonium or an egg.
If Buresch's observations are correct, Helix is not
protandric, but both sorts of germ cells mature at
the same time, and the fate of an indifferent germ
cell depends upon nutrition, that is, its proximity
to a nurse cell.

Demoll (19126) has proposed a new hypothesis
regarding sex determination and has selected certain
events in the oogenesis and spermatogenesis of Helix
poinatia as arguments in its favor. The hypothesis
is that the accessory chromosome (see Chapter IX)
contains the anlagen of the male sexual characters,
whereas the female sexual characters are localized

GERM CELLS OF HERMAPHRODITES 203

in the autosomes. In Helix the oogonia and sperma-
togonia arise from cells that are similar in size and
constitution (Fig. 60, A). When the germ-cell
nuclei reach the bouquet stage, a Nebenkern appears
near the side against which the chromatin threads

Fig. 60. — Helix pomatia. Stages in the difforontiation of male and
female sex cells. A. Young oocyte. B. Later stage of oocyte
showing faint Nebenkern. C. Young spermatocyte. D. Later
stage of spermatocyte showing well-marked Nebenkern. E. Still
later stage of spermatocyte containing Nebenkern consisting of
banana-shaped rods. (From Demoll, 1912.)

become packed. This Nebenkern is probably a
product of the nucleus; it appears in the female
cell only as a slightly darker area of cytoplasm (Fig.
60, B) but in the male cell is more dense (Z)), later
consisting of a number of darkly staining banana-
shaped pieces {E). With the appearance of the
Nebenkern the specific growth of the female cells

204 GERM-CELL CYCLE IN ANIMALS

is initiated. The Nebenkern disappears in the
oocyte soon after the yolk begins to form. The
chromatin threads in the spermatocytes break down
and lose their affinity for dyes, but later reappear.
In the oocyte, on the contrary, the chromatin threads
persist. Demoll concludes from these observations
that the Nebenkern always determines the character
of the germ cells, which, up to its formation, may be
called indifferent germ cells. He further concludes,
that, since in dioecious animals sex is determined by
the accessory chromosomes, in Helix the sexual
specificity of the Nebenkern must be determined
by the accessory chromosomes. Such chromosomes
were described by Demoll (1912a) in a previous
contribution.

A similar idea has been expressed by von Voss
(1914) regarding the differentiation of indifferent
germ cells in a flat-worm, Mesostoma ehrenbergi.
In the embryo of this hermaphrodite the germ gland
is a syncytium containing both the nuclei of future
oogonia and future spermatogonia. The cytoplasm
is apparently homogeneous throughout. The forma-
tion of the oogonia from indifferent germ cells begins
with the appearance of a " germinal- vesicle stage";
this is followed by an increase in the amount of
cytoplasm surrounding them. Since the cytoplasm
appears to be similar in all parts of the syncytium,
differentiation must be initiated by the nucleus,
and the suggestion is made that perhaps the accessory
chromosome may be responsible for the separation
of the germ cells into oogonia and spermatogonia.

GERM CELLS OF HERMAPHRODITES 205

The investigators whose results have been de-
scribed above have thus furnished three theories re-
garding the differentiation of male and female germ
cells in hermaphrodites : (1) In Sagitta, according
to Elpatiewsky, it is an unequal distribution of the
**besondere Korper," (2) in Helix, according to Ancel
and Buresch, it is due to the presence or absence of a
nurse cell in the immediate neighborhood, and (3) in
Helix, Demoll considers it a result of the influence of
the accessory chromosome. It is perfectly obvious
that hermaphrodites offer exceptionally fine material
for the study of the differentiation of germ cells, but
that thus far the results have not furnished an ade-
quate explanation of the phenomenon. The investi-
gations of Boveri (1911), Schleip (1911), and Krueger
(1912) on the chromosomes in hermaphroditic nema-
todes may be discussed more profitably during the
consideration of the chromosome cycle in the next
chapter.

Certain morphological and experimental studies
on the germ glands of amphibians are of interest be-
cause both oogonia and spermatogonia are sometimes
more or less closely associated in a single individual
during the developmental stages, and may persist
even in the adult germ glands of a number of species
which are commonly considered dioecious. Pfluger,
for example, was able to separate the young of the
frog, Rana temporaria, into three groups, males, fe-
males, and hermaphrodites ; the hermaphrodites
developed into either males or females. Similar
results were obtained by Schmidt -Marcel (1908)

206 GERM-CELL CYCLE IN ANIMALS

and Kuschakewitsch (1910), who refer to the her-
maphroditic individuals as intermediates.

There is no consensus of opinion regarding the
origin of the germ cells in amphibians ; one group
of investigators, including Allen (1907) and King
(1908), recognize a definite keimbahn, whereas many
others (Semon, 1891; Bouin, 1900; Dustin, 1907;
Kuschakewitsch, 1910 ; Champy, 1913) believe they
arise from the germinal epithelium or near-by cells.
Very few students have attempted to determine the
stages in or causes of the differentiation of male and
female cells from the primordial germ cells. Kuscha-
kewitsch (1910) concludes from his extensive studies
on the history of the germ cells in frogs that at first
all of the germ cells are indifferent but subsequently
become differentiated in two directions. Champy
(1913) has studied this differentiation in a number
of amphibians and has concluded that if the charac-
teristically lobed or polymorphic nuclei of the pri-
mordial germ cells in Bufo, Hyla, and Rana temporaria
lose their original shape and become spherical and
clear, the germ gland will form an ovary ; but if the
nuclei retain their primitive condition, a testis will
result. Champy believes with Kuschakewitsch that
both sorts of germ cells arise from sexually indifferent
cells, that is, sex is not irrevocably fixed in the fer-
tilized egg. Furthermore Champy' s observations
have led to the conclusion that the germ cells in the
sexually indifferent germ glands are morphologically
identical with primitive spermatogonia. These in-
different germ cells become differentiated into ova

GERM CELLS OF HERMAPHRODITES 207

or spermatozoa as a result of various causes, some
general and others local in nature, which probably
are most influential at certain definite stages in the
cellular activity. A new equilibrium is thereby es-
tablished between the different cell organs which
initiates new processes resulting in differentiation.
The undifferentiated cells in the testis of the adult
appear also to be identical with the primitive sper-
matogonia, and have still the power of producing
either ova or spermatozoa. Thus the male amphib-
ians are also females **en puissance," but the re-
verse is not true. This accounts for the numerous
discoveries of ova in the testes of these animals.

Reports of so-called hermaphroditism in amphib-
ians are abundant in the literature. Cases have
been reported in frogs by Cole (1895), Friedmann
(1898), Gerhartz (1905), Ognew (1906), Yung
(1907), Schmidt-Marcel (1908), Youngman (1910),
Hooker (1912), and many others. Hooker has re-
viewed the literature of the subject. Hermaphrodit-
ism in other amphibians is more rare, but it has
been noted in salamanders by La Vallett St. George
(1895) and Feistmantel (1902). Usually the condi-
tion spoken of as hermaphroditism consists in the
presence of ova in the testis, and it is probable that
true hermaphroditism is rare in these animals as it
is in other vertebrates. In the toad, however, a
condition exists which is of particular interest. The
genital ridge of every toad tadpole 15 to 18 days old
becomes visibly differentiated into two regions, an an-
terior portion which develops into Bidder's Organ, and

208 GERM-CELL CYCLE IN ANIMALS

a posterior region which becomes an ovary or testis.
Bidder's Organ persists in the adult of males, where
it lies just anterior to the testis, but in the females
of Bufo variabilis, B. cinereus, B. clamita, and B.
lentiginosus it disappears at the end of the second
year. Bu^o vulgaris seems to differ from the other
species since here Bidder's Organ persists, becom-
ing small and shrunken during the winter (Ognew,
1906) and regenerating during the summer months
(Knappe, 1886). At first the cells in both the
anterior and posterior portions of the genital ridge
are similar, all possessing a polymorphic nucleus,
and dividing mitotically, but later those of Bidder's
Organ begin to divide amitotically and assume the
characteristics of young oocytes with rounded nuclei.
Knappe (1886) claims that these cells never become
functional ova because they are unable to form yolk.
King (1908), however, does not consider this prob-
able, but traces their differentiation to irregularities
in the synizesis stage.

By most investigators Bidder's Organ is regarded
as a rudimentary ovary. Others believe that the
Amphibia were derived from hermaphroditic ances-
tors and that in the male it is a rudimentary ovary
and in the female a rudimentary testis. This seems
more probable than Marshall's suggestion that this
organ is the result of degenerative processes proceed-
ing backward from the anterior end of the genital
ridge, or than that it represents the remains of a
sex gland possessed by the larvae of ancestral toads
when they were paedogenetic, as Axolotl is at the

GERM CELLS OF HERMAPHRODITES 209

present time. Champy (1913) has found that the
cells of Bidder's Organ in Bufo pantherina pass
through stages in their transformation similar to
those of the primitive germ cells of liana esculenta
which become ova, and is incHned to the view that
the principal difference between the toad and the
intermediate type of young frogs lies in the fact
that in the former the oviform cells are localized in
Bidder's Organ, whereas in the frog they are scattered
throughout the germ gland.

The development of the germ glands in the hag-
fish, Myxine glutinosa, resembles that in the toad
in many respects. Cunningham (1886) and Nansen
(1886) considered Myxine to be a protandric her-
maphrodite. Schreiner (1904), however, was able to
show that every adult is functionally male or female
with a rudimentary ovary anteriorly situated and a
posterior, mature testis, or a functional ovary ante-
rior to a rudimentary testis. These results were con-
firmed by Cole (1905).

Similar conditions have been found by Okkelberg
(1914) in the young of the brook lampre3% E?iio-
sphenus wilderi. Of fifty larvae ranging from 7|
cm. to 20 cm. in length, 46 per cent were true
females, 10 per cent were true males, and 44 per cent
were hermaphrodites. Since male and female adults
are approximately equal in numbers, it was concluded
that the juvenile hermaphrodites become adult
males. In favor of this conclusion is also the fact
that the adult males frequently possess ova in their
gonads which resemble those present in the her-
maphroditic larvae.

210 GERM-CELL CYCLE IN ANIMALS

Regarding the differentiation of the germ cells in
hermaphrodites then we may recognize two principal
views : (1) that there is some material within the
cell which initiates specialization, or (2) that differ-
entiation is due to general or local causes outside
of the germ cells. The former is favored by Elpatiew-
sky (1909, 1910) from studies on Sagitta and by
Demoll (1912) from studies on Helix. The second
view is more widely advocated. The conclusions
derived by Kuschakewitsch (1910) and Champy
(1913) on amphibians, and of Ancel (1903) and
Buresch (1911) on Helix agree in their essential fea-
tures. All of these investigators maintain that the
sex cells pass through an indifferent stage and are
differentiated into oocytes or spermatocytes because
of influences external to themselves. Buresch and
Champy also believe that even in the fully developed
germ glands of the adult these primitive cells are
present. The causes of their differentiation, how-
ever, have not been definitely determined.

CHAPTER VIII

KEIMBAHN-DETERMINANTS AND THEIR SIG-
NIFICANCE

It is customary to be suspicious of any peculiar
bodies revealed to us in fixed and stained material
under high magnification. There can be no doubt,
however, that most, if not all, of the cytoplasmic
inclusions mentioned in the preceding chapters are
realities and not artifacts. Some of them have been
seen in the living eggs ; most of them have been de-
scribed by several investigators; they occur after
being fixed and stained in many different solutions ;
and their presence is perfectly constant. The
genesis, localization, and fate of these bodies are
difficult to determine, and their significance is prob-
lematical ; but the writer has attempted in the follow-
ing pages to draw at least tentative conclusions from
the evidence available and to indicate what still
needs to be done.

A. The Genesis of the Keimbahn-determinants

The writers who have discussed the origin of the
keimbahn-determinants have derived them from
many different sources. In a few cases they are known
to be nuclear in origin, consisting of nucleolar or chro-
matic materials; they are considered differentiated

211

212 GERM-CELL CYCLE IN ANIMALS

parts of the cytoplasm by some investigators; in
some species they are extra-cellular bodies, such as

nurse cells.

The accompanying table indicates the number and
diversity of the animals in which keimbahn-determi-
nants have been described, and shows the increasing
interest that has been given to this subject within re-
cent years, over half of the papers listed having been
published since 1908. Several cases have been re-
ferred to in the text, but omitted from the table be-
cause of insuflacient evidence regarding their connec-
tion with the primordial germ cells. The list as
given includes representatives of the Ccelenterata,
Ch^tognatha, Nematoda, Arthropoda, and Ver-
TEBRATA. The tcrms applied to the various sub-
stances have been chosen evidently because of their
genesis, position in the egg, or supposed function.

Table of Principal Cases of Visible Substances con-
cerned IN Differentiation of Germ Cells (in Chron-
ological Order)

Name of Species,

Name Applied to

AUTHOBITY

Date

Genus, ob Group

Substance

Chironomus nigro-

Dotterkornchen

Weismann

1863

viridis

Miastor

Dottermasse

Metchnikoff

1866

Moina rectirostris

Richtiingskorper

Grobben

1879

Chironomus

Keimwulst

Ritter

1890

Daphnidae

Paracopulations-
zelle

Weismann and
Ischikawa

1889

^quorea

Metanucleolus

Haecker

1892

Ascaris megaloce-

Chromatin

Boveri

1892

phala

KEIMBAHN-DETERMINANTS

213

A. lumbricoides

A. niiiicunda

Chromatin

0. Meyer

189.5

A. labiata '

Cyclops

Aussenkornchen

Haecker

1897

Ektosomen

Haecker

1903

Calliphora

Dotterplatte

Noack

1901

Dytiscus

Anello cromatico

Giardina

1901

Apis mellifica

Richtungskorper

Petrunkewitsch

190^2

Parasitic |
Hymenoptera J

Nucleolo

Silvestri

1906
1908

Chrvsomelidae

Pole-disc

Hegner

1908

Miastor metraloas

polares Plasma

Kahle

1908

Sagitta

besondere
Korper

Elpatiewsky

1909

Guinea-pig

Chondriosom.es

Rubaschkin

1910

Chick

Chondriosomes

Tschaschkin

1910

Lophius

extruded
plasmosome

Dodds

1910

Ascaris

Plasmadifferen-

Boveri

1910

Chironomus

zen
Keimbahn-
plasma

Hasper

1911

Copepoda

Ectosomen

Amma

1911

Polyphemus

Nahrzellenkern

Kiihn

fl911
[1913

Sagitta

Keimbahn-
chromidien

Buchner

1910

Man

Sertoli cell
determinant

Montgomery

1911

Chick

Attraction-
sphere '

Swift

1914

Parasitic

Keimbahn-

Hegner

1914

Hymenoptera

chromatin

a. Nuclear. Nucleoli. It seems certain that
bodies of a nucleolar nature behave as keimbahn-
determinants. There are three or more kinds of
bodies that are spoken of as nucleoli. Of these may
be mentioned (1) the true nucleoli or plasmosomes, (2)
karyosomes or chromatin-nucleoli, and (3) double-nu-

214 GERM-CELL CYCLE IN ANIMALS

cleoli, consisting of usually a single principal nucleolus
(Hauptnucleolus of Flemming), and one or more
accessory nucleoli (Nebennucleoli of Flemming).
Many nucleoli have been described that may perhaps
represent intermediate stages in the evolution of one
of the types mentioned above into another.

The young ovarian egg of most animals contains a
single spherical nucleolus ("Keimfleck," or "germi-
nal spot"), but the number may increase greatly dur-
ing the growth period. Usually during the formation
of the first maturation spindle the nucleolus escapes
from the nucleus into the cytoplasm, where it dis-
appears, often after breaking up into fragments.
Many theories have been advanced regarding the
origin, function, and fate of the nucleoli of the germi-
nal vesicle. They are considered by some of chro-
matic origin, arising as an accumulation of the chro-
matin, or from the chromatin by chemical trans-
formation. Others consider them extra-nuclear in
origin (Montgomery, 1899).

Many functions have been attributed to the nu-
cleoli ; of these the following may be mentioned :
(1) They function as excretory organs (Balbiani,
1864 ; Hodge, 1894) ; (2) nucleoli play an active
role in the cell, since they serve as storehouses of
material which is contributed to the formation of the
chromosomes (Flemming, 1882 ; Lubosch, 1902 ;
Jordan, 1910; Foot and Strobell, 1912) and may
give rise to kinoplasm (Strasburger, 1895) or *'Kine-
tochromidien " (Schaxel, 1910); (3) nucleoH are
passive by-products of chromatic activity; they

KEIMBAHN-DETERMIXANTS 215

become absorbed by active substances (Haecker,
1895, 1899) ; (4) nucleoli represent nutritive material
used by the nucleus into which it is taken from the
cytoplasm (Montgomery, 1899).

Undoubtedly the various bodies known as nucleoli
originate in different ways, have different histories,
and perform different functions.

In the particular cases to be discussed here the
nucleoli are not temporary structures, as is usually
true, but persist for a comparatively long interval after
the germinal vesicle breaks down. What seemed to
be the most important and convincing evidence of
the functioning of a nucleolus as a keimbahn-determi-
nant is that furnished by Silvestri (1906, 1908) in
parasitic Hymenoptera. As shown in Chapter V,
however, the "nucleolo" of Silvestri is really not a
nucleolus but consists of chromatin.

As we have already noted, in a few instances the
nucleolus does not disappear during the maturation
divisions but persists for a time as a "metanucleolus"
(see p. 183). These metanucleoli are evidently of
a different nature from the usual type and are hence
saved from immediate disintegration in the cyto-
plasm. The localization of the metanucleolus in the
egg is the result of either its own activity, or that of
the surrounding cytoplasm, or a combination of these.
Gravity can have no decided effect upon it (Herrick,
1895), since its position is constant, whereas the posi-
tion of the egg with respect to gravity is not. It
also seems hardly possible that oxygenotactic stimuli
are the cause of its localization, as has been suggested

216 GERM-CELL CYCLE IN ANIMALS

by Herbst (1894, 1895) for the migration of the
blastoderm-forming cells from the center to the sur-
face of the eggs of certain arthropods.

Haecker (1897) has suggested that the "Aussen-
kornchen" which appear in the egg of Cyclops during
the formation of the first cleavage spindle may be
nucleolar in nature. Later (1903) this idea was
withdrawn, and more recently Amma (1911) has
likewise been unable to sustain this hypothesis. The
most convincing data furnished by Amma are that in
an allied form, Diaptomus coeruleus (Fig. 49, H), these
granules appear before the cleavage spindle is formed
and before the nucleoli of the pronuclei have disap-
peared.

The remaining forms in which nucleoli have been
considered as keimbahn-determinants are merely
suggestive. In Mquorea, Haecker (1892) traced the
metanucleolus, which arises from the germinal vesicle,
into certain cells of the blastula. Similar bodies
appear in Mitrocoma (Metchnikoff, 1886), Tiara
(Boveri, 1890), Stephanophyes (Chun, 1891), Myzo-
stoma (Wheeler, 1897), and Asterias (Hartmann, 1902),
but their ultimate fate has not been determined.
Meves (1914), however, has traced the middle piece
of the sperm of the sea urchin. Par echinus miliaris,
into one of the cells of the animal half of the egg at
the thirty-two-cell stage. This middle piece is of a
plastochondrial nature.

It seems probable that in all these cases the same
influences may be at work regulating the time, the
place, and the method of localization of the nucleoli.

keimbahn-deter:\il\ants 217

The writer can only conclude (1) that the metanu-
cleoli differ in nature from ordinary plasmosomes,
chromatin-nucleoli, and double-nucleoli ; (2) that
these bodies are definitely segregated in a certain part
of the egg or in a certain blastomere, probably by
protoplasmic movements ; (3) and that their disin-
tegration and the distribution of the resulting frag-
ments or granules are controlled by reactions between
them and the substances in which they are embedded.
Chromatin. In two genera of animals the differ-
entiation of the primordial germ cells is accompanied
by a diminution of the chromatin in the nuclei of
the somatic cells, so that eventually the nucleus of
every germ cell is provided with the full complement
of chromatin, whereas the nucleus of every somatic
cell lacks a considerable portion of this substance,
which remains behind in the cytoplasm when the
daughter nuclei are reconstituted. These two genera
are Ascaris and Miastor. This diminution process
was described by Boveri (1892) in the former and
confirmed by O. Meyer (1895) and Bonnevie (1902),
and by Kahle (1908) in Miastor and confirmed by
Hegner (1912, 1914a). For details of these processes
reference should be made to Figs. 15-16, 51-52, and
pp. 57 and 174. It may be pointed out here that
although the final results are similar the process dift'ers
in the two genera. In Ascaris both ends of each
chromosome are split off, whereas in Miastor approxi-
mately one-half of each daughter chromosome is left
behind to form the *'Chromosomenmitteli)hitte"
(Fig. 16) and later the *'Chromatinreste" (Fig. 18).

218 GERM-CELL CYCLE IN ANIMALS

The elimination of chromatin during the matura-
tion and early cleavage divisions of the egg, as well
as during the mitotic divisions of other kinds of cells,
has often been recorded. For example, Wilson
(1895, p. 458) estimates that only about one-tenth
of the chromatin in the germinal vesicle of the star-
fish is retained to form the chromosomes during the
first maturation division, and Conklin (1902) finds
that "in Crepidula the outflow of nuclear material
occurs at each and every mitosis" (p. 51). Further-
more, Rhode (1911) argues that chromatin-diminu-
tion is a normal histological process, and describes
such phenomena in blood cells, nerve cells, and
cleavage cells of several Amphibia, comparing con-
ditions with the chromatin-diminution in Ascaris
and Dysticus}

Diminution processes similar to those in Ascaris
and Miastor have not been discovered in other ani-
mals, although investigators have been on the watch
for such phenomena and have studied allied species,
e.g., the work of Hasper (1911) on Chironomus and
my own work on the chrysomelid beetles (see pp. 108

^ His conclusion is as follows: "In der Histogenese der allerver-
schiedensten Gewebe tritt uns also die Erscheinung entgegen, dass
die sich entwickelnden Zellen, bzw. Kerne einen Teil ihres Chromatins
abstossen, d. h. also eine Chromatindiminution erfolgt, wenn auch
die Befunde selbst im speziellen von den bisher beobachteten in der
Einleitung beschriebenen Fallen der Chromatindiminution etwas ab-
weichen.

"Eine Chromatindiminution tritt also nicht nur am Anfang und Ende
der Keimbahn, wie es bisher angegeben worden ist, sondern in den ver-
schiedensten Entwicklungsstadien und bei den verschiedensten Geweben
undTieren ein, sie hat also offenbar eine allgemeine Bedeutung." (p. 25.)

KEIMBAHN-DETERMINANTS 219

to 118). If, therefore, there is a similar difference in
all animals in chromatin content between the germ
cells and somatic cells, the elimination of chromatin
from the latter must take place by the transformation
of the basichromatin of the chromosomes into oxy-
chromatin which passes into the cytoplasm during
mitosis, or else by the more direct method advocated
by the believers in the chromidia hypothesis.

The causes of the diminution of chromatin in As-
caris and Miastor are unknown. Recently Boveri
(1910) has concluded from certain experiments on
the eggs of Ascaris (see p. 177) that in this form it is
the cytoplasm in which the nuclei are embedded that
determines whether or not the latter shall undergo
this process. Kahle (1908) does not explain the
cause of the diminution in Miastor. To the writer it
seems more important to discover why the nuclei
of the keimbahn cells do not lose part of their chro-
matin, since the elimination of chromatin during
mitosis is apparently such a universal phenomenon.
I would attribute this failure of certain cells to under-
go the diminution process not to the contents of the
nucleus alone but to the reaction between the nucleus
and the surrounding cytoplasm. As stated in a
former paper (Hegner, 1909a), *'In Calligrapha all
the nuclei of the egg are apparently alike, potentially,
until in their migration toward the surface they
reach the ' Keimhautblastem ' ; then those which
chance to encounter the granules of the pole-disc
are differentiated by their environment, i.e., the
granules, into germ cells. In other words, whether or

220 GERM-CELL CYCLE IN ANIMALS

not a cell will become a germ cell depends on its posi-
tion in the egg just previous to the formation of the
blastoderm."

Similarly in Ascaris the cleavage nuclei are con-
ceived as similar so far as their ''prospective potency"
is concerned, their future depending upon the char-
acter of their environment, i.e., the cytoplasm. In
the egg of Miastor cleavage nucleus IV (Fig. 15) does
not lose part of its chromatin because of the character
of the reaction between it and the substance of the
"polares Plasma." In chrysomelid beetles (Hegner,
1908, 1909, 1914a) and Ckironomus (Hasper, 1911),
however, although no diminution process has been
discovered in the nuclei that encounter the pole-disc
or "Keimbahnplasma," the other nuclei in the egg,
so far as known, are similar in this respect. The
nuclei of the primordial germ cells, however, may be
distinguished easily from those of the blastoderm
cells in chrysomelid beetles, proving conclusively
that a differentiation has taken place either in one
or the other. This differentiation probably occurs in
the nuclei that take part in the formation of the
blastoderm, since the nuclei of the germ cells retain
more nearly the characteristic features of the pre-
blastodermic nuclei, w^hereas those of the blastoderm
cells change considerably.

In some cases the eliminated chromatin may have
some influence upon the histological differentia-
tion of the cell, since it is differentially distributed
to the daughter cells, but in Ascaris and Miastor
no mechanism exists for regulating the distribution

KEIMBAHN-DETERMINANTS 221

of the cast-out chromatin, and there is consequently
no grounds for the hypothesis that "in Ascaris those
cells which become body cells are the ones that in-
clude the cast-off chromosome ends in their cyto-
plasm, and it will pro})a})ly be found that these
ejected chromosome parts engender such cytoplasmic
differentiations as characterize the body cells "
(Montgomery, 1911, p. 192).

Chromidia. To several of the bodies listed in
the table on page 88 as keimbahn-determinants has
been ascribed an origin from the chromatin of
the germinal vesicle. Many cases of the elimination
of chromatin from the nuclei of growing oocytes are
to be found in the literature. Blochmann (1886) dis-
covered a process of "budding" in the oocytes of
Camponotus ligniperda resulting in the formation
of "Nebenkerne." These appear first as small
vacuoles lying near the nucleus ; later they contain
small staining granules and acquire a membrane.
The "Nebenkerne" grow in size and increase in num-
ber, while the nucleus of the oocyte becomes smaller.
Stuhlmann (1886) described a similar phenomenon
in about a dozen different species of Hymenoptera.
The oocyte nucleus in all species examined becomes
localized near the anterior end ; then the small
nuclear-like bodies form around it at its expense.
The time of their i)roduction varies in the differenl
species ; in some they appear in the very young
eggs ; in others not until a much later stage has been
reached. Sometimes they fuse to form a large
"Dotterkern" lying at the posterior pole of the egg;

222 GERM-CELL CYCLE IN ANIMALS

or they may remain separate and later become scat-
tered. Paulcke (1900) also noted nuclear-like bodies
near the oocyte nucleus of the queen bee, and Mar-
shall (1907) has likewise found them in Polistes
pallipes. In this species the nuclear-like bodies
form a single layer around the nucleus; later they
come to lie near the periphery of the oocyte and
finally disappear. Loewenthal (1888) has described
what appears to be chromatin in the cytoplasm of
the egg of the cat, and an elimination of chromatin
was noted by van Bambeke (1893) in the ovarian
egg of ScorpcBTia scrofa. In none of these species,
however, have keimbahn-determinants been dis-
covered.

According to Buchner (1910) the "besondere
Korper" in the egg of Sagitta, and in fact keimbahn-
determinants in most other animals are of a chromid-
ial nature, representing the tropho-chromatin de-
manded by the binuclearity hypothesis. The term
chromidia was introduced by R. Hertwig in 1902 and
apphed to certain chromatin strands and granules
of nuclear origin in the cytoplasm of AdinosphoBrium.
Goldschmidt (1904) transferred the chromidia hy-
pothesis to the tissue cells of Ascaris. Since then
chromidia have been described in the cells of many
animals, including both somatic and germ cells.
Thus far the group of zoologists that favor the
chromidia idea have not received very extensive
backing, but the fact remains that chromatin
particles are in some cases cast out of the nuclei in
the oocytes of certain animals and continue to exist

KEIMBAHN-DETERMINAXTS 223

as such in the cytoplasm for a considerable period.
It is also possible that, as Buchner (1910) maintains,
the keimbahn-determinants may be in reality **Keim-
bahnchromidien."

This view was suggested by the writer in 1909
(p. 274) to account for the origin of the pole-disc
granules in the eggs of chrysomelid beetles. It was
thought that here as in the Hymenoptera (Bloch-
mann, 1886 ; et at.) chromatin granules might be
cast out of the nuclei of the oocytes, and that these
granules might gather at the posterior end to form
the pole-disc. It was also suggested that chromatin
granules from the nurse-cell nuclei might make their
way into the oocyte and later become the granules of
the pole-disc. It should not be forgotten, moreover,
that these granules stain like chromatin. Finally,
mention should be made of the "anello cromatico"
of Giardina (1901) which is associated with the
differentiation of the oocytes in Dytiscus (see p. 1^20,
Fig. 38), and the keimbahn-chromatin which I have
recently described (Hegner, 19146) in the eggs of
the parasitic hymenopteron, Copidosoma (p. 151,
Figs. 46-47).

Conclusion. Certain keimbahn-determinants
may consist of nucleolar material which is derived
from the germinal vesicle and persists until the
primordial germ cells are established. In some cases
the keimbahn cells are characterized by the posses-
sion of the complete amount of chromatin in con-
trast to the somatic cells which lose a part of this
substance. Since, however, the chromatin-diminu-

224 GERM- CELL CYCLE IN ANIMALS

tion process does not occur in many species, it is
probably not a universal phenomenon, and conse-
quently cannot be of fundamental importance. Most
of the evidence, on the other hand, points toward
the conclusion that all of the cleavage nuclei are
qualitatively alike, and that the cytoplasm is the
controlling factor.

b. Cytoplasmic or Extracellular Nutritive
Substances. It was pointed out on a preceding
page (p. 101) that one of the characteristics used to
distinguish primordial germ cells from other embry-
onic cells is the presence within them of yolk material.
In many vertebrates the yolk globules persist in the
primordial germ cells until a comparatively late
stage, and indeed are often so numerous as to practi-
cally conceal the nuclei of these cells. A large num-
ber of the keimbahn-determinants that have been
described are supposed to consist of nutritive sub-
stances. Some of the earliest investigators were
aware of the yolk content of the primordial germ
cells. For example, in Chironomus Weismann (1863)
found four oval nuclei lying in the " Keimhautblas-
tem " at the posterior end of the egg, each of which
is associated with one or two yolk granules ; these
are the "Polzellen." In another Dipteron, Simula
sp., Metchnikoff (1866) records four or five pole-
cells which possess fine yolk granules in their cell
substance. The same author (1866) also states that
when the pseudovum in the psedogenetic larva of
Miastor contains twelve to fifteen nuclei, one of
these, together with the dark yolk-mass in which it

KEIMBAHN-DETERMINANTS 225

lies, is cut off as a cell which gives rise to the pole-
cells.

In certain Daphnid^, Weismann and Ischikawa
(1889) describe a "Paracopulationszelle" wliicli is
derived from the contents of the germinal vesicle
(seep. 163) ; but the recent work of KUhn (1911, 1913)
renders it probable that this body is nothing but
the remains of a nurse cell. The "Dotterplatte"
discovered by Noack (1901) at the posterior end of
the egg of Calliphora (Fig. 34) is considered by this
investigator to consist of yolk elements. In previous
communications (Hegner, 1908, 1909, 1911) the
writer has discussed the probability that the pole-disc
in chrysomelid eggs consists of nutritive material,
and Weiman (1910a) also has offered arguments
for this view.

The granules segregated in certain cleavage cells of
Neritina (Blochmann, 1881), Asplanchna (Jennings,
1896), Lepas (Bigelow, 1902), Siphonaria (Fujita,
1904), and Physa (Wierzejski, 1906) may be of a
nutritive nature, and these cells may be the stem
cells from which the germ cells of these animals
eventually arise. The hypothesis that the nucleoli
consist of food substance also argues in favor of the
idea that the keimbahn-determinants are nutritive.

The importance of these nutritive substances
to the primordial germ cells can be stated with some
degree of certainty. According to some authorities
the primordial germ cells remain in the j)rimitive
condition and do not undergo differentiation at the
same time, or at least at the same rate, as do the

ne GERM-CELL CYCLE IN ANIMALS

other embryonic cells. On this account their yolk
contents are not at first utilized, since their meta-
bolic activities are so slight. This is more especially
true of the vertebrates in which, it has been sug-
gested (Hegner, 1909a, p. 276), the yolk contents
of the germ cells are transformed into the energy of
motion during the characteristic migration of these
cells into the germinal epithelium. Why these
nutritive substances are segregated in the primordial
germ cells is more difficult to answer. Finally, it is
interesting to note that the differentiation of the
indifferent germ cells of Helix arbustorum into sper-
matogonia or oogonia has been found to depend
upon nutrition (Buresch, 1911).^

Yolk Nucleus. There are many bodies in the
cytoplasm of growing oocytes that have been called
yolk nuclei and that may be responsible for the
origin of the keimbahn-determinants. Some of
these bodies have already been considered, but the
term 'y^lk nucleus' has been applied to so many
different cytoplasmic inclusions (Munson, 1912)
that no attempt will be made here to describe them
nor to trace their history.

Mitochondria. The condition of the chondrio-
somes in the primordial germ cells of certain verte-
brates (Rubaschkin, 1910, 1912; Tschaschkin, 1910;
Swift, 1914) and the theories that have been pro-

i"Ob aber eine indifferente Geschlechtszelle sich in mannlicher oder
weiblicher Richtung weiter entwickeln wird, das konnen wir schon sehr
friih sagen, namlich nach der Lage dieser Zelle naher oder weiter von
einer Nahrzelle " (p. 327).

KEIMBAHN-DETERMINANTS 227

posed regarding the role of these bodies in heredity
make it necessary to refer to them briefly here. At
the present time it is difficult to make any definite
statement regarding the origin, nature, and signifi-
cance of the various cytoplasmic inclusions that have
been grouped under the general title of mitochondria.
It seems probable that we are concerned with a num-
ber of different sorts of inclusions, and with various
stages in their evolution. In the guinea pig (Ru-
baschkin, 1910, 1912) and chick (Tschaschkin, 1910)
the chondriosomes of the cleavage cells are spherical
and all similar, but, as development proceeds, those
of the cells which become differentiated to produce
the germ layers unite to form chains and threads,
whereas those of the primordial germ cells remain
in a spherical and therefore primitive condition
(Fig. 31, B). Swift (1914) has found, however,
that in the chick the mitochondria in the primordial
germ cells are not at all characteristic, resembling
those of the somatic cells. The germ cells neverthe-
less can be distinguished from the latter by the pres-
ence of an especially large attraction-sphere (Fig.
31, D). This distinction between the primordial
germ cells and the surrounding somatic cells may
enable us to trace the keimbahn in vertebrates back
into cleavage stages — something that has not been
accomplished as yet.

An examination of the various keimbahn-deter-
minants Hsted in the table (p. 212) has led the writer
to conclude that none of them is of a mitochondrial
nature, but the results obtained by the special methods

228 GERM-CELL CYCLE IN ANIMALS

employed by students who are studying mitochondria
give us good reason to hope that other substances
may be made visible which will help to clear up the
problem of primary cellular differentiation.

Metabolic Products. Among the most difficult
cases to explain are those of Sagitta and certain cope-
pods, since here the keimbahn-determinants ap-
parently arise de novo in the cytoplasm. Buchner's

(1910) contention that the "besondere Korper"
of Sagitta is the remains of the *' accessory fertiliza-
tion cell" of Stevens (1904) is not sustained by either
Stevens (1910) or Elpatiewsky (1910). The idea of
the nucleolar nature of the *'Aussenkornchen " in
Cyclops has been discarded by Haecker (1903)
and the conclusion reached that these granules are
similar to nucleoli in one respect, namely, they are
by-products of activities within the cell. Amma

(1911) has considered this subject at some length,
and after rejecting the possibiHties of these being
of (1) chromatic, (2) nucleolar, (3) chromidial, and
(4) mitochondrial origin likewise concludes that
they are transitory by-products. In this way
the keimbahn-determinants in copepods are satis-
factorily explained, and a similar explanation may
be applied to Sagitta, although with less certainty.

c. Discussion. A review of the literature on the
keimbahn-determinants and the investigation of these
substances in the eggs of insects force me to conclude
that the fundamental organization of the egg is respon-
sible for the segregation of the primordial germ cells,
whereas the visible substances simply furnish evi-

KEIMBAHN-DETERMINANTS 229

dence of this underlying organization. As I have
stated elsewhere (Hegner, 1908, p. 21) regarding Ihe
keimbahn-determinants in beetles' eggs, "the
granules of the pole-disc are therefore either the germ-
cell determinants or the visible sign of the germ-cell
determinants." The writer's experiments have thus
far failed to determine the exact function of these
granules. When the posterior end of a freshly laid
beetle's egg is pricked with a needle, not only the
pole-disc granules flow out, but also the cytoplasm
in which they are embedded (Hegner, 1908). If a
small region at the posterior end is killed with a hot
needle, the pole-disc is prevented from taking part
in the development of the egg, but so also is the sur-
rounding cytoplasm (Fig. 37, c). Eggs thus treated
continue to develop and produce embryos without
germ cells, but as a rule a part of the posterior end
of the abdomen is also absent (Hegner, 1911a). The
pole-disc granules and the cytoplasm containing
them is moved by centrifugal force toward the heavy
end of the egg and is proved to be quite rigid, but
eggs thus treated do not develop sufficiently normally
to enable one to decide whether the pole-disc pro-
duces germ cells in its new environment or not.

That the germ cells of Chironomus arise from a pre-
locaHzed substance was stated by Balbiani (1885) in
these words, "the genital glands of the two sexes
have an absolutely identical origin, arising from
the same substance and at the same region of
the egg'' Ritter (1890) expressed the opinion
that the "Keimwulst" of Chironomus consists of fine

230 GERM-CELL CYCLE IN ANIMALS

granulated protoplasm, an opinion concurred in by
Hasper (1911), who terms it "Keimbahnplasma."
The similar material in Miastor metraloas, the
*'polares Plasma," is considered a special sort of
protoplasm by Kahle (1908), and I can confirm this
for Miastor americana. Further evidence of the
protoplasmic nature of the substances which be-
come segregated in the primordial germ cells is fur-
nished by Boveri's experiments on Ascaris. In
1904 this investigator concluded from a study of
dispermic Qggs that the diminution process is con-
trolled by the cytoplasm and not by an intrinsic prop-
erty of the chromosomes, and that the chromosomes
of nuclei lying in the vegetative cytoplasm remain
intact, whereas those of nuclei embedded in the
animal cytoplasm undergo diminution. This con-
clusion has been strengthened by more recent experi-
mental evidence (Boveri, 1910) both from observa-
tion on the development of dispermic eggs and
from a study of centrifuged eggs (see p. 178, Fig.
53). Boveri's results furnish a remarkable confirma-
tion of the conclusions reached by the writer from a
morphological study of the germ cells of chrysomelid
beetles and expressed in the following words: "All
the cleavage nuclei in the eggs of the above-named
beetles {Calligrapha multipunctata, etc.) are poten-
tially alike until in their migration toward the periph-
ery they reach the 'keimhautblastem.' Then those
which chance to encounter the granules of the pole-
disc are differentiated by their environment, i.e., the
granules, into germ cells; all the other cleavage

KEIMBAHN- DETERMINANTS 231

products become somatic cells." Here, however,
the pole-disc granules were considered the essential
substance.

The appearance of the keimbahn-determinants at a
certain time and in a certain place, and their deter-
minate segregation, point unmistakably to an under-
lying regulating mechanism. These phenomena have
some definite relation to the fundamental organiza-
tion of the egg and require an investigation of our
present knowledge of this subject.

The isotropism of the egg as postulated by Pfluger
and the "cell interaction" idea especially developed
by O. Hertwig and Driesch have given way before
the beautiful researches tending to uphold the hy-
pothesis of "germinal localization" proposed by His
and championed by so many investigators within
the past two decades. The starting point for embry-
ological studies has shifted from the germ layers
to the cleavage cells and from these to the undivided
egg. Organization, which Whitman (1893) main-
tains precedes cell-formation and regulates it, is now
traced back to very early stages in the germ-cell
cycle and held responsible for the cytoplasmic lo-
calization in the egg.

One of the fundamental characteristics of the egg is
its polarity. It has been known for about thirty
years that the eggs of insects are definitely ori-
ented within the ovaries of the adults. Moreover,
gravity and the action of centrifugal force have no
effect upon the polarity of insect eggs (Hegner, 1909^) .
Giardina (1901) has found that during the divisions

232 GERM-CELL CYCLE IN ANIMALS

of the oogonia in Dytiscus a rosette of sixteen cells
is produced of which one is the oocyte and the other
fifteen nurse cells. The rosette thus formed possesses
a definite polarity coincident with the axis of the
oocyte which is identical with that which was present
in the last generation of oogonia. Similarly in
Miastor (Fig. 12) the polarity of the oocyte is recog-
nizable as soon as the mesodermal cells, which serve in
this species as nurse cells, become associated with it.

The germ cells of other animals also possess a
precocious polarity, as evidenced by their implanta-
tion in the germinal epithelium {e.g., Wilson, 1903 ;
Zeleny, 1904, in Cerebratulus), the position of the
nucleus, the formation of the micropyle (Jenkinson,
1911), etc. This is true not only for the inverte-
brates, but, as Bartelmez (1912) claims, "the polar
axis persists unmodified from generation to genera-
tion in the vertebrates and is one of the fundamental
features of the organization of the protoplasm" (p.
310). Furthermore, experiments with centrifugal
force seem to prove that the chief axis of the egg is not
altered when substances are shifted about, but is
fixed at all stages (Lillie, 1909; Morgan, 1909;
Conklin, 1910). Bilaterality also is demonstrable
in the early stages of the germ cells of many animals,
and, like polarity, seems to be a fundamental charac-
teristic of the protoplasm.

It is somewhat difficult to harmonize the various
results that have been obtained, especially by experi-
mental methods, from the study of egg organization.
As the oocytes grow, the apparently homogeneous

KEIMBAHN-DETERMINANTS 233

contents become visibly different in some animals,
and when the mature eggs develop normally these
"organ-forming substances" are segregated in def-
inite cleavage cells and finally become associated
with definite organs of the larva.

Conklin (1905) has shown "that at least five of
the substances which are present in the egg (of
Cynthia) at the close of the first cleavage, viz.,
ectoplasm, endoplasm, myoplasm, chymoplasm,
and chordaneuroplasm, are organ-forming sub-
stances." Under experimental conditions "they
develop, if they develop at all, into the organs which
they would normally produce; and, conversely,
embryos which lack these substances, lack also the
organs which would form from them." "Three of
these substances are clearly distinguishable in the
ovarian egg and I do not doubt that even at this
stage they are differentiated for particular ends"
(p. 220). "The development of ascidians is a mosaic
work because there are definitely localized organ-
forming substances in the egg; in fact, the mosaic
is one of organ-forming substances rather than of
cleavage cells. The study of ctenophores, nemer-
tines, annelids, mollusks, ascidians, and amphibians
(the frog) shows that the same is probably true of all
these forms and it suggests that the mosaic principle
may apply to all animals" (p. 221). The same
writer has also proved from his study on Phallusia
(1911) that these various substances exist even when
they are not visible in the living egg. It is interesting
also to note that Duesberg (1913) finds the "niyo-

234 GERM-CELL CYCLE IN ANIMALS

plasm" of Cynthia to be crowded with plasmosomes,
differing in this respect from other egg regions.

Experiments, especially those of Lillie (1906, 1909),
Morgan and Spooner (1909), Morgan (1909a), and
Conklin (1910), have shown that in many eggs the
shifting of the supposed organ-forming substances
has no influence upon development, and leads to the
conclusion that these visible substances play no
fundamental role in differentiation, but that the
invisible ground substance is responsible for de-
terminate development. The eggs of different ani-
mals, however, differ both in time and degree of
organization, and the conflicting results may be
accounted for by the fact that specification is more
precocious in some than in others.

The most plausible conclusions from a considera-
tion of these observations and experiments are that
every one of the eggs in which keimbahn-determi-
nants have been described consists essentially of a
fundamental ground substance which determines
the orientation ; that the time of appearance of
keimbahn-determinants depends upon the preco-
ciousness of the egg ; that the keimbahn-determi-
nants are the visible evidences of differentiation in
the cytoplasm ; and that these differentiated portions
of the cytoplasm are definitely localized by cytoplas-
mic movements, especially at about the time of
maturation.

KEIMBAHN-DETERMIXAXTS 235

B. The Localization of the Keimbahn-deter-

MINANTS

One of the characteristics of the keinibahn-
determinants is their regular appearance at a certain
stage in the germ-cell cycle according to the species in
which they occur, and their constant localization
in a definite part of the egg, or in one or more definite
cleavage cells. Keimbahn-determinants are recog-
nizable in many insects' eggs before fertilization is
accomplished, and even before the oocyte has reached
its maximum size. We know that in Chironomus
the ''Keimwulst" (Ritter, 1890) or ''Keimbahn-
plasma" (Hasper, 1911) is present when the egg
is laid, at which time the pronuclei as a rule have
not yet fused. This is true also of the "Dotter-
platte" in CaUiphora (Noack, 1901). There can
be little doubt, however, that these substances
are present as such in the eggs before fertilization,
judging from our knowledge of the history of similar
materials in the eggs of other insects. The "pole-
disc" in the eggs of chrysomelid beetles (Hegner,
1908; Wieman, 1910a) and the *'polares Plasma"
in Miastor (Kahle, 1908; Hegner, 1912, 19Ua) are
recognizable some time before fertilization and cannot
therefore arise because of any influence exerted by
the spermatozoon. Moreover, in Miastor the eggs
thus far examined have all been part hen ogenetic.
In parasitic Hymenoptera the Keimbahn-chromatin
appears in both fertilized and parthenogenelic eggs
at an early growth period. In only one animal not

236 GERM-CELL CYCLE IN ANIMALS

an insect has a similar occurrence been noted, namely,
in Polyphemus, where, according to Kuhn (1911,
1913), the keimbahn-determinants consist of the
remains of one or more nurse cells (Fig. 50). In the
Daphnid^ (Weismann and Ischikawa, 1889) the
"Paracopulationszelle" arises from material cast out
by the germinal vesicle ; in Mquora (Haecker, 1892)
the *'Metanucleolus" is likewise derived from the
germinal vesicle ; in Ascaris (Boveri, 1892) chroma-
tin-diminution occurs during the two- to four-cell
stage; in Cyclops (Haecker, 1897, 1903) and other
copepods (Amma, 1911) the " Aussenkornchen " or
*'Ectosomen" become visible soon after fertiliza-
tion (Diaptomus) , but usually not until the pro-
nuclei fuse (other species) ; in Sagitta the "be-
sondere Korper" (Elpatiewsky, 1909, 1910) or
*'Keimbahnchromidien" (Buchner, 1910) appear to
arise de novo after fertilization, although if Buchner's
contention that they are the remains of the accessory
fertilization cells is correct, they should be classed
with the " Nahrzellenkern " described by Kuhn
(1911, 1913) in Polyphemus.

It is thus evident that the keimbahn-determinants
become visible, wherever they have been described,
either just before or just after the eggs are fertilized,
or, in parthenogenetic forms, shortly before matura-
tion and cleavage are inaugurated.

The localization of the keimbahn-determinants at
the time of their appearance seems to be predeter-
mined. In insects the posterior end of the egg is
invariably the place where these bodies occur. In

KEIMBAHN-DETERMIXAXTS 237

species whose eggs undergo total cleavage they are,
under normal conditions, segregated in one definite
blastomere from the two-cell stage up to the thirty-
two-cell stage, as a rule, and are then distril)uted
among the descendants of the single primordial
germ cell. In Ascaris it is normally the cell at the
posterior (vegetative) pole that fails to undergo
the diminution process. It seems therefore that
there must be some mechanism in the Qgg which
definitely localizes the keimbahn-determinants.

The segregation of these substances in one blas-
tomere at the first cleavage division is a result of their
previous localization, but in later cleavage stages
events are more diflficult to interpret. Both Haecker
(1897) and Amma (1911) have attempted to explain
the distribution of the "Ectosomen" in copepods by
postulating a dissimilar infiuence of the centrosomes
resulting in the segregation of these granules at one
end of the mitotic spindle in the dividing stem cell.
According to Zeigler's hypothesis the centrosomes
during unequal cell divison are heterodynamic,
and Schonfeld (1901) believes that the synizesis
stage is due to the attraction of the chromosomes by
the centrosomes. It is well known that in many
cases where unequal cell division occurs one aster
is larger than the other, and this may be the true
interpretation of the phenomena, but to the writer
it seems more probable that the entire cell contents
undergo rearrangement after each cell division,
possibly under the influence of the material elab-
orated within the nucleus and set free during iiiito-

238 GERM-CELL CYCLE IN ANIMALS

sis. Elpatiewsky (1909) also believes in the unequal
attractive force of the centrosomes in Sagitta}

In Ascaris, certain copepods, Sagitta, Polyphemus^
and certain Daphnid^ the keimbahn-determinants
are segregated in one cleavage cell until about the
thirty-two-cell stage, but their substance is dis-
tributed at the next division between the daughter
cells. The insects such as Chironomus, Miastor, and
chrysomelid beetles, where, on account of the super-
ficial cleavage the keimbahn-determinants are not
segregated in blastomeres, the primordial germ
cells from the beginning consist almost entirely
of the keimbahn material or this material plus
the matrix in which it is embedded. Hence in
these cases the keimbahn-determinants are localized
at a determined point during each cleavage stage
instead of being carried about by the movements of
the egg contents or of the blastomeres, but, as in
the eggs that undergo total cleavage, the determi-
nants are distributed between the daughter cells as

^"Nach der vierten Teilung kommt der besondere Korper in den
Wirkungskreis eines Zentrosomos, namlich desjenigen, welcher naher
der Polarfurche liegt. Fast die ganze 'Energie' dieses Zentrosomas wird
fiir die Ueberwindung der vis inertiae des besonderen Korpers ver-
braucht ; dieser wird dera Zentrosoma genahert und umschliesst es wie
mit einer Kappe, so dass er im optischen Durchschnitt stets Hufeisen oder
Sichelform aufweist. Infolge davon wird die wirkung dieses Zentroso-
mas auf das Zellplasma nur sehr schwach, dieses Zentrosoma kann nur
einen kleinen Plasmateil beherrschen, und die resultierende Zelle wird
viel kleiner, als die Schwesterzelle. Diese kleine Zelle, die den beson-
deren Korper bekommen hat, liegt naher zum vegetativen Poles, als
die grossere Schwesterzelle, und stellt die erste Urgeschlechtszelle
G(d^^^), die grossere Schwesterzelle die erste Urentodermzelle E(d^^2)
vor" (p. 231).

KEIMBAHN-DETERxMINANTS r,i9

soon as the primordial germ cells are esta})lis}ie(l.
The reason for this appears to be that localizations
occur in holoblastic eggs at each cleavage and that
not until the thirty-two-cell stage or thereabouts
does the keimbahn material become entirely sep-
arated from other organ-forming substances and
segregated in a single cell. When this point is
finally reached, this keimbahn material must neces-
sarily become divided between the daughter cells.

In practically all known cases the daughter cells
of the primordial germ cells are equal in size and each
receives an equal portion of the keimbahn-de-
terminants (Fig. 37, B). This is certainly to be
expected from their constitution and future historv.
Sagitta, however, differs in this respect, for the remains
of the "besondere Korper" appear to be unequally
distributed between the two daughter cells of the
primordial germ cells (Fig. 54) and both Elpatiewsky
(1909, 1910) and Stevens (1910), therefore, consider
this as probably a differential division whereby in this
hermaphroditic animal the substance of the male
primordial germ cell is separated from the female.
More work is necessary to make certain of this point.

Conclusion. Keimbahn-determinants are def-
initely localized in the egg and in definite cleavage
cells. This localization is first observable just
before or just after the eggs are fertilized, or, in
parthenogenetic forms, shortly before maturation
and cleavage are inaugurated. Some mechanisui in
the egg must be responsible for this localization.
Heterodynamic centrosomes may have some influence

240 GERM-CELL CYCLE IN ANIMALS

so far as the segregation of the keimbahn-determi-
nants in cleavage cells is concerned, but the move-
ment of the egg contents seems to be a more probable
cause of localization.

C, The Fate of the Keimbahn-determinants

It is unfortunately impossible to trace the keim-
bahn-determinants throughout the entire germ-cell
cycle. The question of their fate, however, is an
important one. As we have seen, they become vis-
ibly apparent shortly before or just after the inaugu-
ration of the maturation divisions, and remain intact
for a brief period during the early cleavage stages.
They persist in insects as definitely recognizable
granules (Fig. 37, F) for some time after the primor-
dial germ cells are segregated ; then they gradually
break up into finer particles, leaving no trace of their
existence behind except in so far as they give the
cytoplasm of the germ cells a greater afiinity for
certain dyes. In Chironomus they may still form
distinct masses after the definitive germ glands
have been formed (Fig. 33, D). The ectosomes in
the copepods are temporary bodies which appear
to rise de novo during the formation of each mitotic
figure in the early cleavage stages, then break down
and disappear. Practically all of the other keim-
bahn-determinants persist during early cleavage and
then disappear as distinct visible bodies as soon as the
primordial germ cells are definitely segregated. What
becomes of them during the comparatively long
period between their disappearance in the primordial

KEIMBAHN-DETERMINAXTS ^241

germ cells and their reappearance in the oocytes or
mature eggs can only be conjectured. They seem to
disintegrate into very fine particles which become
thoroughly scattered within the cell body and mixed
with the cytoplasm. It has been suggested (p. 68)
that they may retain their physiological characteris-
tics and become concentrated again in the growing
oocytes into morphologically similar bodies, in-
creasing in the meantime, by multiplication or in
some other way, until they equal in mass those of
the preceding generation of germ cells. On the other
hand, they may all, like the ectosomes of copepods,
be temporary structures produced at a certain time
and place under similar metabolic conditions, and,
becoming associated with particular parts of the
cell contents, thus be constant in their distribution.
Several ideas have been advanced regarding the
fate of the eliminated chromatin in Ascaris. The
ends of the chromosomes which are cast out into the
cytoplasm are not equally distributed among the
daughter cells nor does there appear to be any mech-
anism for their definite unequal division. These
facts argue against the theory that these cast -out
chromatin bodies serve as determinants and also
make improbable the hypothesis that they enal)le
the somatic cells to differentiate, whereas the germ
cells which do not undergo the diminution process
remain in an indifferent condition, since their cyto-
plasm lacks this material (Montgomery, 1911, p. 7\H).
However, the fact that during the early cleavage
divisions in some animals (see p. 218) large amounts

242 GERM-CELL CYCLE IN ANIMALS

of chromatin escape from the nucleus and are dif-
ferentially distributed to the daughter cells is evidence
that nuclear material may play some important role
in the progressive changes of cleavage cells.

It has been shown that in many animals the germ
cells do not multiply for a considerable period
during the early developmental stages. This period
coincides also with that during which the keimbahn-
determinants, as a rule, disappear. For example,
the germ cells of chrysomelid beetles multiply until
there are about sixty -four present, at which time they
constitute a group at the posterior end of the egg and
the embryo has just started to form ; no further
increase in number occurs until the larval stage is
reached and the definitive germ glands are established.
As soon, however, as the embryo has reached a
certain developmental stage, the germ cells migrate
into it, and it looks very much as though they remain
quiescent until the somatic cells are *'able to protect,
nourish, and transport" them.

The number of primordial germ cells during the
**period of rest" is perhaps most definitely known in
Miastor, where, as one group of eight and later as two
groups of four each, they are present throughout a
large part of embryonic development.

In vertebrates also a long period exists during
which division of the primordial germ cells does not
take place (Fig. 6) and at least in several species
certain cell contents (the mitochondria) remain in an
indifferent condition (Rubaschkin, 1910; Tschasch-
kin, 1910 ; Fig. 31, B). These facts all indicate that

KEIMBAHN-DETERMIXANTS 243

these cells remain in a primitive condition and do
not undergo the histological differentiations charac-
teristic of somatic cells, a view which, however, has
been objected to (Eigenmann, 189G). The disap-
pearance of the keimbahn-determinants and the
yolk globules of vertebrates during this period have
suggested that these substances are nutritive in
function, furnishing energy to the migrating germ
cells.

The fact of this long rest period, followed by rapid
multiplication of the oogonia and spermatogonia
during which no important specializations occur, and
later succeeded by the remarkable changes that occur
in both the oocytes and spermatocytes, has led to the
suggestion (Montgomery, 1911, pp. 790-792) that in
the germ-cell cycle there is a series of changes
parallel with that of the somatic cycle. In the
development of both cycles preformation and epi-
genesis proceed at the same time. The chromosomes
seem to be the preformed elements of the germ cells,
since they are apparently the most stable constitu-
ents. The cytoplasm, on the other hand, undergoes
a series of epigenetic changes such as the formation
of an idiozome, the development of mitochondria,
the appearance of a sphere, and the metamorphosis
of the spermatozoon.

Finally we must inquire into the fate of the keim-
bahn-determinants in the male germ cells. Does the
keimbahn material in these cells increase in amount as
has been suggested for the oocytes and is it localized
in the spermatogonia, spermatocytes, or spernuitozoa

244 GERM-CELL CYCLE IN ANIMALS

as a definite, visible substance ? We know from the
investigations of Meves (1911) that the plastosonies
in the spermatozoon are carried into the egg, in the
case of Ascaris, and there fuse with the plastosomes
of the ovum. Whether keimbahn-determinants act
in a similar manner is unknown. There are, how-
ever, certain cytoplasmic inclusions in the male
germ cells that have been compared with similar
structures in the oocytes, for example, the chromatic
body described by Buchner (1909) in the spermato-
genesis of Gryllus (see p. 88), and the plasmosome
which is cast out of the nucleus of the second sperma-
togonia in Periplatieta and disintegrates in the cy-
toplasm (Morse, 1909). That keimbahn-determi-
nants from the spermatozoon are not necessary for
the normal production of germ cells is of course evi-
dent, since some of the species with which we are
best acquainted, for example, Miastor, are partheno-
genetic.

CHAPTER IX

THE CHROMOSOMES AND MITOCHONDRIA OF

GERM CELLS

No account of the germ-cell cycle in animals can be
considered complete without at least a brief reference
to the history of the chromosomes and mitochondria
of germ cells. The chromosomes have for many
years been recognized as the most important visible
bodies in the cell, and their behavior during the germ-
cell cycle has convinced most zoologists that they
may also be regarded as the bearers of hereditary
factors. The mitochondria, on the other hand,
are cellular constituents which have only compara-
tively recently come into prominence in cytological
literature, and ideas concerning their nature and
functions are still in a very chaotic condition.

The Chromosome Cycle in Animals

A few general statements regarding the behavior of
the chromosomes during cell division, maturation,
and fertilization are contained in Chapters I nnd II.
We may recognize a rather definite chromosome cyc-le
as a part of the germ-cell cycle, and it is to certain
events in this chromosome cycle that our attention
will be directed in the following i)aragraphs. It is
best to begin our discussion, as in the gencrnl review

245

246 GERM-CELL CYCLE IN ANIMALS

of the germ-cell cycle (Chapter II), with the par-
thenogenetic or fertilized egg after the maturation
processes have been completed, and to exclude all
references to the accessory chromosome until later.

It may be pointed out first that the number of
chromosomes in the cells of any individual of a
species is, with few exceptions, constant. Thus the
thread worm of the horse, Ascaris megalocephala
var. Mnivalens, has two; A. megalocephala var.
hivalens, four ; the nematod, Coronilla, eight ; the
mole cricket, Gryllotalpa vulgaris, twelve; the
bug, Pentatoma, fourteen ; the rat, sixteen ; the
sea urchin. Echinus, eighteen ; the salamander,
Salamandra maculosa, twenty-four ; the slug, Limax
agrestis, thirty-two ; and the brine shrimp, Artemia,
one hundred and sixty-eight. This number, however,
is reduced one-half during the maturation of the
eggs and spermatozoa so that the mature eggs and
spermatozoa possess only half as many chromosomes
as the other cells in the body ; for example, the body
cells, oogonia, and spermatogonia of the rat are
provided each with sixteen chromosomes, but the
mature eggs and spermatozoa contain only eight.
Parthenogenetic eggs differ from those that require
fertilization, since in these the complete or diploid
number of chromosomes is retained. When cleavage
is inaugurated in such eggs, a spindle is formed, the
chromosomes are halved, and each daughter cell
acquires one-half of each chromosome as in ordinary
mitosis. In fertilized eggs, however, the nucleus
brought in by the spermatozoon fuses more or less

CHROMOSOMES AND MITOCHONDRIA 247

completely with the egg nucleus and the two to-
gether become incorporated in the first cleavage

Fig. 61. — Independence of paternal and maternal chromatin in the
segmenting eggs of Cyclops. A. First cleavage-figure in ('. .strcmtus;
complete independence of paternal and maternal chromosonu-s.
B. Resulting two-cell stage with double nuclei. C. Second cleavage ;
chromosomes still in double groups. D. Blastomeres with double
nuclei from the eight-cell stage of C. brevicornis. (From Wilson,
after Haecker.)

spindle. Each of the two nuclei furnishes an equal
(haploid) number of chromosomes to the first

248 GERM-CELL CYCLE IN ANIMALS

cleavage spindle, and thus the diploid (somatic)
number is regained. These chromosomes may there-
fore be considered as forming two groups, one group
of paternal origin derived from the sperm nucleus,
and one group of maternal origin derived from the
egg nucleus ; in fact the groups supplied by the two
nuclei may remain perfectly distinct (Fig. 61), not
only during the first cleavage division, but also
during subsequent mitoses.

The chromosomes of the fertilized egg and of
the cells to which it gives rise are not always of the
same size and shape, but in many cases are known to
differ morphologically from one another. It is
possible to recognize the different chromosomes
during each mitosis, and the evidence is quite con-
vincing that morphologically similar pairs are present
in every cell and that one member of each pair is
derived from the egg nucleus, the other from the
sperm nucleus. Two principal views are held re-
garding the character of the chromosome divisions
during the early cleavage divisions, (1) that the
chromatin granules, which represent definite de-
terminers, are divided equally between the daughter
chromosomes, and (2) that an unequal distribution
of the granules occurs, thus forming daughter cells
containing qualitatively different chromosomes.
There are no observations which show an unequal dis-
tribution.

One of the changes that takes place in the chromo-
somes at the time of mitosis is the diminution of their
chromatin content brought about by the passage of

CHROMOSOMES AND MITOCIIOXDUIA 249

part of their substance into the cytoplasm. This
phenomenon has been used as an argument in
favor of the theory of nuclear control of cellular
activities. Two special cases of chromatin-diniinu-
tion are known which differ from the usual process ;
these occur in Ascaris and Miastor as described and
figured in Chapters III and VI. In these animals a
large portion of the chromosomes of certain nuclei
is cast out into the cytoplasm, w^hereas all of the
chromatin is retained by others; the latter witli a
complete amount become the nuclei of the germ cells,
the rest with a reduced amount are present in all of the
somatic cells.

During the cellular divisions which result in the
multiplication of the somatic cells and of the ])rimor-
dial germ cells the chromosomes appear at each
mitosis in their normal number and are apparently
divided equally between the daughter cells. There
are, however, certain variations in both the somatic
and germinal mitoses. In the somatic cells only
one-half the normal number may appear; thus in
the snail, Helix pomatia, the number may be twenty-
four instead of the usual forty-eight. T]um'(^ is
reason to believe that each of these twenty-four really
consists of two single (univalent) chromosomes,
and may therefore be considered bivalent. Even
a further reduction in number by the association of
univalent chromosomes has been recorded, in which
case the combined chromosomes are said to be pluri-
valent. Other variations in the number of chronio-
somes, which occur during the maturation of the
germ cells, will be referred to later.

250 GERM-CELL CYCLE IN ANIMALS

Certain cellular phenomena which concern the
chromosome cycle have been described in preceding
chapters and so need only be mentioned here. First,
the occurrence of amitosis in the multiplication of
the germ cells has an intimate relation to the speci-
ficity of the chromosomes, since if nuclei divide en
masse it seems improbable that the chromosomes be-
come equally divided between the daughter nuclei
(see Chapter V, p. 133) ; and second, the formation of
nurse cells from oogonia may be accompanied, as
in Dytiscus (Chapter V, p. 120), by a chromatin-
diminution process which may be regarded as a
differentiation of mother germ cells into somatic
cells (nurse cells) and oogonia, a differentiation re-
sembling the segregation of the primordial germ
cells in the cleavage stages of the ^gg.

The most striking and perhaps the most important
stages in the chromosome cycle occur during the
growth and maturation periods of the germ cells.
As briefly described and figured in Chapter II,
the mitoses which occur during maturation are
meiotic, since the mature germ cells have their chro-
mosome number reduced one-half. The events in this
process most worthy of our attention are those which
take place during the stages known as synapsis
and reduction. Wilson (1912) has summed up the
questions that remain to be solved in the following
words: "The cytological problem of synapsis and
reduction involves four principal questions, as
follows : (1) Is synapsis a fact ? Do the chromatin-
elements actually conjugate or otherwise become

CHROMOSOMES AND MITOCHONDRIA 251

associated two by two ? (2) Admitting the fact of
synapsis, are the conjugating elements chromosomes,
and are they individually identical with those of
the last diploid or pre-meiotic division ? (3) Do
they conjugate side by side (parasynapsis, parasyn-
desis), end to end (telosynapsis, metasyndesis),
or in both ways ? (4) Does synapsis lead to partial
or complete fusion of the conjugating elements to
form 'zygosomes' or 'mixochromosomes,' or are
they subsequently disjoined by a 'reduction-divi-
sion ' ? Upon these questions depends our answer
to a fifth and still more important question, namely,
(5) Can the Mendelian segregation of unit-factors
be explained by the phenomena of synapsis and
reduction ?"

The behavior of the chromosomes during synapsis
in the germ cells of the male is indicated diagram-
matically in Fig. 62, the terms used being those
proposed by von Winiwarter (1901) in his work on
the oogenesis of the rabbit. In the spermatogonia
(Fig. 62, 1) the chromatin is arranged in clumps on
an achromatic reticulum ; in the spermatocyte
(Fig. 62, 2) it breaks up into granules which become
arranged in single rows or filaments (the leptotene
threads). These leptotene threads later become
paired (synaptene stage. Fig. 62, 3) and converge
toward the side of the nucleus near which the centro-
some and centrosphere are situated (Fig. 62, 4)» a
condition known as synizesis. The granules of the
leptotene filaments approach and finally fuse so
as to produce single thick threads (Fig. 62, 5-7) ;

252

GERM-CELL CYCLE IN ANIMALS

this is the pachytene stage. The filaments then
begin to unravel (Fig. 62, 6-7), become distributed

Fig. 62. — Prophases of the heterotype division in the male Axolotl.
1, nucleus of sperm ogonium, or young spermocyte ; 2, early lepto-
tene ; 3, transition to synaptene; 4, synaptene with the double
filaments converging towards the centrosome ; 5, contraction fig-
ure; 6, 7, pachytene; 8, early; 9, later diplotene ; 10, the hetero-
typic double chromosomes; the nuclear membrane is disappearing.
(Frorti Jenkinson, 1913.)

throughout the nucleus, and finally split into two
threads (Fig. 62, 8-9) ; this is the diplotene stage.
The pairs of filaments finally shorten and thicken,

CHROMOSOMES AND MITOCHONDRIA 253

assuming the form of paired chromosomes of various
shapes and sizes (Fig. 62, 10). A spindle then
forms; these *' heterotypic" chromosomes are
drawn upon it ; and each daughter cell receives one
chromosome of each pair.

This mitosis is called heterotypic because it differs
from ordinary indirect nuclear division in two im-
portant respects : (1) the chromosomes are ])resent
in pairs, and entire chromosomes are separated, and
(2) the result is a reduction of chromosomes in the
daughter nuclei to one-half the somatic number.
According to certain investigators {e.g., jNIeves,
1907) the union of the leptotene threads in the synap-
tene stage (Fig. 62, ^) does not occur, but the two
parallel threads are simply the halves of a single
longitudinally split filament which fuse in the
pachytene stage (Fig. 62, 6-7), and separate again
in the diplotene stage (Fig. 62, 8-9). The large
majority of cytologists, however, believe that the
leptotene threads represent chromosomes which
actually fuse in pairs in the pachytene stage and sep-
arate from each other during the heterotypic mitosis.
Furthermore, the chromosomes of each pair are con-
sidered to be homologous, that is, the one derived
from the spermatozoon is morphologically similar, to
its mate, which is derived from the egg nucleus.

Investigators who believe synapsis to be a fact,
that the conjugating elements are chromosomes,
and these chromosomes are identical with those of
the last diploid mitosis are not agreed as to the
method of union and subsequent separation of the

254 GERM-CELL CYCLE IN ANIMALS

chromosomes. The chromosom'es may unite side by
side in parasynapsis or end to end in telosynapsis.
Apparently parasynapsis is the rule, although telosyn-
apsis probably occurs in certain species. The results
are the same in either case.

The next question to be considered is whether the
chromosomes which emerge from the pachytene stage
are the same as those that enter it as leptotene
filaments, or whether there is a complete fusion
into zygosomes or mixochromosomes. It seems
probable that at least a partial fusion occurs
and that the composition of the chromosomes is
changed more or less during synapsis. We know
for certain that the peculiar X-chromosomes which
have been found in many species of animals become
paired in synapsis and later separate in a true
reduction division, and we also have evidence
which furnishes a mechanical means of effecting
a change in the chromosomes during the synaptene
stage. This evidence has led to the formulation of
the chiasmatype theory (Janssens, 1909). Accord-
ing to this theory the chromosomes which pair in
synapsis may twist around each other more or less
(Fig. 63), and cross connections are visible. When
the paired chromosomes later split apart they rep-
resent combinations different from those present
before synapsis, because of these cross connections.
The results of experimental breeding seem to necessi-
tate some such relation as this during synapsis, and
the chiasmatype theory has been used to explain
certain results of hybridization that have not been

CHROMOSOMES AND MITO( HOXDRIA 255

accounted for in any other way (Morgan, 1013,
1914).

The view that the chromosomes are the ])earers of
factors in heredity is based upon several liypotheses,
of which those of their specificity and genetic con-
tinuity will be mentioned here. According to the
hypothesis of chromosome specificity each chromo-
some possesses certain functions of a specific kind

Fig. 63. — Twisting of chromosomes according to the chiasmat>T)e
theory. A. Two twisted chromosomes each divided longitudinally
into two. B. Twisted chromosomes of Batracoseps attenuatus.
(From Janssens, 1909.)

which determine the character of cellular differen-
tiation and thus the structural and physiological
condition of the embryo, larva, and adult. The hy-
pothesis of the genetic continuity was evolved from
that of the individuality of the chromosomes. Ac-
cording to the latter theory the chromosomes that
appear in mitosis do not become scattered during the
resting stage of the nucleus (interkinesis), but re-
tain their identity throughout this period. Lack
of evidence has resulted in the substitution of the
hypothesis of genetic continuity, according to
which there is a definite relation between the chro-
mosomes of successive mitotic divisions.

Much of the cytological literature of the past dec-
ade deals with the history of the X- or sex-chromo-

25Q GERM-CELL CYCLE IN ANIIVMLS

somes. For many years the number of chromosomes in
the cells of the individuals of a species was considered
constant and even. Henking, however, in 1891,
discovered in the bug, Pyrrhocoris, sl single chro-
mosome which did not divide in one of the sperma-
tocyte divisions, but passed to one of the daughter
cells and hence into only one-half of the spermatozoa.
Paulmier (1899) observed similar conditions in
the squash bug, and since then one or more odd
chromosomes have been discovered in a large number
of animals belonging to many different phyla. In
1902, McClung suggested that these pecuHar chro-
mosomes might be sex-determinants, and subsequent
discoveries have fully demonstrated that they are
intimately associated with the phenomena of sex.
Most of our knowledge of this subject is due to the
investigations of cytologists in this country, es-
pecially Montgomery (1898, 1906, 1911), McClung
(1899, 1902, 1905), Stevens (1905, 1906, 1910),
Wilson (1905, 1906, 1911, 1912), and Morgan (1909,
1911, 1913, 1914). A few of the principal types of
sex-chromosome distribution are as follows :

Type I. One X-chromosome. This, the simplest
type, has been recently demonstrated in a remarkable
fashion by Mulsow (1913) in a nematoid worm,
Ancyr acanthus. Here the chromosomes can be seen
not only in stained material but also in the living
germ cells. The diploid number of chromosomes in
male worms is eleven (Fig. 64, A), in female worms,
twelve {E) . Two sorts of spermatozoa are produced,
one-half with five and the other half with six chromo-

Fig. 64. — Behavior of chromosomes during inaturatioii, fertiliziition.
and cleavage of Ancyr acanthus cystidicola. {From Mulsoir, IIJIS.)
A. Spermatogonium with eleven chromosomes. B. First matura-
tion (spermatocyte) division. The single chromosome finally joins
one group. C. The four spermatids arising from one spcrniatooyte ;
two with six chromosomes, and two with five. D. Two sperma-
tozoa drawn while alive ; one with six chromosomes, and one with
five. E. Oogonium with twelve chromosomes. F. Second matura-
tion (oocyte) division. The black mass above is the first polar
body; the set of six black chromosomes are those of the second
polar body; the six dotted chromosomes are those of the egg.
G. Fertilized (male producing) egg ; sperm nucleus above with five
chromosomes; egg nucleus below with six chromosomes. //. fertil-
ized (female producing) egg; both egg and sperm nuclei with .six
chromosomes. /. Cleavage stage of male producing egg : the central
cell with nucleus containing eleven chromosomes. J. Cleavage

Prot^nor <?

Pro/eno/

t'€

Fig. 65. — Maturation in Pro^enor. Male above. A. Spermatogonium.
B. Synapsis. C. First maturation division. Z), D'. Second mat-
uration division. E, E' . Two sorts of spermatozoa.

FemaU below. A. Oogonium. B. Synapsis. C. First matura-
tion division. D. Second maturation division. E. Egg nucleus
and two polar bodies all alike in chromosome content. First polar
body is dividing. {From Morgan's Heredity and Sex, published by
the Columbia University Press.) (258)

CHROMOSOMES AND MITOCHONDRIA 259

somes (Fig. 64, D). The nuclei of all the inalure
eggs exhibit six chromosomes. When fertilized
the spermatozoon nucleus can be recognized, since
it lies near the end away from the polar bodies. On
the average one-half of the eggs are fertilized by
spermatozoa containing five chromosomes and one-
half by spermatozoa containing six. The results
are as follows : A zygote resulting from the fusion
of an egg with six chromosomes and a spermatozoon
with six chromosomes possesses twelve chromosomes
and develops into a female (Fig. 64, //) ; and a
zygote formed by an egg with six chromosomes and
a spermatozoon with five chromosomes contains
eleven chromosomes, and hence gives rise to a nuile
(Fig. ,64, G). The events during the maturation
processes in such a case are similar to those in the
bug Protenor, as illustrated in Fig. Q5.

Type II. One X-chromosome and one Y-chro-
mosome. In the bug, Lygoeus bicrucis, and a num-
ber of other species the number of chromosomes in
both male and female is the same, but two sex-chro-
mosomes of different sizes are present in the male.
As shown in Fig. 66, the eggs are all alike, contain-
ing six ordinary and one X-chromosome. The sper-
matozoa are of two sorts: one-half with the larger,
or X-chromosome, the other one-half with the smaller,
called by Wilson the Y-chromosome. The zygotes,
consequently, produce males if one X-chromosome
and one Y-chromosome are present, and fenudes
if two X-chromosomes occur.

Type III. Two chromosomes of equal size

260

GERM-CELL CYCLE IN ANIMALS

i-yp'

<a.eits

•*:?%

• •••

Ly^aetts" ^

••

»!*^

Fig. 66. — Maturation in Lygoeus. Male above. Female below. Let-
tering as in Fig. 65. (From Morgan s Heredity and Sex, published
by the Columbia University Press.)

CHROMOSOMES AND MITOCHONDRIA 2^1

(Fig. 67). In the bug, Oncopelius fa.sriatus, the
number of chromosomes (10) in both male and
female is the same, but they are of ecjual size in
both sexes. It is probable, however, that one of
those of the male represents an X-chromosome and
the other a Y-chromosome as in Type II, although
they are not visibly different.

Type IV. One X-chromosome attached to an
ordinary chromosome. There are a number of
cases on record in which the X-chromosome is
attached to an ordinary chromosome as in Ascaris
megalocephala. Probably on this account the sex-
chromosome was overlooked in these species for
many years. The resulting zygotes, as Fig. 08
show^s, are comparable to those of Type I (Fig. 65).

Type V. Spermatozoa alike, but eggs of two sorts.
In a few animals it has been found that the eggs are
dimorphic and the spermatozoa all alike, as repre-
sented in Fig. 69.^

Numerous variations have been discovered in
the number and size of the X- and Y-chromosomes ;
some of these are illustrated in Fig. 70. When more
than one X-chromosome is present they act as a unit,
and two sorts of zygotes are produced as in other cases.

Chromosome cycles of more than ordinary interest
have been described in the honeybee, in phyloxerans
and aphids and in certain hermaphrodites. It
has long been known that the female honeybees
(queens and workers) develop from fertilized eggs

1 The recent contributions of Tennent iind Baltzcr make the occurrence
of this type seem very doubtful.

262

GERM-CELL CYCLE IN ANIMALS

OnccpettoAf 3

to*.*

*.o°.

fl .^^

•••

Wlii//-

Onccpettu^ 9

JT'

Fig. 67. — Maturation in Oncopeltus. Male above. Female below.
Lettering as in Fig. 65. (From Morgans Heredity and Sex, pub-
lished by the Columbia University Press.)

CHROMOSOMES AND MITOCHONDRIA 263

Escorts cT

«\}

fik

^sootri^ 9

//I

#«

Fig. 68. — Maturation in .4scans. Male al)Ove. Female below. Let-
tering as in Fig. 05. {From Moruan's Heredity and Sex, publmfud
by the Columbia University Press.)

264

GERM-CELL CYCLE IN ANIMALS

Reduction
Division

Ripe Eggs

Q»i P'^^f^r Body

Oogonium

^\y

1// <

Spermat-
ogonium

'-/V

s^/^

Fertilized

Egg

Fertilized

Egg

Fig. 69. — Diagrams showing the behavior of the chromosomes during
maturation and fertilization in the starfish, Echinus. One kind of
spermatozoon is formed, but the ripe eggs differ, one containing a
large X-element, the other a small Y-element. (From Schleip, 1913.)

Protenor, A^iasa Syromastes, Homo Ascaris lumhricoides

Nazara
viridula

Euschisfus
coemis

Nazara
hilaris

i i

Thy ant a
calceata

Rocconoia, Prionides, Gelasfocoris Acholla
Fitchia Sinea multispinosa

Fig. 70. — Diagram showing the number and size relations of the X- and
Y-chromosomes in a number of animals. {From Wilson, 1911.)

CHROMOSOMES AND MITOCHONDUIA ^2G5

and the drones parthenogenetically. The liistory of
the chromosomes has here been worked out by
Nachtsheim (1913). The primary oocyte contains
sixteen chromosomes in the form of eight tetrads ; the
mature egg and polar bodies are each provided witli
eight chromosomes (Fig. 71, E) ; the inner half of the
divided first polar body fuses with the second polar
body, forming a *'Richtungskopulationskern" (Fig.
71, F) which does not give rise to the male germ cells
as Petrunkewitsch (1901) claimed, but degenerates.

The cleavage nucleus in the parthenogenetic egg
which produces the male shows sixteen chromosomes
which divide to form thirty-two or sixty-four in
the somatic cells, but do not increase in number in
the spermatogonia. The first maturation division
is unequal, and a "polar body" without any chroma-
tin is pinched off (Fig. 71, A-C, Rh). The sperma-
tids are likewise of two sorts; the smaller (Fig. 71,
C, Rk2) contain as many chromosomes as the larger
(16), but degenerate, while the larger transform into
spermatozoa. The fertilized (female) eggs possess
the same number of chromosomes as the partheno-
genetic eggs, plus an equal number which is brought
in by the spermatozoon. The cleavage nucleus
exhibits thirty-two chromosomes wliich may become
sixty-four in the somatic cells, Init unite two by
two to form sixteen in the oogonia.

Phylloxera caryoBcaulis will serve to illustrate
the chromosome cycle in a species with a life cycle
composed of parthenogenetic females which alter-
nate with sexual males and females (Morgan, 1909,

266

GERM-CELL CYCLE IN ANIMALS

1910). The eggs laid by the stem-mother (see Chap-
ter I, p. 24) in the spring possess four ordinary and

B C

•• •.

;*^A??!-

D >-^-

Fig. 71. — Stages in the spermatogenesis and oogenesis of the honeybee.
A, B. First maturation division in the miale. C. Second matura-
tion division in the male. Three cells are produced : the first (RKi)
without chromatin ; the second {RK-i) with chromatin, but small
and functionless ; and the third a functional spermatid. {After
Meves, 1907.)

D. First maturation division in the female showing polar body
with eight dyads, and secondary oocyte with eight dyads. E. Sec-
ond maturation division in the female showing the divided first
polar body, the second polar body, and female pronucleus each with
eight monads. F. Outer end of first polar body disintegrating ;
inner half of first polar body uniting with second polar body, and
female pronucleus. {After Nachsheim, 1913.)

two sex chromosomes. These eggs give rise to
parthenogenetic females with the same number of

CHROMOSOMES AND MITOCHOxXDRlA ^207

chromosomes, and generation after generation of
such females appear during the summer; l)ut in
the autumn, females, whose eggs must be fertilized
before they will develop, and males are produced.
The chromosomes of these eggs are distributed during
maturation as shown in the diagram (Fig. 7^2).
The eggs that develop into the females possess the
usual number of chromosomes, but those that give
rise to males cast out in the polar body one chromo-
some that fails to divide, and hence are provided
with one chromosome less than the others. During
the maturation of the germ cells of these males two
sorts of spermatozoa are formed, one with three
chromosomes, the other with only two ; the latter
degenerate. Therefore, since only one sort of
spermatozoa is functional, the fertilized winter
eggs are all alike and all give rise to females (stem-
mothers) the following spring.

The chromosome distribution in certain nema-
todes resembles somewhat that of the phylloxerans.
Here, however, we have to deal with organisms that
are peculiar in several respects. Maupas (1900)
has shown that in the genus RhahcUiis the number
of males per 1000 females ranges from 45.0 to 0.15
according to the species ; and that these few males
do not copulate with the females and hence are func-
tionless. Furthermore, the females are not true
females, but hermaphrodites. Kruger (1912) dis-
covered that in Rhahditis aherrcuis the nuclei of the
spermatozoa did not fuse with that of the v^^:^^, except
in one instance, but disappeared in the cytojilasm ;

268

GERM-CELL CYCLE IN ANIMALS

hence the spermatozoa simply initiate development.
The chromosome cycle of Rhabditis nigrovenosa has
been studied by Boveri (1911) and Schleip (1911).

rHYLLOXERA CARYMCAVLJS

T'cCiA. T'^:cac^ A A Q\

*y«rrut/ t^ ft2ft^eii -^sn.

C^D

c:rP

<Seouu(/ J^/z/ey yO Ik^

Ti'^a/f Sfii/ruiU.

o ) j-^^

Sf>t/mtalcaftc

Fig. 72. — Chromosome cycle in Phylloxera carycBcaulis. (From Mor-
gan's Heredity and Sex, published by the Columbia University Press.)

CHROMOSOMES AND MITOCHONDRIA 269

This nematode is a parasite in the king of the frog
for part of its Hfe cycle; during tliis period it re-
sembles the female, but is really hermaphroditic.
These hermaphrodites give rise to free-living indi-
viduals which are true males and females; the
eggs of the latter when fertilized develop into para-
sitic hermaphrodites. The oogonia and sperma-
togonia of the hermaphroditic parasites possess
twelve chromosomes (Fig. 73, ^1). The nucleus of
the mature egg is provided with six (B) . Two sorts
of spermatozoa are formed, one-half with six chromo-
somes, the other half with five ; the latter result from
the casting out of one chromosome (E) in a manner
similar to that described above in Phylloxera. The
eggs fertilized with the spermatozoa containing
six chromosomes (F) produce free-living, true fe-
males, whereas those fertilized by the spermatozoa
with five (G) develop into free-living, true males.
The hermaphroditic condition is regained as follows :
The free-living females give rise to eggs all with
six chromosomes ; the males, whose spermatogonia
contain eleven chromosomes, produce spermatozoa
with six or five chromosomes; those with the latter
number, however, are not functional, hence all
fertilized eggs must be provided with twelve chromo-
somes and develop into the hermaphroditic parasites.
The chromosome cycle in pteropod mollusks as
worked out by Zarnik (1911) seems even more re-
markable than that described for nematodes. The
hermaphroditic species, C resets acicula, possesses
twenty chromosomes, sixteen large ordinary chromo-

270

GERM-CELL CYCLE IN ANIMALS

Fig. 73. — Rhabditis nigrovenosa. Stages in maturation, fertilization,
and cleavage. A. Oogonium with twelve chromosomes. B. Sec-
ond maturation division. Pronucleus and second polar body each
with six chromosomes. C. Primary spermatocyte. D. Division of
primary spermatocyte. E. Second spermatocyte division ; one
chromosome delayed. F. Two spermatozoa each with six chromo-
somes. G. Cleavage spindle of egg showing two groups of chromo-
somes ; one with six contributed by the egg, the other with five
contributed by the sperm. {After Schleip, 1911.)

CHROMOSOMES AND MITOCIIOXDRIA ^71

somes (shown in black in Fig. 74), two lar^e sex-
chromosomes (dotted), and two small sex-clironio-

Spermato-
gonium

Spermato-
cyte 1. Ordn.

Spermato-
cyte 2. Ordn

Spermien

Oogonium.

/^^p^\\ Oocyte 1. Ordn.

C /• J Oocyte 2. Ordn.

Reifes Ei.

Fig. 74. — Diagrams showing the chromosome cycle in the pteropod
mollusk, Creseis acicula. In order to sinipHfy thi- (Ha^rams each
black chromosome is made to represent ei^ht ordinary chromosomes.
{After Zarnik, 1911.)

somes (dotted). The spermalo^'oiiia outer the mat-
uration period in this condition. The nuniljcr of

272 GERM-CELL CYCLE IN ANIMALS

chromosomes is reduced in the first division, resulting
in two secondary spermatocytes each with eight
large ordinary chromosomes, and one large and one
small sex-chromosome. During the second division
the small sex-chromosome does not divide, but passes
intact into one spermatid ; thus two sorts of sperma-
tozoa are formed, one with eight large ordinary and
one sex chromosome and the others with eight
large ordinary chromosomes and two large sex-
chromosomes. The spermatozoa with only one
sex chromosome is not functional. The oogonia
differ from the spermatogonia and somatic cells in
the possession of sixteen large ordinary chromosomes
and four small sex-chromosomes ; two of the latter
arise by the diminution of the chromatin in two of
the large sex-chromosomes. The maturation divi-
sions are of the usual sort, and all of the eggs are
alike, containing eight large ordinary chromosomes
and two small sex-chromosomes. Fertilization, as
indicated in Fig. 74, always results in a zygote with
sixteen large ordinary chromosomes, two large sex-
chromosomes, and two small sex-chromosomes, which
develop into a hermaphroditic individual.

Although we know very little about the chromo-
somes of man, the data available seem to indicate
that here also there are chromatin bodies concerned
with sex-determination. The following table indi-
cates the state of our knowledge at the present time.

Guyer (1910) was the first to announce the dis-
covery of accessory chromosomes in man. He found
twenty-two chromosomes in the spermatogonia,

CHROMOSOMES AND MITOCIIOXDRIA 273

Table Showing the Number of Chromosomes in Man

ACCORDING TO VaRIOUS INVESTIGATORS

Diploid
Number

Haploid Number

Investigator

Date

Bardelehen

1892

Flemming

1897

18 (15 or 19) 1

Wilcox

1900

Duesberg

190(1

Farmer, Moore, and AValker

1900

Moore and Arnold

1900

12 or 10

Giiver

1910

12 or 10

Montgomery

1912

24(?)

Gutherz

1912

23 or 24

Winiwarter

19 b2

34 (33, 38)

Wieman

1913

which became ten bivalent and two accessories in
the primary spermatocytes. The latter pass un-
divided to one pole (Fig. 75, A), and hence two classes
of spermatozoa result, one with ten ordinary chromo-
somes, and the other with ten ordinary and two
accessory chromosomes. Winiwarter (1912), on the
other hand (Fig. 75, D-E), reports forty-seven
chromosomes in the spermatogonia and two classes
of spermatozoa, one with twenty-three and the
other with twentv-four. The number in the female,
according to Winiwarter, is probably forty-eight,
and hence all mature eggs are alike so far as chromo-
some number is concerned, each being provided
with twentv-four. If these data are coniirmed, it is
evident that sex in man is determined at the time
of fertilization and cannot be influenced by changing
the environment.

1 Wilcox doesn't state whether this is the reduced or diploid number.
T

274 GERM-CELL CYCLE IN ANIMALS

The above illustrations indicate that there is some
internal mechanism which controls sex, and that
certain chromosomes are, in at least many cases,

yi',"': »■•,■■.«";-"•■**•.

•■'-—.>

\-sxy:

Fig. 75. — Chromosomes in man. A. First spermatocyte division show-
ing two accessories passing early to one pole. B. Two contiguous
spermatids, one without and the other with two accessories. C. Two
secondary spermatocytes ; the one above with an accessory. D. Sec-
ond spermatocyte with twenty-four dyads. E. Second spermatocyte
with twenty-three dyads. \A-B, from Guyer, 1910; C-E, from
Winiwarter, 1912.)

factors in sex-determination. Several hypotheses
have been suggested as to the relation of these
chromosomes to sex, such as that sex is determined
by the quantity of chromatin present in the zygote.

CHROMOSOMES AND MITOC IIOXDRI V ^275

No view, however, has won general acceptance, but
it seems probable that there are fundamental inter-
relations between the different parts of the cell which
regulate the behavior of the chromosomes. We
must, therefore, look further for an explanation of
sex-determination. It has been suggested that
differences in metabolism may be responsible for the
fundamental differences between the sexes. Ac-
cording to this view changes in metabolism may
control the behavior of the sex-chromosomes, or the
presence of the sex-chromosomes in every cell in
the body may influence the metabolism "in such a
way that the organism is caused to become of one
sex rather than of the other, in consequence of its
type of metabolism " (Doncaster, 1914, p. 515).

The Mitochondria of Germ Cells

The study of the relative importance of the nucleus
and the cytoplasm in heredity has been given a new
impetus within recent years by the more accurate
examination and description of certain cytoplasmic
inclusions of both germ cells and somatic cells known
as mitochondria, chondriosomes, plastosomes, chro-
midia, etc. Some of the best recent evidence that
part of the germ-plasm may be located in the cyto-
plasm is afforded by the work of Bcnda, Meves,
Regaud, Duesberg, and others on the history of
these mitochondrial bodies during maturation, fer-
tilization, early cleavage, and cellular differentiation.
As long as forty years ago the cytoplasm of the
germ cells was known to contain bodies other than

276 GERM-CELL CYCLE IN ANIMALS

the nucleus ; these bodies have been given various
names such as spherules (Kunstler, 1882), cytomi-
crosomes (La Valette St. George, 1886), bioblasts
(Altmann, 1890), and ergastoplasm (Bouin, 1898).
In 1897 and 1898 Benda noticed the constant pres-
ence of certain granules in the male germ cells of a
number of vertebrates and was able to trace their
history from the spermatogonia until they formed the
spiral filament in the tail of the spermatozoa. These
observations were extended the following year
(1899) so as to include all stages in the development
of the eggs and spermatozoa of many vertebrates
and invertebrates and also various tissue cells such
as striated muscle-fibers, leucocytes, marrow-cells,
etc. This work attracted wide attention chiefly
for two reasons: (1) the history of the granules
was carefully worked out and the various stages
accurately described, and (2) special, rather com-
plicated, staining methods were devised which were
supposed to color the mitochondria so that they
could be distinguished from all other cell inclusions.
From 1899 until the present time an ever increasing
number of investigators have attacked the problems
presented by the mitochondria, or referred to these
structures incidentally when working upon other his-
tological or cytological problems. The study of mito-
chondria received its greatest impetus, however,
in 1908, when Meves published a paper on these
structures in the chick embryo entitled ''Die Chon-
driosomen als Trager erblicher Anlagen." In this
paper the chick embryo is described from the fifteen-

CHROMOSOMES AND MITOCHONDRIA 2

■: / /

hour stage up to the three-days-nine-liour si age.
The cells of the earHest stage studied contained mito-
chondria (Fig. 76) which were differently arranged
in the germinal layers : the ectoderm and entoderm
cells contained, for the most part, rods and threads,
the granules being scarce, and the mesoderm cells
were characterized by numerous granules and few
rods and threads. At the three-day stage the mito-
chondria of the neuroblasts became difficult to
stain by the usual method, but did stain like neuro-
fibrils. These and other observations led Meves
to the conclusion that the mitochondria are of con-
siderable importance in cellular differentiation and
are in fact the bearers of hereditary Anlagen.

Since this paper of Meves appeared, the zoological
periodicals have been flooded with the results of in-
vestigations of the mitochondria in almost every
sort of germ and somatic cell, both normal and
abnormal, and in Protozoa and Metazoa, In-
vertebrates and Vertebrates. No report on
spermatogenesis, oogenesis, or early embryonic de-
velopment is complete without reference to the mito-
chondria. In plants, also, cellular bodies have been
described of a mitochondrial nature (Meves, 1904 ;
Duesberg and Hoven, 1910; Guilliermond, 1911).

A large number of new terms have been coined
for the purpose of describing these cytoj)lasniic in-
clusions. Some of them are as follows: (1) mito-
chondria, applied by Benda (1897, 189cS) to certain
granules with definite staining reactions : ('2) chon-
driosomes, proposed by jVIeves (1908) for both single

278 GERxM-CELL CYCLE IN AxNIMALS

^'"'■ll--}-^}^'''^.^^^^^ in the embryonic cells of the chick A In
cells of the primitive strenk- R Jt^a;.-^- "^c luilk. jt. m

C In connective tissue cet, D. Int"a' "LrSf 'T'Z^ 'f^'
oerg, wi^, a, B, C, E, after Meves; D, F, after Duesherg.)

CHROMOSOMES AND MITOCHONDRIA 279

granules and chains of granules ; the latter were also
called chondriokonts ; (3) plastosomes (plastochon-
dria, plastokonta), employed by Meves (1910) be-
cause of their supposed role in histogenesis ; (4)
eclectosomes, selected by Regaud (1909) as a general
physiological expression for chondriosonies ; (5) chon-
driotaxis, used by Giglios-Tos and Granata (1908)
to describe the parallel arrangement of chondrio-
konts ; (6) chondriodierese, proposed by the same
authors for the division of the chondriokonts during
cell division ; (7) karyochondria, coined by AVildman
(1913) for cytoplasmic inclusions derived from the
basichromatin of the nucleus ; (8) chromidia, a term
considered by Goldschmidt (1904) and others to in-
clude the mitochondria.

We are here especially interested in the mitochon-
dria of the germ cells, their origin, fate, and signif-
icance, but our ideas regarding the importance of
these bodies in heredity depend somewhat upon their
behavior in somatic cells. As already stated,
Benda (1903) observed mitochondria in both germ
cells and somatic cells. Since then they have been
recorded in Protozoa, in almost every sort of somatic
cell in Metazoa, and in many plant cells (Fig. 77).
Excellent reviews have been published by Benda
(1903), Faure-Fremiet (1910), Prenant (1910), and
Duesberg (1912). These reviews have led to the
conclusion already expressed by Regaud (1909,
p. 920) that "it is probable that they (mitochondria)
exist in all cells, at least at certain stages in their
activities." Among the somatic differentiations to

280

GERM-CELL CYCLE IN ANIMALS

which mitochondria are supposed to give rise are
neurofibrils and myofibrils. Meves (1907, 1908)
considered it probable that neurofibrids were trans-
formed chondriosomes, and Hoven (1910) seemed to
have proved it, but Marcora (1911) and Cowdry
(1914) find that the neurofibrils arise independently,

^yJi.

A B C

Fig. 77. — Mitochondria in the cells of a plant, Pisum sativum.
A. Young germ cell. B. Young germ cell dividing. C. Old cell
containing vacuoles. {From Duesberg and Hoven, 1910.)

although mitochondria are present in the nerve
cells. Duesberg (1910) is quite positive that the
myofibrils of striated muscle fibers are produced
by the metamorphosis of chondriosomes from em-
bryonic muscle cells, and has recently (Duesberg,
1913) strengthened his position by the discovery
that the myoplasm described by Conklin (1905)
in the egg of the Ascidian, Cynthia, is well supplied
with chondriosomes.

Mitochondrial structures have been studied in
both living and preserved cells. Faure-Fremiet
(1910) describes them in living cells (Fig. 78, D) as

CHROMOSOMES AND MITOCHOXDRIA 281

small, transparent, slightly refringent granuk's of a
pale gray tint, either homogeneous or else vesicular
with fluid contents and a thin, denser, rci'ringcuL
periphery. Rod-like mitochondria were likewise
observed by Montgomery (1911) in the living male
germ cells of Euschistus (Fig. 78, A-B) which had
been teased out in Ringer's solution ; and this in-

A O

Fig. 78. — Division of mitochondria. A-B. Mitochondrial rods divid-
ing during first maturation division in Euschistus. C. Stages in
division of mitochondrial l)ody in Hudromctra. D. Sinniltnnt'oua
division of micronucleus and mitochondria in Carchcsium (in vivo).
(A-B, from Montgomery, 1911; C, from Wilke, 1913; D, from
Faure-Fremiet, 1910.)

vestigator concluded that in preserved material "we
have been working with images that are very close
to the living. ..." More recently Lewis [{\u\
Lewis (1914) have made careful .studies of niiloclion-
dria in living cells from chick embryos. Granules
were here seen *'to fu.se together into rods or chaius,
and these to elongate into threads, wliich iu turn
anastomose with each other and may nnilc into a
complicated network, which iu turn may again
break down into threads, rods, loops, and rings."
Even more remarkable are the movements within the

282 GERM-CELL CYCLE IN ANIMALS

cell described by the same investigators. "The
mitochondria are almost never at rest, but are con-
tinually changing their position and also their shape.
The changes in shape are truly remarkable, not only
in the great variety of forms, but also in the rapidity
with which they change from one form to another. A
single mitochondrium may bend back and forth
with a somewhat undulatory movement, or thicken
at one end and thin out at the other with an appear-
ance almost like that of pulsation, repeating this
process many times. Again, a single mitochondrium
sometimes twists and turns rapidly as though
attached at one end, like the lashing of a flagellum,
then suddenly moves off to another position in the
cytoplasm as though some tension had been re-
leased." Mitochondria may also be stained intra
vitam, especially with dahlia violet and Janus green.
Most of the fixing solutions ordinarily used for cy to-
logical purposes destroy the mitochondria. The
methods which seem to give the best results have
osmic acid or formalin as a basis, such as those de-
vised by Altmann (see Lee, 1905, p. 43), Benda
(Lee, 1905, p. 223), Meves (1908), and Regaud
(1908, p. 661). Benda (1903) claimed that all
cellular structures which stained violet by his method
were of a mitochondrial nature ; but this has not been
found to hold true. Undoubtedly the many bodies
which have been discovered in cells are of several
sorts, and only by a thorough study of their staining
qualities, morphological aspects, and biological roles
can they be identified. Benda's crystal violet

CHROMOSOMES AND MITOC IIOXDRIA 283

stain seems to be more selective than any other for
mitochondria and is of great value for this reason.

Mitochondria most often appear as spherical
or elongated granules about 0.001 mm. diameter.
These granules may become arranged in a series,
thus forming a chain, and the granuK's in a chain
may fuse into a homogeneous rod. Different forms
are present in different kinds of cells or even in the
same cell at various stages in its evolution or func-
tional activity. Some investigators (Prenanl. 1!)10)
maintain that the homogeneous rod is the i)rimitive
condition and that the granules are formed by the
disintegration of such rods; to others just the
reverse seems to be true (Rubaschkin, 1910; Dues-
berg, 1912).

The chemical constitution of the mitoelionchia
has been studied by a number of investigators.
Regaud (1908) has shown that the miloehondria of
the seminal epithelium are not histocluMnically
identical. He distinguishes three sorts of granuk's :
(1) those which resist the action of acetic acid and
are stainable without being previously innncrsed in
a solution of potassium bichroniale, (2) granuk's
which resist acetic acid but require int(Mis(^ ctiromisa-
tion, and (3) granules which (k) not resist acetic-
acid and demand ckromisation. Faure-Frenn'el ,
Mayer, and Scliiiffer (1<)09) have studied the mito-
chondria by microchemical and conipaiali\c niclliods
and reached the conclusion that llicy are lecillial-

bumins.

Mitochondria have been noted in all stages of

284

GERM-CELL CYCLE IN ANIMALS

the male germ-cell cycle, especially in mammals,
mollusks, and insects, and appear to be continuous
from one generation of cells to the next. During

Fig. 79. — Behavior of the mitochondria during the fertilization and
early cleavage of the egg of Ascaris. A. Egg into which a sperma-
tozoon has penetrated. B, C. The mixing of the mitochondria of
the egg and spermatozoon. D. Division stage of the first two blas-
tomeres. (After Meves, 1911 and 1914.)

mitosis the plastosomes lie outside of the spindle
(Fig. 79, D) ; they may divide autonomously as
claimed by Faure-Fremiet (1910) in Protozoa (Fig.

CHROMOSOMES AND MITOCHONDRIA ^285

78, D) and Wilke (10V2) in tlie spermatocytes
of Ilydrometra or en masse, as in the spermatogenesis
of Euschistus (Fig. 78, A-B), thus undergoing a sort
of paramitosis (Montgomery, 1911) and Xofouecfa
(Browne, 1913). In the former cases each (hiughter
cell is supposed to receive one-half of each gramde;
in the latter the distribution is largely by chance,
but apparently equal (Cowdry, 1914). Accor( Hug
to certain observers the centrosomes exert an in-
fluence upon the mitochondria as indicated by the
aggregation of these bodies around the asters (Faure-
Fremiet, 1910; Meves, 19U) ; but others have been
unable to find any confirmatory evidence in their
material (Montgomery, 1911). Duesberg (1908)
has pointed out that since there is no rest period
between the two maturation divisions there nuist be
a quantitative reduction of plastosomes in the sj)er-
matids; a quartering of the mitochondria couKl
not, however, be observed by :Montgomery (191 '■2)
in Peripatus. Montgomery (1911) has suggested
that the relative amount of the mitochondrial sub-
stance received ''might determine the sex-i)rcp()n-
derance character of the sperm, a matter unfor-
tunately very difficult to test."

Faure-Fremiet recognizes four ty]H\s of mitochon-
drial distribution in the germ cells: (1) filaments or
masses that do not undergo i)r()found mori)h()logical
changes (Fig. 80) ; (2) one or more masses which
transform into a definite morphological element,
the Nebenkern; (3) masses which only ])artially
change into a Nebenkern or yolk nucleus ; (4) bodies

286

GERM-CELL CYCLE IN ANIMALS

which transform entirely or in part into deuto-
plasmic granules of a fatty nature.

The origin of the mitochondria in male cells can-
not be stated definitely, since certain investigators
(Goldschmidt, Buchner, Wassilieff, etc.) claim that
they arise from the nucleus ; others (e.g.,M.eves, Wilke,
Duesberg) consider them to be integral parts of the

cytoplasm ; and a third
group (Montgomery,
Browne, Wildman)
looks upon some of
them as the results
of chemical interaction
between the nucleus
and the cytoplasm.

Less is known con-
cerning the mitochon-
dria during oogenesis
than during sperma-
togenesis, but certain bodies have been described in
the ova of a number of animals which exhibit all of
the characteristics of the mitochondria of male cells.
As in the latter, they have been considered chromidial
by some and of cytoplasmic origin by others.

The importance of the mitochondria depends
largely upon their functions. Those of the egg have
been observed by Russo (1907), Loyez (1909),
Faure-Fremiet (1910), Van Durme (1914), Hegner
(1914a), and others to transform directly into yolk
globules. According to Van der Stricht (1904),
Lams (1907), etc., they produce yolk elements in-

FiG. 80. — Four stages in the formation
of the spermatozoon of Enteroxenos
showing the distribution of the mito-
chondria (M). {After Bonnevie.)

CHROMOSOMES Ax\l) MlTO( IIONDUIA ^287

directly; and It is the opinion of Movos, Dueshcr^S
and their followers that they play an iniporlaiit mle
in fertilization. Likewise in the spermatozoa ideas
differ regarding their functions. I^cnda (IS!)!))
believed them to be motor organs; Kollzofl' (l!)(l(i),
from a study of the spermatozoa of Decapods,
maintains that they represent elements whidi form
a sort of cellular skeleton; Rcgaud (1J)()!)) claims
that they are the particular crlhihir organs which
exercise a *'fonction eclectique," extracting and
fixing substances in the cell, and should Iherefore he
called " eclectosomes " ; and Meves (1907, 1908)
holds that they are cytoplasmic constituents cor-
responding to the chromosomes of the nuch'us.
Meves (1907, 1908) came to the conclusion that tliere
must be hereditarj^ substances in the cyl()j)la>ni,
and by the method of elimination decide(j in faxor
of the mitochondria. In his stu(hes on fertihzation
and cleavage in Ascaris (Meves, 1911. 1!)M-) he has
shown that granules from the spermatozoon (Fig.
79) fuse with similar granules in the v^^:,^^, as described
previously by L. and R. Zoja (1891), and thai (lie.se
granules are plastosomes. The distiibntion of the
fused granules is followed until the am|)hiaster is
formed in the two-cell stage; here the ])lastosomes
are mainly grouped about the centrosomes, although
a few are scattered about in the cytoplasm (Fig. 79,
D).

Although there are many who believe Meves and
his followers to be correct in their contention that
the plastosomes are the bearers of hereditary charac-

288 GERM-CELL CYCLE IN ANIMALS

teristics in the cytoplasm, just as the chromosomes are
the bearers of hereditary characteristics in the
nucleus, still there are many objections to this view,
such as the fact that part or all of the plastosomes
may be cast out of the spermatid {e.g., in the opos-
sum, Jordan, 1911 ; and in Peripatus, Montgomery,
1912). It is obvious from the foregoing account
that there are a number of opposing views regarding
the origin, nature, and role of the various cytoplasmic
inclusions which have been considered mitochondria.
Are they constant, necessary constituents of the
living protoplasm, or are they inactive lifeless bodies
which may be included under the term metaplasm ?
If they constitute a part of the living protoplasm,
do they form the skeleton of the cell, do they take
part in the metabolic activities of the cytoplasm
or nucleus, or do they play a role in the process of
differentiation, and should they be considered as
the hereditary substance of the cytoplasm ? If
they are simply metabolic products, are they excretory
in nature, or reserve materials set aside for the later
use of the cell ? And finally, do they arise from the
nucleus, are they strictly cytoplasmic, or do they
originate through the interaction of nucleus and
cytoplasm ? It is impossible in a short space to
give an adequate account of the arguments pro and
con, and so we must refer the reader to the compre-
hensive reviews mentioned above. The conclusion,
however, is perfectly safe that we shall have to await
the results of further investigations before we can
come to a definite decision. In the meantime we

CHROMOSOMES AND M1T()( IIONDKIA 289

should thank the mitochondria for focusing llie
attention of cytologists upon tlie cytoplasnn'c cl(»-
ments, since the belief is becoming more and more
general that hereditary phenomena are the result of
interactions between nucleus and cytophism and that
the latter may play a more important role than is
usually supposed.

CHAPTER X

THE GERM-PLASM THEORY

In discussing the germ-plasm theory it is necessary
to distinguish between this hypothesis and that of
the morphological continuity of the germ cells. The
facts and theories involved have grown up to-
gether. Owen (1849) was perhaps the first to
point out the differences between germ cells and body
cells. "Not all of the progeny of the primary impreg-
nated germ cell, " he writes, "are required for the for-
mation of the body in all animals ; certain of the de-
rivative germ cells may remain unchanged and become
included in that body which has been composed of
their metamorphosed and diversely combined and
confluent brethren; so included, any derivative
germ cell or the nucleus of such may commence and
repeat the same processes of growth by imbibition,
and of propagation by spontaneous fission, as those
to which itself owed its origin. ..." Galton (1872)
was among the earliest to recognize the necessity
for two sorts of materials in the individual metazoon,
"one of which is latent and only known to us by its
effects on his posterity, while the other is potent,
and constitutes the person manifest to our senses."
He at that time believed in the inheritance of ac-
quired characters and conceived the egg as a struc-

290

THE GERM PLASM TilKOin 291

tureless body from wliicli holli I lie body .ind the ova
of the individual evolve; and considered lliese ova
to consist of contributions partly from I he v<n^ and
partly from the body which develoix'd from the e^'^'.
Later Jager (1877) stated the idea of gerim'nal con-
tinuity more definitely. He maintained that |)art
of the germ-plasm (Keini Protoplasma) of the
animal forms the individual, and the rest is re-
served until sexual maturity, when it forms the re[)ro-
ductive material. The reservation of this j)hyl()-
genetic substance he termed the "continuity of
the germ-plasm" ("Continuitiit des Keimproto-
plasmas"). To Weismann (1885) is usually givtMi
the credit for originating the germ-j)lasui theory,
but while w^e are undoubtedly iiKh'bted lo him for
the great influence the hyj^othesis of germinal con-
tinuity has had upon the trend of biological in\ (•>({-
gations within the past thirty years, we must con-
sider Jager as the first to clearly enunciate I he i(N a.
Jager (1878) also expressed a belief in the mor-
phological continuity of the germ cells of succeed-
ing generations, but this idea was first definitely
stated by Nussbaum (1880), whose investigations
of the germ cells in the trout and I'log led him to
conclude that the cleavage cells t'orni two groups
independent of each other. One group contains
the cells which multi})ly and dill'erent iate and Ihns
build up the body of the individual, but do not |)i'o-
duce germ cells; the other group takes no pari in
the formation of the body and undergoes no ditl'eren-
tiations, but mult ij)lics by simple division. The germ

292 GERM-CELL CYCLE IN ANIMALS

cells are thus not derived from the individual in
which they lie, but have a common origin with it.
The segregated germ cells or species substance is
therefore distinct and independent of the individual ;
this accounts for the constancy of the species. We
may distinguish between the two ideas by defining
them as follows :

(1) Germinal continuity, or the germ-plasm
theory. "In each ontogeny a part of the specific
germ-plasm contained in the parent egg-cell is not
used up in the construction of the body of the off-
spring, but is reserved unchanged for the formation
of the germ cells of the following generation"
(Weismann, 1891, p. 170).

(2) Morphological continuity of the germ cells.
The developing egg produces by division two sorts of
cells, germ cells which contain the germ-plasm and
somatic cells which protect, nourish, and transport
the germ cells until they leave the body to give
rise to the succeeding generation.

No case of a complete morphological continuity
of germ cells has ever been described. Such an
occurrence would necessitate the division of the egg
into two cells, one of which would give rise to all
of the body cells and nothing else, the other only to
germ cells. The behavior of the germ-plasm in such
a case would be as follows (Weismann, 1904, p. 410) :
"The germ-plasm of the ovum first doubles itself
by growth, as the nuclear substance does at every
nuclear division, and then divides into two similar
halves, one of which, lying in the primordial somatic

THE GERM PLASM TIIKORV 293

cell, bec'oiiu's al oner acllxc and breaks up into
sinallor and smaller ^r()U])s oi" (Ictcrnniiaiits corre-
sponding to the building up of llic Ixxly, wliilc tlir
germ-plasm in the other ri'iuains in a more or less
'bonnd' or 'set' condition, and is only actJNc to the
extent of gradually stamping as germ cells the cells
which arise from the primordial germ cell."

According to Weismann this actually occurs in
Dipterous insects, but there is no evidence in the
literature to warrant this statement. It is conse-
quently necessary to imagine the germ-plasm as
present but not definitely localized in a germ cell
until some time after the two-cell stage has been
reached. Thus in hydroids Weismanu explains the
situation as follows: "Here the j)rimordial germ
cell is separated from the ovum by a long series of
cell-generations, and the sole possibility of explaining
the presence of germ-plasm in thi> i)rimor(lial germ
cell is to be found in the assumi)lion that in the
divisions of the ovum th(* whole oi' the gerni-pla>m
originally contained in it was not broken up into
determinant groups, but that a i)art, i)erhaps the
greater part, was handed on in a latent state from
cell to cell, till sooner or later it reached a cell which
it stamped as the primordial germ cell."

Evidence that the germ-])lasm does become sooner
or later localized in the i)rimor(lial germ cell has accu-
i mulated rai)idly within recent years. In the pa^lo-

genetic fly, Miastor (see (1iai)ter III), the fir>t (vU
to be cut oflF from the egg is the i)rimordial germ cell
(Fig. 17, p.g.c). although at this time there are

294 GERM-CELL CYCLE IN ANIMALS

eight nuclei in the egg. As determined by Kahle
(1908) and confirmed by the writer (Hegner, 1912,
1914a), this primordial germ cell gives rise to sixty-
four oogonia and to no other cells. This is the nearest
approach to a complete morphological continuity
of the germ cells that has yet been described, and
since this primordial germ cell must contain the germ-
plasm of the succeeding generation, the condition
in this fly is really comparable to that of the hypo-
thetical case cited above, only in Miastor the cell
set aside for reproductive purposes is much less than
one-half of the egg, the somatic part of the egg being
not a single cell, but a syncytium containing seven
nuclei.

We may therefore look for the germ-plasm of
Miastor in the primordial germ cell. So far as we
know there are only two sorts of materials in this
cell, that contained in the nucleus, and the darkly
staining part of the egg which becomes recognizable
just before maturation occurs, is situated at the pos-
terior pole, and has been termed the pole-plasm
(Fig. 13). If the primordial germ-cell multiplies by
simple division and if there is an equal distribution
of the contents at every mitosis, then the sixty-
four oogonia must each possess one sixty-fourth of
both the nucleus and the pole-plasm of the primordial
germ cell plus any materials that have been added
during the period of multiplication. An enormous
enlargement occurs during the growth period both of
the nucleus and of the cell. The pole-plasm cannot
be recognized at this time, but again becomes

THE GERM PLASM THEORY ^2U5

evident just before nialuration ; it lias increased in
amount to approximately sixty-four limes its iornicr
mass. How this increase lias been brou^'lit about is
not known, but it has been su^^f^^esied (p. G8) that
preexisting particles of ])()lc-plasni may grow and
divide, or the dilution of the j)ole-plasm caused l»y
the growth of the egg might start into action some
catalyst which would cause the production of niort*
substance Hke the pole-plasm and cease its activity
when the amount of ])()le-plasm characteristic of
the mature egg had accumulated and brought it to a
state of equilibrium. In the midge, C/iirunomu.s,
the primordial germ cell is segregated even earlier
than in Miastor, namely, at the four-cell stage.
The later historv of the germ cells is not so well
known in this species, however, as in Minstor.
The data presented in Chapters V and \'I ))i()\e
that a definite and early segregation of germ ceils is
known in a sufficient number of groups to indicate
that the process is quite general among animals.
The morphological continuity of the germ cells,
how^ever, cannot be established with such a degree of
certainty in the vertebrates, and although mu.^L
investigators believe that the gvr\^^ cells wvv con-
tinuous, still the entire keind)ahn has never been
traced as accurately as it has in many invertebrates.
Fortunately almost every new investigation contains
additional data and more refined methods which lead
us to hope that some tini(^ in the nivir future the
primordial germ cells cvcmi here may be traced back
to early cleavage stages.

^296 GERM-CELL CYCLE IN ANIMALS

One of the distinguishing features of many primor-
dial germ cells is the presence within their cytoplasm
of certain stainable bodies to which I have applied
the term "keimbahn-determinants." Although, as
pointed out in Chapter VIII, these inclusions do not
appear to consist of the same sort of material in
the eggs of different species and hence their signif-
icance is problematical, still they seem to be asso-
ciated with that particular part of the egg sub-
stance which becomes the cytoplasm of the primor-
dial germ cells. For this reason, if for no other,
the keimbahn-determinants are of the greatest
value, since they enable us to determine the position
of this germ-cell substance during the stages before
the primordial germ cells are established. It is
therefore possible to trace the germ-cell substance
in such cases as Sagitta (Fig. 54), where there is no
morphological continuity of the germ cells. What
relation the keimbahn-determinants have to the germ-
plasm is not yet definitely known.

There have, of course, been many objections to
the germ-plasm theory. The history of the germ
cells in the Coelenterata, upon which Weismann
(1882) based a large part of his argument, is consid-
ered by Hargitt (see p. 95) to be directly opposed
to the hypothesis. According to some zoologists
there is no essential difference between the repro-
ductive cells and the various sorts of somatic cells ;
they have all arisen as the result of division of labor,
and the germ cells have been differentiated for pur-
poses of heredity just as the muscle cells have been

THE GERM-PLASM THEORY 297

differentiated for causing motion and the nerve cells
for receiving and conducting stimuli. That the irerm
cells remain in a primitive condition during a large
part of the embryonic i)erio(l is accounted for hy
the fact that thev become functional at a compara-
tively late stage in ontogeny (Eigenmann, ISDO).
Asexual reproduction by means of fission or budding
has seemed to some to invalidate the theory of ger-
minal continuity, but as Montgomery (IDOd, ]>. 8''2)
has pointed out, "Perhaps in all cases products of
asexual generation contain germ cells. If this were
so, it might then be the case that the incapacity
of any part of the body of an animal to reproduce
asexually, or even to regenerate, would l)e due to
the absence of germ cells in it — but this is merely
a suggestion." The probability that the regenerat-
ing pieces of coelenterates and the artificial Plas-
modia formed by dissociated sponge cells contain
germ cells has already been noted (]). 79), but there
are cases of the regeneration of sex organs that are
not so easily explained. For example, Janda ( 191 ••2)
has found that if the anterior part of the hermaph-
roditic annelid, CriodrUus laruinn, is n^moved, a
new anterior end will regenerate containing both
ovaries and testes, although not always in their
normal positions.

The study of the germ cells in the cestode Moniczia
expansa convinced Child (11)0()) that germ cells may
develop from tissue cells. In this species the germ cells
are derived from the parenchymal syncytium, which
has undergone a considerable degree of cytoplasmic

298 GERM-CELL CYCLE IN ANIMALS

differentiation and therefore consists of real tissue
cells. Those parenchymal cells that encounter
certain conditions become germ cells. Later (1906)
the same author gave an account of the development
of spermatogonia in the same animal from the dif-
ferentiated muscle cells. These studies, together with
the results from experiments on regeneration, have led
Child (1912) to the belief ''that this germ-plasm
hypothesis and the subsidiary hypotheses which
have grown up about it are not only unnecessary
and constitute an impediment to biological thought,
which has retarded its progress in recent years to a
very appreciable extent, but furthermore, that they
are not in full accord with observed facts and can
be maintained only so long as we ignore the facts."
He further maintains that if protoplasm is a physico-
chemical substance it is capable of changing its con-
stitution in any direction according to the conditions
imposed upon it, and that therefore the continuous
existence of a germ-plasm with a given specific
constitution is vmnecessary.

The evidence in favor of the germ-plasm theory
is so strong that the arguments thus far advanced
against it have had but little influence. If, then, we
accept germinal continuity as a fact and consider
the germ-plasm to be a substance that is not con-
taminated by the body in which it lies, but remains
inviolate generation after generation, we should next
inquire as to the nature of this substance. The
generally accepted idea is that the chromatin of the
nucleus represents the physical basis of heredity. In

THE GERM-PLASM IIIEORY 299

favor of this view are the i'acts thai (Inriiiii- mitosis
the number and shape of the cliroinosonies are con-
stant in every species (variations soinetiines occur)
and the complex series of processes in indirect nuclear
division seems to be for the sole purpose of dividing
the chromosomes equally between the daughter
cells; even during the intervals (interkinesis) be-
tween successive mitoses the chromosomes may be
recognized in certain species as prochromosomes
(see Digby, 1914, for review of literature). During
the maturation of the germ cells chromosomes
seem to play the most important role, uniting in
synapsis, and separating in the reducing division.
The chromosomes of the minute, motile spernui-
tozoa equal in number those of the comparatively
enormous, passive egg; the spermatozoon consists
almost entirely of chromatin, and this is tlu^ only
substance present in the zygote that is e(iually
contributed by both egg and spernuitozoiin. The
processes following the penetration of the spermato-
zoon into the egg bring about a combination of the
chromosomes of the two gametes into a single
nucleus; in certain animals at least some characters
depend upon the presence of a certain chromosome,
the X-chromosome ; in certain cases of ])olyspcrmy
the addition of extra male chromosomes seems to
be the cause of tlu^ abnormal dcn-c^lopnuMit of the egg.
These and many other facts of chromosome be-
havior that have been discovered by observations
and experiments have convinced most biologists
that the chromatin is the germ-plasm.

300 GERM-CELL CYCLE IN ANIMALS

It is becoming more and more evident, however,
that the cytoplasm cannot be entirely excluded. As
noted in Chapter IX, the mitochondria appear to be
constant cell elements and may actually constitute
a part of the essential hereditary substance. Even
if these particular cytoplasmic bodies do not repre-
sent germ-plasm, still, as pointed out by Guyer (1911)
and others, cytoplasm as well as nuclear material is
necessary to explain the phenomena which we call
heredity. It was shown in Chapter I that the most
important primary constituents of protoplasm are
the proteins, and the idea is rapidly becoming general
that the mechanism of heredity consists of (1) fun-
damental species substances, probably mainly pro-
tein in nature, together w4th (2) equally specific
enzymic substances which regulate the sequences of
the various chemical and physical processes incident
to development (Guyer, 1911, p. 299). The chro-
mosomes have been suggested as enzymatic in
nature (Montgomery, 1910), but enzymes are sup-
posed merely to accelerate reaction already initiated,
and hence the substrate must be of as great importance
as the enzymes which work upon it. But the sub-
strates must be extremely numerous to supply each
species with its specific proteins. That there are
enough configurational differences in corresponding
protein molecules to supply the number for the
thousands of animal species is certain, since some
comparatively simple proteins may possess thousands
of millions of stereoisomers. Thus the study of
heredity substance involves primarily a knowledge

THE GER:\I PLASM TIIKOKY 801

of the nature and reactions of I lie clieinical constitu-
ents of protoplasm, for, as AVilsou flDbJ, p. (Ui) says,
*'The essential conclusion tluit is iudicatcd l)\ cvto-
logical study of the uuclear substance is, thai it is an
aggregate of many different chemical couipouents
which do not constitute a mere mechanical mixture,
but a complex organic system, and which inuhTgo
perfectly ordered processes of segregation and dis-
tribution in the cvcle of cell life."

Some of the strongest evidence that the germ-
plasm must include cytoplasmic constituents is
afforded by the observations and experiments ch^aling
with the differentiation of the germ cells, especially
during early embryonic development. ^Fhe writer's
morphological and experimental studies of chrysom-
elid beetles seem to prove that the nuclei during
the cleavage stages are all potentially alike and that
it is the cytoplasm which decides their fate. Boveri's
experiments on the eggs of Ascaris likewise show
that the cytoplasm determines the initiation of the
chromatin-diminution process and controls the differ-
entiation of the germ cells. Furthernu)re, nuich of
the data in the preceding cha])ters indicates that the
non-nuclear substance which will beconu' segregated
within the primordial germ cell is present in a nu)re
or less definite region in the nndivided egg, being
gradually localized and separated from the other v\i^;^
substances as cleavage progresses. 'V\\v })osili()n of
this germ-cell substance can in many ca.ses be (U'ter-
mined because of the presence ol niclusions ot vari-
ous sorts, but whether these keimbahn-determinants

302 GERM-CELL CYCLE IN ANIMALS

constitute an important part of the germ-plasm or
play a minor role in heredity is still uncertain.

Modern cytological studies and the results of ex-
perimental breeding both help to solve the prob-
lems of the combination and subsequent distribution
of the determiners or factors within the germ-plasm.
In fact, it has been maintained by certain geneticists
that "The modern study of heredity has proven
itself to be an instrument even more subtle in the
analysis of the materials of the germ cells than actual
observations on the germ cells themselves " (Morgan,
1913, p. v). Those who do not wish to commit
themselves as to the physical or chemical nature of
the germ-plasm are content to speak of determiners,
factors, or genes without connecting them with any
particular substances. The behavior of the chro-
mosomes, however, enables us to explain so many of
the facts of heredity that, as stated above, these
bodies are generally considered to constitute the
essential hereditary substance.

The study of heredity was wonderfully stimulated
by the recognition in 1900 by Correns, Von Tscher-
mak, and de Vries of the results of Mendel's (1866)
investigations on plants. One of the simplest of
Mendel's experiments is that which he performed
with differently colored peas (Fig. 81). A pea bear-
ing green seeds was crossed with a pea bearing yellow
seeds. The first (Fi) generation of peas resulting
from this cross all bore vellow seeds. When the in-
dividual plants of this generation were inbred, three-
fourths of the resulting {F2) generation were yellow

THE GERM -PLASM THEORY

303

and one-fourth green. This proved that the seeds
of the first generation (Fi), althougli yellow, si ill
possessed within them the factor for grtH'iiness in a
latent condition. Green was therefore called a re-

Fig. 81. — Diagram to illustrate Mondel's law of aogregution. Individ-
uals (zygotes) are represented by suptTiinposed circles, \vho.se colors
stand for the factors involved. Gamctrs (Kcrni cells) are repreaeutcd
by single circles. {From Morgan, 1U14-)

cessive character and yellow a dominant character.
As a result of breeding the (F2) secumi generation it
was found that all of the green seeds produced plants
which bore green seeds; that is, these plants were
pure green and "homozygous" as regards color;
whereas the plants which bore yellow seeds couKl l)e

304 GERM-CELL CYCLE IN ANIMALS

separated into two groups ; one, containing on the
average one-third of these plants, was pure yellow and
homozygous as regards color ; the other two-thirds,
although yellow, contained green in a latent condi-
tion and were therefore impure yellows and "hetero-
zygous" as regards color. The conclusion reached
was that the eggs and spermatozoa produced by the
first (Fi) generation (see Fig. 81) were pure yellow or
pure green and that chance combinations during
fertilization resulted in the three classes of individ-
uals in the second (F2) generation ; that is, one-fourth
pure yellow, one-fourth pure green, and one-half with
dominant yellow and green recessive. Evidently
the factors for yellow and green repulsed each other
during the maturation so that they became localized
in different germ cells.

Such a characteristic as the color of the seeds of
these peas is kno^vn as a unit character, and the sepa-
ration of the factors of such a character during
maturation is referred to as the principle of segrega-
tion, Mendel further discovered that if the seeds
were also wrinkled or round, such characters behaved
independently of the color characters. These and
other experiments described by Mendel opened
the way for new lines of investigation which have
yielded results of vast importance from the stand-
point of heredity and evolution.^

Soon after Mendel's results were "rediscovered"

^ For more detailed accounts of experiments and theories that have
been pubhshed wathin the past fourteen years the reader is referred
to the books of Bateson (1909, 1913) and Punnet (1911).

THE GERM-PLASM TIIEORV

30^

it was pointed out by Guycr (100^2), Sutloii fl!)03),
and others that the distribution of the achill cliar-
acteristics of hybrids which were found by Mendel
to reappear in the offspring in rather (k'finilc propor-

||. |i. \l' \\ ]\' J8-

Fig. 82. — Diagrams to show the pairs of ( hroiiiDsoiurs and their Ik»-
havior at the time of maturation of the (•««. Three pairs of eliromo-
somes are represented ; three from one parent, three from the other.
The six possible modes of separation of these three are shown in
the lowest line. (From Morgan, WI4)

tions, could be explained if these cliaraeteristies are
located in the chromosomes. During- synapsis, as
already explained (p. 44), homologous maternal and
paternal chromosomes are sui)posed to pair and then
separate in the reduction division. Tt seems ])robable
that the pairs of chromosomes do not occnpy any

306

GERM-CELL CYCLE IN ANIMALS

definite position on the spindle at this time, but, as
indicated in Fig. 82, the distribution of the maternal
and paternal chromosomes to the daughter cells is
entirely a matter of chance. If the homologous
maternal and paternal chromosomes really are dis-
tributed by chance to the eggs and spermatozoa
following synapsis, then the number of combinations
possible are as follows (Sutton, 1903) :

Somatic Series

Reduced Series

Combinations in
Gametes

Combinations in
Zygotes

256

'25Q

65536

4096

16777216

262144

68719476736

The only direct evidence that such distribution
of chromosomes takes place is that furnished re-
cently by Carothers (1913) from a study of the
spermatogenesis of three Orthopterous insects,
Brachystola magna, Arphia simplex, and Dissosteira
Carolina. Miss Carothers, while working in Pro-
fessor McClung's laboratory, discovered a tetrad in
the first spermatocytes of these insects which consists
of two unequal dyads (Fig. 83). During the two mat-
uration divisions the four parts of this tetrad pass
to the four spermatozoa, and consequently two sorts
of spermatozoa are produced so far as this chromo-
some is concerned, one-half with one of the larger
elements of the tetrad and one-half with one of the

THE (;i:R.M PLASM TIIKORY 307

smaller elements. These diU'ereiillv .sized (1\ .ids an-
considered by Carotliors as "distinct pliy.siolo^n'cal
individuals, representing^ resj)ecli\ <*ly I he j)at('riial
and maternal contribution to the formation of some
character or characters; and, as each can he iden-
tified, they furnish an excellent means of tracing tlie
process of segregation and recombination '' (p. 499).
It was at first assumed that eacli of tlie pairs of
chromosomes which unite in synapsis was respon.sible
for a single adult ^

character, birt ^ U

the number of d A /^/"^^i^O*^

Mendelian char- ^rp 0% Q P9 VAAj^

acters is known ^

, I , • Fig. 83. — Arjjftia simplex. Chromosomes ol

LO oe greater m ^^.g^ spermatocyte, o = accessory chromo-
Certain cases than '^o"'^ • ^ = unequal dyad. {Frum Caruthers,

the number ot

chromosomes. Fortunately, it has been f()nn<l that
the characters, instead of unch'rgoing iuih'peiKh'ut as-
sortment, mav become hnked so that certain of them
almost always occur together in the oil*sj)ring. The
relation of these facts to the constitution of the
chromosomes mav best be il hist rated bv reference to
the studies of Morgan and his stu(h'nts on the fruit -
fly, DrosopJiila. Over one hundred nuitants of tliis
species have been discovered by these in\estigators.
So far as studied, the characters of lliese Hies seem to
form three groups. "The characters in t he first gronj)
show sex-linked inheritance. They follow the sex-
chromosomes. The second group is less extensive.
Since the characters in this group arc linked to each

308 GERM-CELL CYCLE IN ANIMALS

other, we say that they He in a second chromosome.
The characters of the third group have not as yet
been so fully studied, except to show that they are
linked. We place them in the third chromosome
without any pretensions as to which of the pairs of
chromosomes are numbered II and III.

*' The arrangement of these characters in groups
is based on a general fact in regard to their behavior
in heredity, viz., A member of any group shoivs linkage
with all other members of that group, but shows inde-
pendent assortment with any member of any other
group.'* If the factors which determine these groups
of characters are situated in the chromosomes, as the
hypothesis demands, we should expect each group
to act as a unit in heredity. Occasionally, however,
the characters of a group appear to act independently,
and there must thus be an interchange of factors at
the time of synapsis. As already stated (p. 254), an
interchange of substances between chromosome pairs
during synapsis is possible and even probable. Mor-
gan explains the degree of crossing over of characters
in the following way : The factors which determine
the characters are arranged in the chromosomes in
a linear series ; those factors that are near together
will have less chance of being separated than those
that lie farther apart. The relative distances be-
tween these factors can be judged by the frequency
of interchange as determined by breeding experi-
ments. It has thus been possible to locate certain
factors in the chromosomes more or less accurately
and to predict with some degree of certainty the re-

THE GERM-PLASM TIIKOin 309

suits of hybridization. Tims If \]\v j)()siti()n of a
newly discovered factor is deteniiiiicd 1)\ compariMjii
with another particular known factor, it i\ pos^flijc
to ''calculate the results for all other known factors
in the same chromosome." ]\Ior<(an's ideas re^'ard-
ing the organization of the chromosomes coincide
with those expressed hy AVeismann in one r<'spect,
that is, they are assumed ''to have definite strnctnres
and not to be simply bags filled with a hoino^^^eneons
fluid." Wilson (191^2, p. (),S) also regards the chro-
mosomes as "componnd })o(Iies, consisting of diller-
ent constituents which undergo dilferenl modes of
segregation in different species."

Students of genetics now consider the iinhvidnal
as built up of a number of unit characters represented
in the germ-plasm by factors, and when two different
germ-plasms unite (amphimixis) the factors do not
mix, but remain uncontaminated. The germ-plasm
of offspring which develop from fertilized vfi^i^s is
supposed to consist of an assortment of factors
brought about during synapsis and rednctionas indi-
cated in Fig. 84. The factors (or genes) in the germ-
plasm occur in pairs called alleloni()ri)hs,' and one of
the pair may be regarded as donn'nant, the other re-
cessive, as, for exani})le, the yellow and green color> of
pea seeds. Thns the appearance of the individual
depends upon the character of its dominant factors.
Any attempt to account for the origin of new species

1 According to some investigators, especially in Kngland. tlie pn'senct*
of a factor should be considered one ailelonuirph and its absence jus the

contrasting factor.

310 GERM-CELL CYCLE IN ANIMALS

Fig. 84. — Diagrams illustrating the union of two stocks with paired
factors A, B, C, D, and a, b, c, d, to form pairs Aa, Bb, Cc, Dd.
Their possible recombinations are shown in the sixteen smaller
circles. {After Wilson.)

must accept these facts of heredity as a basis. If
evolution is a fact, new species must have arisen from
time to time. This may have occurred by the drop-
ping out of old factors or the addition of new factors.
There seems to be sufficient evidence that factors
are sometimes left out, but there are very few cases
of the addition of new factors. Our ideas of a pro-
gressive evolution demand the addition of new factors,
but whether this is brought about by changes within
the germ-plasm or is the result of external influences
is not known.

D. H. HILL LIBRARY

North C^rc^'i-- "•-'- -"nlUne

REFERENCES TO LITERATURE

Allen, B. M. 1906. The Origin of the Sex-cells of Chrysemys.
Anat. Aiiz. Bd. 29.

1907a. A Statistical Study of llie Sex-cells of Cliryscmys

marginata. Anat. Anz. Bd. 30.

19076. An Important Period in tlie History of the Sex-
cells of Rana pipiens. Anat. Anz. IJd. .'»1.

■ 1909. The Origin of the Sex-cells of Ainia uml Lepidos-

tcus. Anat. Record. Vol. 3.

1911. The Origin of the Sex-cells of Aniia and Lcpidos-

teus. Journ. Morph. Vol. '■I'-Z.

Altmann, R. 1890. Die Elementarorganismen und ihre
Beziehungen zu den Zellen. Leipzig.

Amma, K. 1911. Ueber die Differenzierung der Krinihahn-
zellen bei den Copepoden. Arch. Zellf. Bd. (I.

Ancel, p. 1903. Histogenese et structure de la glande herma-
phrodite d'Helix pomatia. Arch. Biol. T. ID.

Balbiani, E. G. 1864. Sur les mouvements tjui se manifestent
dans la tache germmative chez quehiues aniinaux. C. IL
Soc. Biol. Paris.

188'-2. vSur la signification des cellules polaires des in.sects.

C. R. Acad. Sci. Paris. T. !).>.

1885. Contribution a Tetude de la formation des organos

sexuelles chez les insects. Receuil. Zool. Suisse. T. 'i.

Bambeke, C. van. 1893. Contribution a riiistoire dr la con-
stitution de I'ceuf. "2. Elimination d'rlemrnts iMuh'airt-s
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1913. Chromosomes inMan.Amer. Journ. Anal. \ol. 14.

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336 GERM-CELL CYCLE IN ANIMALS

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INDEX OF AUTHORS

All numbers refer to pages. An asterisk (*) after a pnur niinilMT in-
dicates that the title of a contribution by the author will U- fmiiul uu
that page.

Allen, B. M., 32, 100, 102, 206, 311.*
Altmann, 276, 282, 311.*
Amma, 140, 163/., 216, 228, 311.*
Ancel, 195-197, 311.*

Baer, van, 192.

Balbiani, 107, 214, 229, 311.*

Baltzer, 261.

Bambeke, van, 222, 311.*

Bancroft, 21.

Bardeleben, 273, 311.*

Bartelmez, 232, 311.*

Bateson, 312.*

Beard, 100, 312.*

Beckwith, 135, 312.*

Benda, 40, 276, 277, 279, 282, 312.*

Beneden, van, 80, 82, 87, 88.

Berenberg-Gossler, 98, 100, 102,

312.*
Bessels, 118, 312.*
Bigelow, 172, 186, 225, 312.*
Blockmann, 185, 221, 225, 312.*
Bonnevie, 177, 286, 313.*
Bouin, 206, 276, 313.*
Boveri, 174 /., 184, 193, 195, 217,

230, 268, 301, 313.*
Brandt, 118, 313.*
Brauer, 83, 313.*
Brown, 3.
Browne, 285, 313.*
Buchner, 123, 140, 180, 187, 195,

222, 286, 313.*
Bunting, 87, 314.*
Buresch, 195, 199/., 226, 314.*

Calkins, 26, 314.*

Carothers, 306, 307, 314.*

Carter, 74, 314.*

Castle, 192, 314.*

Caulleiy, 195. 314.*

Champy, 195, 206, 209, 3U.»

Child, 130, 188, 297, 314.*

Chun, 184, 315.*

Cohn, 3.

Cole, 207, 209. 315.*

Conklin, 218, 232. 23.3. 234. 315.*

Correns, 302, 316.*

Cowdry, 280. 285, 315.*

Cunningham, 20!). 315.*

Debaisieux. 120. 121. 122. 315.*

Delage, 195. 315.*

Delia Vallo, 12.315.*

DemoU, 195. 202/.. 316.

Desii, 70, 76, 316.*

DickrI, 144, 316.*

I)i^M)y, 299, 316.*

Dobell, 28, 316.*

Dodds. 102-103. ;U(5.*

Dontaster, 275. 316.*

Downing. 83-85, 97. 188. 316.*

Drie.sch. 161. 231. 316*

Duesberg, 104. 23:{. 273. 2S0. tsX

31(i.*
Dujardiii, 3.
Diirmc, van. 286. 316.*
Dustin. 99. 206. 317.*

KinvnlM-rg. S2. 317.*
Kigenmanii. 100. 243. 297. 317.

•• •

337

338

INDEX OF AUTHORS

Elpatiewsky, 26, 140, 179 /., 195,

228, 317.*
Escherich, 107, 317.*
Evans, 75, 317.*

Farmer, 273.

Faure-Fremiet, 13, 279, 283, 285,

317.*
Feistmantel, 207.
Felt, 52, 317.*
Fiedler, 73, 317.*
Firket, 99, 317.*
Fischer, 12.

Flemming, 214, 273, 318.*
Fol, 186, 318.*
Foot, 123, 137, 214, 318.*
Friedmann, 207, 318.*
Frischholz, 97, 318.*
Fuchs, 163, 169, 318.*
Fujita, 186, 225, 318.*
Fuss, 100, 318.*

Galton, 290, 318.*

Gardiner, 157, 318.*

Gates, 160, 318.*

Gerhartz, 207, 318.*

Giardina, 120-122, 223, 231, 318.*

Giglios-Tos, 279, 319.*

Goette, 75, 95, 319.*

Goldschmidt, 222, 279, 286, 319.*

Gorich, 73, 319.*

Govaerts, 120, 123, 128, 319.*

Graber, 107, 319.*

Granata, 279.

Grimm, 107, 310.*

Grobben, 163, 170, 319.*

Gross, 137, 319.*

Gudematsch, 194, 319.*

Guenther, 83, 319.*

Gunthert, 121, 122, 128, 319.*

Guilliermond, 277, 319.*

Gutherz, 273, 320.*

Guyer, 272, 300, 305, 320.*

Hadzi, 83, 320.*
Haeckel, 320.*

Haecker, 36, 73, 12'4, 140, 163 /.,

184, 215, 320.*
Hallez, 112, 320.*

Hargitt, C. W., 86, 88, 95, 296, 320.*
Hargitt, G. T., 96, 320.*
Harm, 88, 89, 98, 320.*
Harman, 136, 320.*
Harmer, 161, 321.*
Hartmann, 216, 321.*
Harvey, 1.
Hasper, 104, 107, 110, 140, 218, 230,

235, 321.*
Hegner, 33, 51, 107, 140, 219, 225,

235, 286, 294, 321.*
Heider, 79.

Henking, 106, 256, 321.*
Herbst, 216, 321.*
Herold, 118, 321.*
Herrick, 215, 322.*
Hertwig, O., 82, 231, 322.*
Hertwig, R., 83, 222, 322.*
Heymons, 186, 194, 322.*
His, 231.

Hodge, 214, 322.*
Hogue, 179, 322.*
Holmes, 137, 322.*
Hooke, 2, 207, 322.*
Hoven, 277, 280, 322.*

Ijima, 76.

Ischikawa, 86, 163, 170, 322.*

Jager, 291, 322.*
Janda, 297, 322.*
Janssens, 254, 255, 322.*
Jarvis, 100, 322.*
Jenkinson, 50, 232, 323.*
Jennings, 186, 225, 323.*
Jordan, 214, 288, 323.*
Jorgensen, 72, 74, 78, 323.*

Kahle, 51, 107, 140, 230, 235, 294,

323.*
Kellicott, 50, 323.*
Kellogg, 118, 323.*
King, 195, 206, 208, 323.*

INDEX OF AUTHORS

339

Kite, 6, 323.*

Kleinenberg, 80, 82, 83, 161, 323.*

Knappe, 208, 323.*

Kolliker, 373.

Koltzoff, 323.*

Korotneff, 83, 323.*

Korschelt, 79, 324.*

Kossel, 8.

Kowalevsky, 107, 324.*

Kruger, 195, 267, 324.*

Kiihn, 140, 163/., 225, 236, 324.*

Kulesch, 105, 324.*

Kunstler, 276, 324.*

Kuschakewitsch, 100, 195, 206, 324.*

Lams, 286, 324.*

Lang, 157, 324.*

La Valette St. George, 207, 276, 324.*

Lecaillon, 109, 111, 324.*

Lewis, 281, 324.*

Leuckart, 51, 107, 324.*

Levene, 11.

Leydig, 82, 325.*

Lieberkiihn, 73, 325.*

Lillie, 188, 232, 234, 325.*

Loeb, 13, 21, 325.*

Loewenthal, 222, 325.*

Loyez, 286, 325.*

Lubarsch, 130, 325.*

Lubosch, 214, 325.*

Maas, 73, 76, 78, 325.*
McClendon, 172, 185, 325.*
McClung, 256, 325.*
McGregor, 134, 135, 326.*
Malpighi, 3.
Mangan, 187, 326.*
Marchal, 161, 326.*
Marcora, 280, 326.*
Marshall, A, 326.*
Marshall, W., 75, 326.*
Marshall, W. M., 222, 326.*
Maupas, 267, 326.*
Mayer, 283.
Megusar, 113, 326.*
Meinert, 51, 326.*

Mendel, .302/., .326.*
Metschnikolf. 51, 107, 1H.3, 221. .S2(J.»
Moves, 134, 216. 244. 200, 2K4. 2H7.

326.*
Meyer. 170. 327.*
Mimiiin, 70, 72. 327.*
Mohl, von, 3.
Montgomery, 129. l.U, 195, 214.

241, 285. 297, .300, 327.*
Moore, 273, 328.*
Morgan, 192, 232, 255. 2(55, 3<)2.

307, 309, 328.*
Morse, 1.37, 244, 328.*
Muller, F., 1J)5. 328.*
Muller, K.. 77, 80, .328.*
Muller-Cale. 172, 328.*
Mulsow. 256, 257.
Munson, 226, 329.*

Nachtsheim,143, 145, 205, 200.;{^t».'
Xoack, 107, 109. Ill, 225. 2,35. 329.*
Nussbamn, 83, 100, 291, 329.'

Ognew, 208, 329.*
Okkeberg, 209, 329.*
Ostwald, 9.
Owen, 290, 329.*

Patterson, 157, 161. .329.*
Pauleke, 120, 122, 222, 329.*
Paulmier, 250. 329.*
Payne, 138, .329.*
Pei-seneer, 195, 329.*
Petrunkewitseh. 143. 145. 205, 3.3<».*
PBiiger, 205. 231.
Pick. 194, .3.30.*
Prenant, 279, 2S3. .330.*
Preusse. 137, 330.*
Punnet, 330.*

Rath, v(.m. 134, i:i5. 3.30.*

Regaud, 279, 282. 330.*

Rhode. 218. ,330.*

Richards, 1.3(5. 3.30.*

Ritter. 107, lOS. 22!>. 235. ;U0.'

RolxTtson. 101. .3.3(1.*

Robin, 107, 330.*

340

INDEX OF AUTHORS

Rosel V. Rosenhoff, 72, 331.*

Rosner, 161, 331.*

Roux, 141.

Rubaschkin, 98, 100, 103, 226, 283,

331.*
Ruckert, 99, 331.*
Russo, 286, 331.*

Samassa, 170, 172, 331.*

Sauerbeck, 194, 331.*

Schapitz, 100, 331.*

Schaxel, 214, 331.*

Schaffer, 283.

Schleip, 195, 268, 331.*

Schleiden, 3.

Schmidt-Marcel, 205, 207, 331.*

Schmiedeberg, 12.

Schneider, 83, 331.*

Schonemund, 194, 331.*

Schonfeld, 237, 331.*

Schreiner, 209, 332.*

Schulze, 76, 80, 193, 332.*

Schwann, 3.

Selenka, 157, 332.*

Semon, 206, 332.*

Siebolt, von, 193, 332.*

Silvestri, 143, 145, 215, 332.*

Simon, 194, 332.*

Smallwood, 87, 98, 332.*

Spooner, 234.

Steudel, 12.

Stevens, 140,180, 195,228, 256, 332.*

Strasburger, 214, 332.*

Stricht, van der, 188, 286, 333.*

Strobell, 123, 137, 214.

Stuhlmann, 221, 333.*

Suckow, 118, 333.*

Surface, 157, 333.*

Sutton, 305, 306, 333.*

Swarezewsky, 26, 333.*

Swift, 33, 103, 226, 333.*

Tannreuther, 83, 333.*
Tennent, 261.
Thallowitz, 88, 333.*
Trembley, 82, 333.*

Tschaschkin, 98, 102, 226, 302, 333.*
Tschermak, 333.*

Uffreduzzi, 194, 333.*

Vander Stricht, 187, 333.*
Varenne, 82, 333.*
Vejdovsky, 334.*
Voeltzkow, 107, 334.*
Vollmer, 172, 334.*
Voss, von, 204, 334.*
Vries, de, 302, 334.*

Wager. 83, 334.*

Wagner, 51, 334.*

Waldeyer, 98, 130, 334.*

Walker, 159, 334.*

WassiUeff, 286.

Weismann, 25, 82, 88, 97, 107, 113,

144, 296, 309, 334.*
Weltner, 73, 75, 77, 334.*
Wheeler, 33, 100, 109, 144, 157,

185, 193, 335.*
Whitman, 231, 335.*
Wieman, 124, 138, 225, 273, 335.*
Wierzejski, 75, 186, 225, 335.*
Wijhe, van, 99, 335.*
Wilcox, 273, 335.*
Wildman, 279, 335.*
Wilke, 285, 336.*
Wilson, E. B., 4, 21, 133, 224, 232,

250, 301, 309. 336.*
Wilson, H. v., 75, 77, 80, 336.*
Winiwarter, 129, 132, 251, 273, 336.*
Winter, de, 119, 120.
Woods, 100, 336.*
Wolff, 2.
Wulfert, 89, 98, 336.*

Youngman, 207.
Yung, 207, 336.*

Zamik, 195, 269, 336.*
Zeigler, 237.
Zeleny, 232, 336.*
Zoja, 287, 336.*
Zykoff, 75, 336.*

INDEX OF SUBJECTS

All numbers refer to pages. Words in iUilics are names <.f famili«-s.
genera, species, or of higher divisions. Xunibers in thick tyiH* are num-
bers of pages on which there are figures.

Aborting spindle, 157.

Accessory chromosome, 134, 202.

Acidophile, 11.

Actinospherium, 222.

Ageniaspis, 146.

Allelomorph, 309.

Alternation of generations, 23.

Alveolar structure of protoplasm,
4.

Amoebocyte, 71, 73, 79.

Amia, 32, 33.

Amitosis, 13-14, 133-139, 250.

Amphiaster, 15.

Amphibia, amitosis, 134-135; her-
maphroditism, 205 ff.

Amphimixis, 309.

Amphiuma, 135.

Amyloplastid, 7.

Anaphase, 15, 16.

Anello cromatico, 121, 123, 223.

Animal pole, 20.

Aptera, life cycle, 22.

Arcella, 26.

Archaeocyte, 70-73.

Archoplasm, 5, 7.

Arenicola, 188.

Armadillo, polyembryony in, 161.

Arphia, 307.

Arfhropoda, 212.

Ascaris, 122, 174 /., 217 /., 230,
241, 301 ; maturation in, 2G1,
263 ; mitochondria in, 284.

Asexual larvae, 149.

Asplanchna, 186, 225.

Aster, 16.

Asterias, 6.

Attraction-sj)here, 5, 7, ii7.

Amelia, 183.

Aussenkornelu-n. 164. 213. 210. 228.

Axolotl, 159, 208.

Bacteria, 4, 186-187.

Basophile, 1 1 .

Bat, 188.

Besondere Kiirpcr. 180. isj /T .

213, 228, 23!).
Bidder's organ, 207.
Binary fission. 17.
Binuclearity hy|)othesi.s, 27.
Bioblast, 276.
Bivalent chrDmosomes. 44.
lilastotomy, 161.
Bryozoa, 161.
Budding, 17, 22, 23. 69. 161, 297.

Calligrapinu 109. 111. 2.30.

Calliphora, 107. 111/.. 235.

Camponotus, ^-^l .

Canfliocamptu.t, 165.

Cat, 187.

Cell, 2-16; definition. 3; divisit»n.

13-16; lineage, 29; shapr. 4;

size. 4; tiieory, 3.
( entrifuged egg.s, 173.
C'entro.sonu'. 6, 7. 14, 15. 164. 169.

237, 238.
Cerchratiihis, 232.
Ct'stoiUi. \'M\ 137.

341

342

INDEX OF SUBJECTS

Chcetognatha, 212.

Characters, dominant, 303 ; linked,
307 ; recessive, 303 ; unit 303.

Chiasmatype theory, 254.

Chick, 33^ 100, 103, 227, 281.

Chironomus, 108-109, 110, 224, 229,
235.

Chloroplastid, 7.

Cholesterin, 8, 12, 13.

Chorion, 113.

Chondriodierese, 279.

Chondriokont, 279.

Chondriosome, 7, 102, 103, 168,
227 /., 275, 277.

Chondriotaxis, 279.

Chromatin, 5, 7, 11-12; as germ-
plasm, 299 ; as keimbahn-deter-
minants, 211 ^.

Chromatin-diminution, 47, 56, 57,
139-141, 174/., 217/., 249.

Chromatin-nucleolus, 5, 7.

Chromidia, 26, 123, 168, 221 /.,
279.

Chromidial net, 26.

Chromosome, 6, 7, 14, 15, 243, 299 ;
accessory, 106 ; cycle, 245-275 ;
diploid, 43; division, 248; in
fertilization, 49; haploid, 43;
individuality, 255 ; in man,
272 /. ; and Mendelism, 305 ;
number, 246; from nucleolus,
214 ; in parthenogenesis, 246 ;
univalent, 249.

Chrysemys, 32.

ChrysomelidcB, 109.

Ciona, 192.

Cladocera, 163/.

Clathrina, 70.

Clava, 88, 135.

Cleavage, 29, 115.

Cockroach, 194.

Coelenterata, 80-98, 212.

Coleoptera, 109-143.

Colloid, 9.

Colony, 17.

Compsiliira, 107, 109.
Conjugation, 17.
Copepoda, 165/
Copidosoma, 146/
Copulationszelle, 163.
Corps enigmatique, 187.
Crepidula, 218.
Crustacea, 163-173.
Crystalloid, 9.

Cyclops, 124, 164/, 228, 247.
Cyviatog aster, 100.
Cynthia, 233, 280.
Cyst formation, 125-129.
Cytomicrosome, 276.
Cytoplasm, 6, 143, 179, 224 /.,
300/

Daphnidce, 163.
Death, natural, 25.
Determination of sex, 118.
Determiner, 302.
Diaptomus, 165.
Differentiation, 76, 141-143.
Dioecious, 18, 190.
Diploid, chromosomes, 248.
Diplotene, 252.
Diptera, 107.
Dispermic, 177, 178.
Dominance, 303.
Dotterplatte, 109, 115, 225, 235.
Drosophila, 307/
Dyad, 45, 46, 306, 307.
Dytisciis, 120-124, 121, 223.
Dzierzon theory, 143.

Earthworm, 161, 190, 191.

Eclectosome, 279.

Ectosome, 166, 167 ff., 213, 237.

Egg, 19, 20.

Encyrtus, 145.

Enzyme, 300.

Ephydatia, 75.

Epigenesis, 2, 243.

Ergastoplasma, 276.

Eudendrium, 86.

IXDKX OF SLIUKCTS

Euschisiis, 281.
Evolution, 310.

Factor, 302, 309.
Female sex, 18.

Fertilization, 44, 47-49, 2oG, /.
Fission, 22.

Frog, hermaphroditism in. 20.5 (f.
Fusion, of chromosomes, 234; (tf
oocytes, 152. 1.5.)^.

Gel, 5, 6, 9.

Gemmule, 18, 74-75, 76, 79.

Genes, 302, 309, 310.

Genetics, 309.

Genetic-continuity of chromosomes.

255.
Germ cell, 19-22, 101, v.s. somatic

cell, 290-297.
Germ-cell cycle, 28-49.
Germinal continuity. 292.
Germinal epithelium the(»ry. 98.
Germinal localization, 231.
Germinal spot, 214.
Germinal vesicle, 19, 20, 54.
Germ-plasm, in A.scariji, 177, in

Hijdra, 8.3-85 ; in Miastnr, 29.3 ; in

polyembryony, 102; in sponj^rs.

80.
Germ-plasm theory. 290-310.
Gonochorism, 18, 191.
Gonocyte, 71, 73.
Gonothyrcpa, 89.
Gonotome theory, 97.
Grafdla, 157.
Gryllus, 123, 244.
Guinea-pig, 102. 103. 104. 227.
Gynandromorj)h, 1!>3 19i.

Haploid, 247.
Hauptnudeolus, 214.
Helix, 195. 190/.. 220.
Hemipfcra, amitosis in. 137.
Hermaphrodite. 18. 189-210. 20!).
Ilclcrucopc, 105.

n«'teroty|)ic mitosis, 40. 252. f.'i.'i.

Ilrterozygou.s, iJOi.

Ilonuilogou.s chroinoHomrA, 4J3,

Hornotypic mittwis, 40.

Iloinozyguus. ;{<»;{.

Iloncy-lM-i'. 14.3 I Vl. 2r,i /.. 266.

Hyaloplasm, 4. 6.

Ili/dra. 82-85. 159.

Ili/ilrartinia, 87.

Hy<lroid. lif.- cyclr at. 23.

Illfdrophilus, \ 13.

Ilydmzoa, 85-98.

Hymcnopieru, 14.3-103, 221. 235.

chrumosomrs.

IdiochroMi.itin. 28.
lndi\'i(lualit>- of

255.
Interkinesis, 299.
Isotropism, 231.

Jellv-fish. 23.

KarviK'hondria, 279.
Karyokintvsis, 1.3. 1 1. 15
Kar\dlyinpli. (!.
Karyosomr. 6. 7. 213.
Kciiuhalm. in .tji/uorf,!, ]s:\. 184,

Aiiipfiihia, 200 Jf.; ( hidoccra,

103 /. ; Coprpixlii. 105 ff.\

insiM-t-s. 100-103; neinat«Hli*«.

174 17!»; Sagilla. 179/.
Keinil».din-drt<Tininants. 19. 211-

2U. 2!»«i. .301 ; ^;.u.->i>. 211 234;

l.xalization. 2;U 2KI; futr. 240-

244.
K«-iinl»ahn(hn»midirn. 22.3
K«'irni)ahniliron»atiii. 152/. 223.
KrimLalmplasina. 108. 110. 115.

23(1. 2:U.
K«-iml>ahn7.rlle. 1(>1.
Krimlh-ik. 2U.
K.-iinhautl.lastrm. 113. lU
Kriinsljil ti". 95.
K.imwtiUt. 108. 110. 11.'.. ^^3.'.
Keimzonc, 95.

344

INDEX OF SUBJECTS

Kinetochromidia, 214.
Kinoplasm, 214.

Lamprey, 100, 209.

Larva, 23.

Lecithin, 8, 12.

Lejpas, 172, 225.

Lepidoptera, 118.

Lepidosteus, 32, 33, 101.

Leptinotarsa, 37-41, 111, 125-129,

138-139.
Leptotene, 251, 252.
Life cycles, 22^.
Linin, 5, 7.

Linked characters, 307.
Locust, 23.
Lophius, 102.
LygoBus, 259.
Lymncea, 192.

Macrogamete, 27.

Male, 18.

Man, chromosomes of, 272 ff. ;

hermaphroditism in, 194.
Maturation, 41-47, 129, 256 #.
Medusa, 23.
Mesostoma, 204.
Metabolism, and sex, 275 ; and

Keimbahn-determinants, 228.
Metagenesis, 23.
Metanucleolus, 183, 215.
Metaplasm, 5, 7, 8.
Metaphase, 15, 16.
Metazoa, 1, 18.
Miastor, 51-68, 107, 217 /., 235,

293-294.
Microgamete, 27.
Microsome, 6.

Middle piece, of sperm, 21, 216.
Migration, of germ cells, 31-34,

101-102, 116, 226.
Mitochondria, 5, 13, 39, 40, 226/.,

275-289 ; methods, 282-283 ;

Ascaris, 284; chick, 278; di^ii-

sion of, 281, 284 ; function of.

286 /. ; in living cells, 280, 281 ;

in plants, 277, 280 ; reduction of,

285; and sex, 285.
Mitosis, 13, 14-16.
Mitrocoma, 183.
Mixochromosomes, 251.
Moina, 163.
Mollusk, 185, 191.
Monad, chromosome, 45, 46.
Moniezia, 136, 297.
Monoecious, 18, 191.
Monospermy, 48.
Mosaic development, 233.
Moulting, 23.
Musca, 107.
Myofibril, 280.
Myxine, 209.
Myzostoma, 37, 185, 193.

Nahrzellenkem, 170.

Nebenkem, 203, 221, 285.

Nebennucleolus, 214.

Nematodes, chromosomes of, 267 ff.

Nepa, 137.

Neratina, 186, 225.

Netzapparat, 103, 104.

Neurofibril, 280.

Nuclear sap, 6.

Nucleic acid, 11.

Nuclein, 11.

Nucleolo of Silvestri, 145 ff.

Nucleolus, 5, 6, 13, 167, 213/.

Nucleoprotein, 8, 11.

Nucleus, 3, 13-16.

Nurse cells, 35-36, 53, 119-121, 150,

151, 201, 202.
Nutritive substances, 225 ff.

(Enothera, 160.
Oncopeltus, 261, 262.
Oocyte, 38, 39, 40-41.
Oogenesis, 42, 256 /.
Oopfhora, 145, 146.
Ophryotrocha, 37.
Opossum, 288.

IXDKX OF SUBJECTS

345

Organ-forniinp substances, 233.
Organization of rgg, l!), 2«J, iiH Jf.
Oxyphils, 11,

Pachytene, 252.
Paedogenesis, 18, 52.
Paracopulations/A'lle, 212, 225.
Paramecium, 27.
Paranucleus, 1(53.
Paraplasm, 7.
Parasitism, 191-192.
Parasynapsis, 254.
Parthenogenesis, 18, 47, 145, 240,

265.
Pea, 302, 303.
Peden, 191.
Pennaria, 87.
Peripatus, 285, 288.
Petromyzoru 33.
Phalliisia, 233.
Phosphatid, 8, 12.
Phylloxera, 2G5 /.
Physa, 18G, 225.
Pig, 194.
Planoccra, 157.
Pla7iorbi.s\ ISO.
Plasmodia, artificial, 77-7S.
Plasmosome, 5, 7, 102, 103, 213.
Plastid, 5, 7.
Plastochondria, 279.
Plastokonta. 279.
Plastosome, 7. 244, 275. 279.
Polar body, 47, 143-144.
Polares Plasma (see pole-plasm).
Polarity, 19, 107. \n, 179, 231/.
Pole-cell, 110. 111. 117.
Pole-disc, 1(>!>, 114. 117. U2. Jl!).

225, 229. 2.3.-).
Pole-plasm, 53 55. 228, 230, 235,

294-295.
Polistes, i'i.'i.
Polychoerus, 1 57 .
PolyembryouN , 145/., Kil.
Polyp, 23!
Polyphemus, 170/.. 230.

I Polyspenny, 48. 115. 299.

Porifcni, iW) Jf.
I Pol.ito JMftl*' {gee L< ptinolartaK

Prrl)l,i.sto«l«Tiiiic tni<lfi. 114.

Prt'«lrlfrniinali<»n. 2.

Preformation. 2. 243.

Prochrom(».some, 299.

Progenninativr c«-ll. 190. 197.

Proiiiorplioltigy. l!).

I'riiplia.sc. 14.

Protandry. 193.

Protein. 8. 10.

Protenor, 12.3. 258.

Pr(»t(.gyny. 192-193.

Protoplasm. 3 13.

Protozoa, 1. 17. 25.

Pteropod. 2<»;». 271.

Pupa. 23.

Pyrrhocori.t, 25(5.

Rana, 32.

Recessive charjicter. .304.

He<lu(lion of clintmosomc*. 4.3,

253.
Regeneration. 79 SO. 297.
RepnKlu<tion. 17 is.
Hot if era. ISO.
Hhalxliti.-*, 207. 270.
Ri(lifung.sropuIati«)nNkem. 1 U

Sagitta. 179/.. 195. 228.

Salatiiandra, 134.

SarciMJe, 3.

SrorpcFua, 222.

Sea urchin. 210.

Secondary .sex «-hnmrters. ISO.

Segregation of germ <flls. 21*.

Scif-copiiiation. l!»2.

Si'lf-fertilizjition. 192.

S<Tt..li cell. .35. 129-1.33.

Sex. 18. 189.

Sc\ chromosonie. 255/.

S'X detennination, 271.

Sol. 5. 0. 9.

Sorite. 70, 79.

346

INDEX OF SUBJECTS

Spermatogenesis, 42, 256 ff.
Spermatogonia, 127.
Spermatozoon, 19-22, 48.
Spherule, 276.
Spireme, 14, 15.
Spongilla, 73.
Spongioplasm, 4, 5.
Sporulation, 17.
Squash bug, 256.
Starfish, 6.
Statoblast, 18.
Statocyte, 70, 71, 73.
Stem-cell, 175.

Stone-fly, hermaphroditic, 194.
Synapsis, 44, 122, 250/., 305.
Synaptene, 251, 252.
Synizesis, 43, 237, 251, 252.

TcBTiia, 136, 137.
Telophase, 15, 16.
Telosynapsis, 254.
Testis, 41.
Tethya, 70, 76, 79.

Tetrad, 44, 45.

Tipulides, 107.

Toad, hermaphroditic, 207-208.

Tokocyte, 71, 73, 79.

Trophochromatin, 28.

Unit character, 304.
Uterine spindle, 157.

Vacuole, 5, 8.
Vegetative pole, 20.
Vertebrate, 32, 95-105, 212.
Vitelline membrane, 113, 114,
Vitellophag, 114.

X-chromosome, 255 ff., 264, 299.

Y-chromosome, 259 ff., 264.
Yolk, in germ cells, 101, 224.
Yolk nucleus, 19, 226, 285.

Zygosome, 251.
Zygote, 1, 48.

'T^HE following pages contain advertisements of
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An Introduction to Zoology

By
ROBERT W. HEGNER, Ph.D.

Assistant Professor of Zoology in the University of Michigan

A TEXT-BOOK INTENDED FOR THE USE OF STUDENTS
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Genetics. An Introduction to the Study of Heredity
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An Outline of
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The First Principles of Heredity

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CONTENTS

Introduction — Evolution in General. Palacontol<ijr>'.

Section I — Inorganic Evolution. Geographical Dinribinion

The P: volution of Matter. Part II Theories of Kvolution.

Section II — Organic Evolution Section III — Suj>«ior • •■ • i\ 1. .•..-.

Part i - The Facts of Evolution. Social I

Morphology. Conclusion — The Formula of Evoluiim.

Embryology. The Philowphy of Chanec

Classification.

THE MACMIM.AN COMP.ANV

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