Stages in the development of Ictalurus nebulosus

6

X

STAGES IN THE
DEVELOPMENT OF

Ictalurus
nebulosus I

5); PHILIP B. ARMSTRONG, M.D.

College of Medicine at Syracuse
State University of New York
and Marine Biological Laboratory
Woods Hole, Massachusetts

Illustrated by

JULIA SWOPE CHILD

Marine Biological Laboratory
Woods Hole, Massachusetts

1962
SYRACUSE UNIVERSITY PRESS

university of

Connecticut

libraries

UNIVERSITY of

CONNECTICUT

LIBRARY

K

^«St OaA

\
\

B

at
a

J,

STAGES IN THE
DEVELOPMENT OF

Idalurus nebuhsus

By PHILIP B. ARMSTRONG, M.D.

College of Medicine at Syracuse
State University of New York
and Marine Biological Laboratory
Woods Hole, Massachusetts

Illustrated by

JULIA SWOPE CHILD

Marine Biological Laboratory
Woods Hole, Massachusetts

SYRACUSE UNIVERSITY PRESS SjQj 1962

PRESS

Air

Library of Congress Catalog Card 63-11037

Copyright © 1962 by Syracuse University Press

Syracuse, New York

ALL RIGHTS RESERVED

The development and publication of this normal series was sup-
ported by U.S. Public Health Service grants B-643 and GM-10758.

MANUFACTURED IN THE UNITED STATES OF AMERICA

STAGES IN THE DEVELOPMENT OF
ICTALURUS NEBULOSUS

Ictalurus nebulosus, the brown bullhead or
horned pout, is generally distributed through-
out the Eastern and Central United States from
Maine and the Dakotas on the north into the
Gulf States on the south. It has been introduced
successfully into streams and ponds on the West
Coast and into the waters of several European
and Far Eastern countries.

Under favorable conditions adult bullheads
attain a length of a foot or more with occasional
specimens 18 inches in length weighing 3 to 4
pounds. When crowded they are much smaller.
Ictalurus nebulosus is very tenacious of life,
surviving under conditions which will not sup-
port other native fishes. It is commonly found
in small ponds, lakes, and sluggish streams and
does well even in a muddy habitat. The adults
are frequently caught on a drop-line during the
daytime but are more active at night. The eyes
are small but the olfactory and cutaneous senses
are highly developed. They are omnivorous,
the diet depending on what is available. The
adults are readily collected in a baited minnow
trap with an adequate entrance and with reason-
ble care will survive well in aquaria.

The reproductive habits of Ictalurus nebu-
losus have been observed in both natural sur-
roundings and in aquaria. The eggs are usually
deposited in a prepared nest, an open shallow
excavation among weeds, or a burrow excavated
by the adults under the roots of a water plant
which will support the roof of the nest. A com-
mon site for such a burrowed nest is horizontally
for as much as three feet into the bank of the
stream or pond. The bullhead resembles the
wren in selecting a nesting site, taking advantage
of protected locations in pails, stovepipes, rub-
ber tires, cinder blocks— or under boards, stones,
or other articles thrown by chance into the

water. Rarely are the eggs laid in an open site
without preparation. Depending on the nature
of the nest, the floor is invariably some firm
material— coarse sand, gravel, or stone. When
the nest is found in the muddy bottom of a
pond, it is located where the mud is a shallow
layer on a sandy or gravel bottom to which the
nest is excavated. Drainage tiles provide attrac-
tive nesting sites for bullheads. Tiles are par-
ticularly useful in ponds supporting a heavy
growth of weeds in which it is difficult to locate
the natural nesting sites. The tiles should be
distributed separately around the margin of the
pond in two or three feet of water, their loca-
tions indicated by a suitable marker.

The eggs are adhesive when laid and form a
spongy pale yellow mass which adheres to the
floor of the nest. Each egg through its chorionic
membrane has adhesion discs with several adja-
cent membranes which sometimes show up on
the surface of the chorionic membranes when
they are torn from each other (see figure of
Stage 20) . The micropyle is located as the center
of a stellate structure seen in some membranes
(Stage 1 et al.). When the eggs are laid in indi-
vidual drainage tiles it is frequently necessary
to gently scrape the egg mass free of the tile to
which it adheres. If the floor of the nest is sand
or gravel, grains of sand and small pebbles are
found adhering to the egg mass. There are from
several hundred up to a thousand and more eggs
in a single egg mass. The adults, particularly
the male, guard the eggs. Frequently an ob-
server can flush the adult off a nest by wading
in a pond a few yards off shore, locating as close-
ly as possible the point at the bank from which
the adult emerges. Close inspection of the bank
at this point will usually reveal the opening of
the nest. The collector can reach into the bur-

STAGES IN THE DEVELOPMENT

row, and by careful exploration the egg mass,
which is soft and yielding, can be obtained. It
must be handled gently. (Sometimes a second
fish is flushed from the nest and rushes past the
collector's arm as he attempts to find the egg
mass.)

The adults work over the eggs very actively
(Breder, 1935), fanning them with their fins,
taking them in their mouths and ejecting them,
and in general handling them rather roughly.
Apparently almost constant agitation is re-
quired to assure normal development of the
eggs, since eggs removed to still water usually
will die within a day or so. Two simple methods
were devised for handling the eggs; either will
usually give normal development of the major
share of the eggs of a cluster. The cluster may be
placed in a seven-inch finger bowl in tap water
flowing from a rubber tube of which the lower
three inches rests on the bottom of the finger
bowl directed to give a swirling flow of the
water in the bowl. There should be a flow of
water adequate to agitate the eggs slightly but
not to carry them with the current. A constant
flow of water can best be obtained through a
petcock without a washer. In a washered outlet,
the flow reduces as the washer swells. The sec-
ond method is illustrated in Figure 1, which
shows a strainer to which a metal tube is
soldered in such a position that the water flows
from below through the egg mass resting in the
strainer. The egg masses should be examined
at least twice a day and any dead eggs removed,
since these quickly develop mold which extends
to the adjacent eggs and kills them.

The laying season starts about the same time
in central New York State and on Cape Cod,
Massachusetts— i.e., about the middle of May.
The latest that newly laid eggs were found on
Cape Cod was July 28. Adults in aquaria will
lay spontaneously, with the opening and closing
dates of the spawning somewhat later than with
fish in the wild. Females early in the winter
carry eggs which in appearance and size are close

to maturity. We induced spawning and obtained
fertilized eggs out of season by injecting into a
pair of bullheads crushed pituitaries from ripe
female alewives, but additional work must be
done to standardize the procedures if uniform
success is to be achieved.

The eggs measure about 3 mm. in diameter.
Those from the same egg mass are remarkably
uniform in size but there are differences in size
from one egg mass to another. The subchor ionic
space is wide and accounts for about one-third
of the diameter of the egg. The chorionic mem-
brane is transparent, naked, and relatively thin
for the size of the egg. The membrane can be
removed readily without injury to the develop-
ing embryo merely by tearing the membrane
with fine forceps. After the blastopore is closed,
the embryos survive in running tap water with-
out any special treatment; in fact, they are less
susceptible to death than if left in the mem-
branes where the mechanics of aeration are
more difficult. The embryos hatch spontaneous-
ly in Stage 43 and will swim actively on the floor
of their container. In the earlier swimming
stages, the embryos can be kept in an open
dish with a gentle stream of water circulating
through the dish. In the more advanced stages,
they become negatively geotropic and are lost
from an open dish, but they can be kept in a
battery jar closed with a strainer fitted with a
tube through which water can be introduced
from above (Figure 2) . The strainer must be

FIGURE 1

FIGURE 2

OF ICTALURUS NEBULOSUS

of such a size as to fit snugly in the top of the
battery jar.

The embryos leave the nests and seek food
when the yolk has been absorbed. For a few
days they swim as a compact school in shallow
water guarded by the adults which swim about
below the school. After a few days the school
breaks up and the larvae disappear into deeper
water.

To identify stages of the embryos, it is ad-
vantageous to examine them with transmitted
and with reflected light. Light transmitted
through the embryo from a white substage mir-
ror is especially effective in bringing out the
details of pigmentation. The developing blood
vessels and circulation are revealed best by the
use of a regular substage mirror. In lighting the
embryo from above, strong illumination, such
as that from a zirconium lamp, is required to
bring out the finer details. Also all available
features of the developing embryos, both mor-
phological and physiological, should be taken
^ into consideration for accurate identification of

stages.

The embryos in the same egg mass, if properly
handled, show remarkable synchrony in their
development. However, minor but perceptible
variations do occur from mass to mass, possibly
because the development of the various features
of the embryo do not depend on a single com-
mon factor. Because of this, embryos will be
included in the same stage in spite of some such
minor variation from the pertinent illustration.
In the illustrations the embryos are magnified
1 7 times.

Some of the special features of development
are described below.

CLEAVAGE

Stages 1 through 7

Cleavage in the bullhead egg is meroblastic

and meridional as is characteristic of the teleosts,

the cleavages being limited to the blastodisc, not

involving the yolk. The diameter of the early

blastodisc is only about one-eighth the circum-
ference of the egg. The successive cleavages fol-
low each other at 35 to 40 minute intervals and
extend well through the disc. The blastomeres
formed at each cleavage are of equal size and
the cell divisions at any one cleavage are closely
synchronous. From the margins of the disc the
protoplasm thins out abruptly and extends
around the yolk as a thin continuous layer, as
in Fundalus. There is no defined periblastic
ridge such as occurs in Serranus (Wilson 1891).
Cleavage continues to be meridional in the bull-
head at least through the 32-cell stage and pos-
sibly through the 64-cell stage, but it is quite
probable that in this latter cleavage some of the
central cells divide horizontally.

BLASTULA

Stages 8 through 15
The subchorionic (perivitelline) space in the
bullhead egg is wide and the chorionic mem-
brane thin and yielding. This permits the for-
mation of a high blastula which is not seen in
those teleosts with narrow subchorionic spaces
such as Trutla, Fundulus, and Opsanus. The
early blastulae (Stages 8 and 9) very much re-
semble the morula stage of some of the inverte-
brates, the individual cells being large enough
to give the morula effect. The blastula remains
high (Stages 10 and 11) through additional cell
divisions and then gradually flattens out (Stages
12 to 15) with the formation of the blastocoele
which is apparent in the living egg (Stage 15).
The high blastula (Stage 1 1) has a diameter even
less than that of the one-cell stage, whereas the
flattened blastula (Stage 15) has a diameter more
than half again that of the one-cell stage. As the
blastula begins to flatten out in Stage 12, there
is the beginning formation of marginal periblast
cells, the process continuing through Stage 13,
resulting in the formation of roughly two rows
of such cells placed at a distance from the blasto-
derm. These periblast cells disappear later as
the blastoderm extends over the yolk.

STAGES IN THE DEVELOPMENT

GASTRULA
Stages 1 6 through 1 9
When the blastoderm has expanded to cover
about one-fourth of the yolk, there develops,
through mitotic activity, an accumulation of
cells at its margin (Stage 16) resulting in the
formation of the germ ring. In Stage 17 the
anterior lip of the blastopore is seen as an extra
accumulation of cells forming the embryonic
shield which becomes more prominent in Stages
18 and 19, seen best as viewed on the horizon
of the egg.

NEURULA
Stages 20 through 24
During Stages 20 to 24 the blastoderm con-
tinues to extend over the yolk. This culminates
in the closure of the blastopore late in Stage 24.
The blastoderm covers about half the yolk in
Stage 20. The embryonic shield in this stage
bears considerable resemblance to the open
neural plate of the amphibia. However, true
concrescence as in the amphibia does not occur.
There is a median longitudinal cellular pro-
liferation with a narrowing of the neural plate
resulting in the formation of a definitive pre-
somitic embryonic axis (Stage 21). After the
formation of the embryonic axis neurulation
proceeds rapidly. In Stage 22 the main divisions
of the brain can be discerned, the optic vesicle
is rudimentary, and the first somites appear.
There follows continued growth and further
differentiation of the divisions of the brain and
of the optic vesicle and a continuing addition of
somites in a caudal direction. In Stage 24 there
is a bilateral accumulation of cells, the rudiment
of the otic vesicle, at the level of the anterior
hind brain.

TAIL BUD EMBRYO

Stages 25 and 26
In Stage 25 (ca. 14 somites) the tail bud is well
denned. Around its periphery, the ectoderm is
directly continuous with that of the yolk sac.

The blastopore is closed, but its location is
marked by a slight depression in the yolk. Al-
though the main divisions of the brain can be
discerned, the brain ventricles are not yet
apparent.

With continued growth, the tip of the tail
bud develops as a projection free of ectodermal
continuity with the yolk sac as in Stage 26 (ca. 17
somites). In this stage the pericardial sac is
developing and the heart rudiment forms as a
small bit of tissue in the pericardial sac under
the head. However, the heart rudiment does
not beat when it first forms.

ORGANODIFFERENTIATION

Stages 27 through 53

This period includes development extending
through to the complete absorption of the yolk.
The embryos prior to Stage 43, when hatching
normally occurs, were removed from the chori-
onic membranes for clearer observation and
illustration.

In Stage 27, the heartbeat is just discernible C
in transmitted light. It is faint but regular.
Also, slow weak contractions of the anterior
somites result on gross mechanical stimulation
of the embryos. No cross striations are seen in
the developing muscle.

In Stage 28, the heartbeat is still faint but
readily visible. Also there are spontaneous con-
tractions of the anterior somitic muscle. The
first formation of the brain ventricles is apparent
in the region of the hind brain indicated by the
formation of the roof of the fourth ventricle,
seen in the lateral view of the embryo.

In Stage 29, muscle contractility is still limited
to the anterior somites. There has been an in-
crease in the size of the heart, but the circulation
of the blood is not established until Stage 30
and at first is sluggish and principally intraem-
bryonic. Sometimes at this stage there is seen
in the lower part of the otocyst an aggregation
of minute granules, the earliest indication of
the otoliths.

OF ICTALURUS NEBULOSUS

In Stage 31 a sluggish circulation is present
over the surface of the yolk sac, with the cells
of the communicating blood islands moving
slowly toward the venous end of the heart.
There are now in each otocyst two miniscule
otoliths, each composed of a number of minute
granules. During Stages 32 and 33 the opercu-
lum and gill arches differentiate, but circulation
through the gill arches does not appear until
late Stage 35 when it is established through the
first arch.

At Stage 35 the head has extended forward,
carrying with it the operculum, resulting in a
gap between the operculum and the first gill
arch. During Stages 35 and 36 there is an open
communication through the pharynx from side
to side. This closes in Stage 37 by the backward
growth of the operculum.

It is in Stage 37 that the first melanin pigment
is seen in the eye, sparsely scattered in the upper
posterior quadrant. Blood vessels form in the
eye in Stage 38 with an active circulation. Also
at this time a circulation is established through
the liver which is located along the posterior
margin of the left anterior cardinal vein lateral
to the body axis.

In Stage 40, circulation is established in the
maxillary and lateral mandibular barbels, in the
pectoral fin and in the caudal fin, the latter by a
single vessel extending posteroventrally from
the aorta. Also in Stage 40, bile is present in the
gut at its anterior end and movements of the
lower jaw first appear.

During the period of organodifferentiation
there is a progressive extension caudally of con-
tractility of the myotomes. At Stage 29 the tail
flexes laterally to the side of the head, at Stage 37
it sweeps over the dorsum of the head. Also in
Stage 37, the muscles of the anterior somites are
cross striated. In late Stage 41, progressive un-

dulatory swimming occurs, the embryo com-
monly swimming on its side. Photokinetic re-
sponse to the "off" of light occurs first in Stage
42, with a well-defined negative phototactic
response demonstrable in Stage 44.

Spontaneous hatching occurs in Stage 43, with
most of the embryos of an egg mass emerging in
three to four hours but with delay of some until
Stage 44. By the time Stage 44 is reached, there
is yellow bile in all but the terminal end of the
gut.

The subsequent stages are marked by an in-
crease in size of the embryos and their ap-
pendages, development of the circulation into
the fins, and an extension of the pigment to
finally produce what is essentially the adult
pattern. The embryos are positively geotropic
until Stage 52, when some negative geotropism
appears. This becomes more marked as the em-
bryos progress into Stage 53. This negative geo-
tropism is probably correlated with the develop-
ment of the bilobed swim bladder which can be
demonstrated readily on dissection of fixed
embryos. Also in these late stages the embryos
apparently adapt to intensities of light which
earlier evoked a negative phototactic response.

At 20 to 21° C. the development described
above through Stage 53 is completed in seven-
teen days. The table below gives the time
sequence of this development.

Time Sequences,

Development of Ictalurus nebulosus

Stage Time

Stage Time

Stage

Time

1 1.2 hrs.

25 2 days

43

8 days

2 1.6 hrs.

29 3 days

46

9 days

4 3 hrs.

34 4 days

48

10 days

8 6 hrs.

37 5 days

50

12 days

14 9 hrs.

39 6 days

52

15 days

18 1 day

40 7 days

53

17 days

BIBLIOGRAPHY

Bachmann, Freda M. 1914. The migration of germ
cells in Amiurus nebulosus. Biol. Bull. 26: 351-
366.

Breder, C. M., Jr. 1933. On the genesis of oral incuba-
tion in fishes. Anat. Rec. 57: Sup. 62-63.

. 1935. The reproductive habits of the common

catfish, Ameiurus nebulosus (Le Sueur) , with a
discussion of their significance in ontogeny and
phylogeny. Zoologica N.Y. 19: 143-185.

1939. Variations in the nesting habits of

Ameiurus nebulosus (Le Sueur) . Zoologica N.Y.

24: 367-378.
Cable, Louella E. 1927. The food of bullheads. Anat.

Rec. 37: 170.
Eycleshymer, Albert C. 1901. Observations on the

breeding habits of Ameiurus nebulosus. Amer.

Nat. 35:911-918.
Gudger, E. W. 1916. The gaff-topsail (Felichthys felis)

a sea catfish that carries its eggs in its mouth.

Zoologica N.Y. 2: 125-158.
Hasler, Arthur D., Roland K. Meyer and Howard M.

Field. 1939. Spawning induced prematurely in

trout with the aid of pituitary glands of the carp.

Endocrinology. 25: 978-983.
Kendall, William Converse. 1902. Habits of some of

the commercial catfishes. Bull. U.S. Fish Comm.

22: 401-409.

Lee, Genevieve. 1937. Oral gestation in the marine
catfish, Galeichthys felis. Copeia 1937. No. 1. 49-56.

Oppenheimer, Jane M. 1937. The normal stages of
Fundulus heteroclitus. Anat. Rec. 68: 1-15.

Ramaswami, L. S. and B. I. Sundararaj. 1957. Inducing
spawning in the Indian catfish Heteropneustes
with pituitary injections. Acta anat. 31: 551-562.

Ryder, John A. 1883. Preliminary notice of the de-
velopment and breeding habits of the Potomac
catfish, Amiurus albidus (Le Sueur) Gill. Bull.
U.S. Fish Comm. 3: 225-230.

Schiche, Otto E. 1921. Reflexbiologische Studien an
Bodenfischen. I Beobachtungen an Amiurus nebu-
losus Les. Zool. Jb., Abt. Allg. Zool. u. Physiol.
38: 49-112.

Smith, Hugh M. and L. G. Harron. 1902. Breeding
habits of the yellow catfish. Bull. U.S. Fish Comm.
22: 151-154.

Solberg, Archie Norman. 1938. The development of a
bony fish. Progr. Fish Cult. No. 40, 1-19.

Swarup, H. 1958. Stages in the development of the
stickleback, Gasterosteus aculeatus(L). J.Embryol.
exp. Morph. 6: 373-383.

Wilson, H. V. 1891. Embryology of the sea bass (Ser-
ranus atrarius). Bull. U.S. Fish Comm. 6: 209-277.

Witschi, Emil. 1956. Development of Vertebrates.
W. B. Saunders Co., Philadelphia. 588 pp.

i^^CS^

Br

; Bail • ■

Hi

n

;•- SIS

Hi I

STAGES IN THE
DEVELOPMENT OF

Ictalurus nebulosus

By PHILIP B. ARMSTRONG, M.D.

College of Medicine at Syracuse
State University of New York
and Marine Biological Laboratory
Woods Hole, Massachusetts

Illustrated by

JULIA SWOPE CHILD

Marine Biological Laboratory
Woods Hole, Massachusetts

SYRACUSE UNIVERSITY PRESS

SIU 1962

PRESS

Library of Congress Catalog Card 63-1 1037

Copyright © 1962 by Syracuse University Press

Syracuse, New York

ALL RIGHTS RESERVED

The development and publication of this normal series was sup-
ported by U.S. Public Health Service grants B-64 3 and GM-10758.

/I75"

MANUFACTURED IN THE UNITED STATES OF AMERICA

)

STAGES IN THE DEVELOPMENT OF
ICTALURUS NEBULOSUS

Ictalurus nebulosus, the brown bullhead or
horned pout, is generally distributed through-
out the Eastern and Central United States from
Maine and the Dakotas on the north into the
Gulf States on the south. It has been introduced
successfully into streams and ponds on the West
Coast and into the waters of several European
and Far Eastern countries.

Under favorable conditions adult bullheads
attain a length of a foot or more with occasional
specimens 18 inches in length weighing 3 to 4
pounds. When crowded they are much smaller.
Ictalurus nebulosus is very tenacious of life,
surviving under conditions which will not sup-
port other native fishes. It is commonly found
in small ponds, lakes, and sluggish streams and
does well even in a muddy habitat. The adults

| are frequently caught on a drop-line during the
daytime but are more active at night. The eyes
are small but the olfactory and cutaneous senses
are highly developed. They are omnivorous,
the diet depending on what is available. The
adults are readily collected in a baited minnow
trap with an adequate entrance and with reason-
ble care will survive well in aquaria.

The reproductive habits of Ictalurus nebu-
losus have been observed in both natural sur-
roundings and in aquaria. The eggs are usually
deposited in a prepared nest, an open shallow
excavation among weeds, or a burrow excavated
by the adults under the roots of a water plant
which will support the roof of the nest. A com-
mon site for such a burrowed nest is horizontally
for as much as three feet into the bank of the
stream or pond. The bullhead resembles the
wren in selecting a nesting site, taking advantage
of protected locations in pails, stovepipes, rub-
ber tires, cinder blocks— or under boards, stones,

| or other articles thrown by chance into the

water. Rarely are the eggs laid in an open site
without preparation. Depending on the nature
of the nest, the floor is invariably some firm
material— coarse sand, gravel, or stone. When
the nest is found in the muddy bottom of a
pond, it is located where the mud is a shallow
layer on a sandy or gravel bottom to which the
nest is excavated. Drainage tiles provide attrac-
tive nesting sites for bullheads. Tiles are par-
ticularly useful in ponds supporting a heavy
growth of weeds in which it is difficult to locate
the natural nesting sites. The tiles should be
distributed separately around the margin of the
pond in two or three feet of water, their loca-
tions indicated by a suitable marker.

The eggs are adhesive when laid and form a
spongy pale yellow mass which adheres to the
floor of the nest. Each egg through its chorionic
membrane has adhesion discs with several adja-
cent membranes which sometimes show up on
the surface of the chorionic membranes when
they are torn from each other (see figure of
Stage 20) . The micropyle is located as the center
of a stellate structure seen in some membranes
(Stage 1 et al.). When the eggs are laid in indi-
vidual drainage tiles it is frequently necessary
to gently scrape the egg mass free of the tile to
which it adheres. If the floor of the nest is sand
or gravel, grains of sand and small pebbles are
found adhering to the egg mass. There are from
several hundred up to a thousand and more eggs
in a single egg mass. The adults, particularly
the male, guard the eggs. Frequently an ob-
server can flush the adult off a nest by wading
in a pond a few yards off shore, locating as close-
ly as possible the point at the bank from which
the adult emerges. Close inspection of the bank
at this point will usually reveal the opening of
the nest. The collector can reach into the bur-

STAGES IN THE DEVELOPMENT

row, and by careful exploration the egg mass,
which is soft and yielding, can be obtained. It
must be handled gently. (Sometimes a second
fish is flushed from the nest and rushes past the
collector's arm as he attempts to find the egg
mass.)

The adults work over the eggs very actively
(Breder, 1935), fanning them with their fins,
taking them in their mouths and ejecting them,
and in general handling them rather roughly.
Apparently almost constant agitation is re-
quired to assure normal development of the
eggs, since eggs removed to still water usually
will die within a day or so. Two simple methods
were devised for handling the eggs; either will
usually give normal development of the major
share of the eggs of a cluster. The cluster may be
placed in a seven-inch finger bowl in tap water
flowing from a rubber tube of which the lower
three inches rests on the bottom of the finger
bowl directed to give a swirling flow of the
water in the bowl. There should be a flow of
water adequate to agitate the eggs slightly but
not to carry them with the current. A constant
flow of water can best be obtained through a
petcock without a washer. In a washered outlet,
the flow reduces as the washer swells. The sec-
ond method is illustrated in Figure 1, which
shows a strainer to which a metal tube is
soldered in such a position that the water flows
from below through the egg mass resting in the
strainer. The egg masses should be examined
at least twice a day and any dead eggs removed,
since these quickly develop mold which extends
to the adjacent eggs and kills them.

The laying season starts about the same time
in central New York State and on Cape Cod,
Massachusetts— i.e., about the middle of May.
The latest that newly laid eggs were found on
Cape Cod was July 28. Adults in aquaria will
lay spontaneously, with the opening and closing
dates of the spawning somewhat later than with
fish in the wild. Females early in the winter
carry eggs which in appearance and size are close

to maturity. We induced spawning and obtained
fertilized eggs out of season by injecting into a
pair of bullheads crushed pituitaries from ripe
female alewives, but additional work must be
done to standardize the procedures if uniform
success is to be achieved.

The eggs measure about 3 mm. in diameter.
Those from the same egg mass are remarkably
uniform in size but there are differences in size
from one egg mass to another. The subchorionic
space is wide and accounts for about one-third
of the diameter of the egg. The chorionic mem-
brane is transparent, naked, and relatively thin
for the size of the egg. The membrane can be
removed readily without injury to the develop-
ing embryo merely by tearing the membrane
with fine forceps. After the blastopore is closed,
the embryos survive in running tap water with-
out any special treatment; in fact, they are less
susceptible to death than if left in the mem-
branes where the mechanics of aeration are
more difficult. The embryos hatch spontaneous-
ly in Stage 43 and will swim actively on the floor
of their container. In the earlier swimming
stages, the embryos can be kept in an open
dish with a gentle stream of water circulating
through the dish. In the more advanced stages,
they become negatively geotropic and are lost
from an open dish, but they can be kept in a
battery jar closed with a strainer fitted with a
tube through which water can be introduced
from above (Figure 2) . The strainer must be

FIGURE 1

FIGURE 2

OF ICTALURUS NEBULOSUS

of such a size as to fit snugly in the top of the
battery jar.

The embryos leave the nests and seek food
when the yolk has been absorbed. For a few
days they swim as a compact school in shallow
water guarded by the adults which swim about
below the school. After a few days the school
breaks up and the larvae disappear into deeper
water.

To identify stages of the embryos, it is ad-
vantageous to examine them with transmitted
and with reflected light. Light transmitted
through the embryo from a white substage mir-
ror is especially effective in bringing out the
details of pigmentation. The developing blood
vessels and circulation are revealed best by the
use of a regular substage mirror. In lighting the
embryo from above, strong illumination, such
as that from a zirconium lamp, is required to
bring out the finer details. Also all available
features of the developing embryos, both mor-
phological and physiological, should be taken
into consideration for accurate identification of
stages.

The embryos in the same egg mass, if properly
handled, show remarkable synchrony in their
development. However, minor but perceptible
variations do occur from mass to mass, possibly
because the development of the various features
of the embryo do not depend on a single com-
mon factor. Because of this, embryos will be
included in the same stage in spite of some such
minor variation from the pertinent illustration.
In the illustrations the embryos are magnified
17 times.

Some of the special features of development
are described below.

CLEAVAGE

Stages 1 through 7

Cleavage in the bullhead egg is meroblastic

and meridional as is characteristic of the teleosts,

the cleavages being limited to the blastodisc, not

involving the yolk. The diameter of the early

blastodisc is only about one-eighth the circum-
ference of the egg. The successive cleavages fol-
low each other at 35 to 40 minute intervals and
extend well through the disc. The blastomeres
formed at each cleavage are of equal size and
the cell divisions at any one cleavage are closely
synchronous. From the margins of the disc the
protoplasm thins out abruptly and extends
around the yolk as a thin continuous layer, as
in Fundulus. There is no defined periblastic
ridge such as occurs in Serranns (Wilson 1891).
Cleavage continues to be meridional in the bull-
head at least through the 32-cell stage and pos-
sibly through the 64-cell stage, but it is quite
probable that in this latter cleavage some of the
central cells divide horizontally.

BLASTULA

Stages 8 through 15
The subchorionic (perivitelline) space in the
bullhead egg is wide and the chorionic mem-
brane thin and yielding. This permits the for-
mation of a high blastula which is not seen in
those teleosts with narrow subchorionic spaces
such as Trutta, Fundulus, and Opsanus. The
early blastulae (Stages 8 and 9) very much re-
semble the morula stage of some of the inverte-
brates, the individual cells being large enough
to give the morula effect. The blastula remains
high (Stages 10 and 11) through additional cell
divisions and then gradually flattens out (Stages
12 to 15) with the formation of the blastocoele
which is apparent in the living egg (Stage 15).
The high blastula (Stage 1 1) has a diameter even
less than that of the one-cell stage, whereas the
flattened blastula (Stage 15) has a diameter more
than half again that of the one-cell stage. As the
blastula begins to flatten out in Stage 12, there
is the beginning formation of marginal periblast
cells, the process continuing through Stage 13,
resulting in the formation of roughly two rows
of such cells placed at a distance from the blasto-
derm. These periblast cells disappear later as
the blastoderm extends over the yolk.

STAGES IN THE DEVELOPMENT

GASTRULA
Stages 1 6 through 1 9
When the blastoderm has expanded to cover
about one-fourth of the yolk, there develops,
through mitotic activity, an accumulation of
cells at its margin (Stage 16) resulting in the
formation of the germ ring. In Stage 17 the
anterior lip of the blastopore is seen as an extra
accumulation of cells forming the embryonic
shield which becomes more prominent in Stages
18 and 19, seen best as viewed on the horizon
of the egg.

NEURULA

Stages 20 through 24
During Stages 20 to 24 the blastoderm con-
tinues to extend over the yolk. This culminates
in the closure of the blastopore late in Stage 24.
The blastoderm covers about half the yolk in
Stage 20. The embryonic shield in this stage
bears considerable resemblance to the open
neural plate of the amphibia. However, true
concrescence as in the amphibia does not occur.
There is a median longitudinal cellular pro-
liferation with a narrowing of the neural plate
resulting in the formation of a definitive pre-
somitic embryonic axis (Stage 21). After the
formation of the embryonic axis neurulation
proceeds rapidly. In Stage 22 the main divisions
of the brain can be discerned, the optic vesicle
is rudimentary, and the first somites appear.
There follows continued growth and further
differentiation of the divisions of the brain and
of the optic vesicle and a continuing addition of
somites in a caudal direction. In Stage 24 there
is a bilateral accumulation of cells, the rudiment
of the otic vesicle, at the level of the anterior
hind brain.

TAIL BUD EMBRYO

Stages 25 and 26
In Stage 25 (ca. 14 somites) the tail bud is well
denned. Around its periphery, the ectoderm is
directly continuous with that of the yolk sac.

The blastopore is closed, but its location is ^^
marked by a slight depression in the yolk. Al- ^F
though the main divisions of the brain can be
discerned, the brain ventricles are not yet
apparent.

With continued growth, the tip of the tail
bud develops as a projection free of ectodermal
continuity with the yolk sac as in Stage 26 (ca. 17
somites). In this stage the pericardial sac is
developing and the heart rudiment forms as a
small bit of tissue in the pericardial sac under
the head. However, the heart rudiment does
not beat when it first forms.

ORGANODIFFERENTIATION
Stages 27 through 53

This period includes development extending
through to the complete absorption of the yolk.
The embryos prior to Stage 43, when hatching
normally occurs, were removed from the chori-
onic membranes for clearer observation and
illustration.

In Stage 27, the heartbeat is just discernible ^&
in transmitted light. It is faint but regular.
Also, slow weak contractions of the anterior
somites result on gross mechanical stimulation
of the embryos. No cross striations are seen in
the developing muscle.

In Stage 28, the heartbeat is still faint but
readily visible. Also there are spontaneous con-
tractions of the anterior somitic muscle. The
first formation of the brain ventricles is apparent
in the region of the hind brain indicated by the
formation of the roof of the fourth ventricle,
seen in the lateral view of the embryo.

In Stage 29, muscle contractility is still limited
to the anterior somites. There has been an in-
crease in the size of the heart, but the circulation
of the blood is not established until Stage 30
and at first is sluggish and principally intraem-
bryonic. Sometimes at this stage there is seen
in the lower part of the otocyst an aggregation
of minute granules, the earliest indication of
the otoliths.

OF ICTALURUS NEBULOSUS

In Stage 31a sluggish circulation is present
over the surface of the yolk sac, with the cells
of the communicating blood islands moving
slowly toward the venous end of the heart.
There are now in each otocyst two miniscule
otoliths, each composed of a number of minute
granules. During Stages 32 and 33 the opercu-
lum and gill arches differentiate, but circulation
through the gill arches does not appear until
late Stage 35 when it is established through the
first arch.

At Stage 35 the head has extended forward,
carrying with it the operculum, resulting in a
gap between the operculum and the first gill
arch. During Stages 35 and 36 there is an open
communication through the pharynx from side
to side. This closes in Stage 37 by the backward
growth of the operculum.

It is in Stage 37 that the first melanin pigment
is seen in the eye, sparsely scattered in the upper
posterior quadrant. Blood vessels form in the
eye in Stage 38 with an active circulation. Also
at this time a circulation is established through
the liver which is located along the posterior
margin of the left anterior cardinal vein lateral
to the body axis.

In Stage 40, circulation is established in the
maxillary and lateral mandibular barbels, in the
pectoral fin and in the caudal fin, the latter by a
single vessel extending posteroventrally from
the aorta. Also in Stage 40, bile is present in the
gut at its anterior end and movements of the
lower jaw first appear.

During the period of organodifferentiation
there is a progressive extension caudally of con-
tractility of the myotomes. At Stage 29 the tail
flexes laterally to the side of the head, at Stage 37
it sweeps over the dorsum of the head. Also in
Stage 37, the muscles of the anterior somites are
cross striated. In late Stage 41, progressive un-

dulatory swimming occurs, the embryo com-
monly swimming on its side. Photokinetic re-
sponse to the "off" of light occurs first in Stage
42, with a well-defined negative phototactic
response demonstrable in Stage 44.

Spontaneous hatching occurs in Stage 43, with
most of the embryos of an egg mass emerging in
three to four hours but with delay of some until
Stage 44. By the time Stage 44 is reached, there
is yellow bile in all but the terminal end of the
gut.

The subsequent stages are marked by an in-
crease in size of the embryos and their ap-
pendages, development of the circulation into
the fins, and an extension of the pigment to
finally produce what is essentially the adult
pattern. The embryos are positively geotropic
until Stage 52, when some negative geotropism
appears. This becomes more marked as the em-
bryos progress into Stage 53. This negative geo-
tropism is probably correlated with the develop-
ment of the bilobed swim bladder which can be
demonstrated readily on dissection of fixed
embryos. Also in these late stages the embryos
apparently adapt to intensities of light which
earlier evoked a negative phototactic response.

At 20 to 21° C. the development described
above through Stage 53 is completed in seven-
teen days. The table below gives the time
sequence of this development.

Time Sequences,

Development of Ictalurus

NEBULOSUS

Stage Time

Stage Time

Stage

Time

1 1.2 hrs.

25 2 days

43

8 days

2 1.6 hrs.

29 3 days

46

9 days

4 3 hrs.

34 4 days

48

10 days

8 6 hrs.

37 5 days

50

12 days

14 9 hrs.

39 6 days

52

15 days

18 1 day

40 7 days

53

17 days

8

BIBLIOGRAPHY

Bachmann, Freda M. 1914. The migration o£ germ
cells in Amiurus nebulosus. Biol. Bull. 26: 351-
366.

Breder, C. M., Jr. 1933. On the genesis of oral incuba-
tion in fishes. Anat. Rec. 57: Sup. 62-63.

. 1935. The reproductive habits of the common

catfish, Ameiurus nebulosus (Le Sueur) , with a
discussion of their significance in ontogeny and
phylogeny. Zoologica N.Y. 19: 143-185.
-. 1939. Variations in the nesting habits of

Ameiurus nebulosus (Le Sueur) . Zoologica N.Y.

24: 367-378.
Cable, Louella E. 1927. The food of bullheads. Anat.

Rec. 37: 170.
Eycleshymer, Albert C. 1901. Observations on the

breeding habits of Ameiurus nebulosus. Amer.

Nat. 35:911-918.
Gudger, E. W. 1916. The gaff-topsail (Felichthys felis)

a sea catfish that carries its eggs in its mouth.

Zoologica N.Y. 2: 125-158.
Hasler, Arthur D., Roland K. Meyer and Howard M.

Field. 1939. Spawning induced prematurely in

trout with the aid of pituitary glands of the carp.

Endocrinology. 25: 978-983.
Kendall, William Converse. 1902. Habits of some of

the commercial catfishes. Bull. U.S. Fish Comm.

22: 401-409.

Lee, Genevieve. 1937. Oral gestation in the marine
catfish, Galeichthys felis. Copeia 1937. No. 1. 49-56.

Oppenheimer, Jane M. 1937. The normal stages of
Fundulus heteroclitus. Anat. Rec. 68: 1-15.

Ramaswami, L. S. and B. I. Sundararaj. 1957. Inducing
spawning in the Indian catfish Heteropneustes
with pituitary injections. Acta anat. 31: 551-562.

Ryder, John A. 1883. Preliminary notice of the de-
velopment and breeding habits of the Potomac
catfish, Amiurus albidus (Le Sueur) Gill. Bull.
U.S. Fish Comm. 3: 225-230.

Schiche, Otto E. 1921. Reflexbiologische Studien an
Bodenfischen. I Beobachtungen an Amiurus nebu-
losus Les. Zool. Jb., Abt. Allg. Zool. u. Physiol.
38:49-112.

Smith, Hugh M. and L. G. Harron. 1902. Breeding
habits of the yellow catfish. Bull. U.S. Fish Comm.
22: 151-154.

Solberg, Archie Norman. 1938. The development of a

bony fish. Progr. Fish Cult. No. 40, 1-19. ^

Swarup, H. 1958. Stages in the development of the ^P -
stickleback, Gasterosteus aculeatus(L). J.Embryol.
exp. Morph. 6: 373-383.

Wilson, H. V. 1891. Embryology of the sea bass (Ser-
ranus atrarius). Bull. U.S. Fish Comm. 6: 209-277.

Witschi, Emil. 1956. Development of Vertebrates.
W. B. Saunders Co., Philadelphia. 588 pp.

PLATE I - STAGES IN THE DEVELOPMENT OF ICTALURUS NEBULOSUS

S

Copyright © 1962 by Syracuse University Press, Syracuse, New York

V

PLATE II - STAGES IN THE DEVELOPMENT OF ICTALURUS NEBULOSUS

10

11

12

13

14

\

•

/

v

15

16

\

17

X

Copyright © 1962 by Syracuse University Press, Syracuse, New York

V

\_

*

PLATE III - STAGES IN THE DEVELOPMENT OF ICTALURUS NEBULOSUS

18

/

'

/

19

20

20

21

21

I

22

22

Copyright © 1962 by Syracuse University Press, Syracuse, New York

ۥ

0 -

PLATE IV - STAGES IN THE DEVELOPMENT OF ICTALURUS NEBULOSUS

23

23

24

24

25

i

26

26

27

..>

/

Copyright © 1962 by Syracuse University Press, Syracuse, New York

e

PLATE V - STAGES IN THE DEVELOPMENT OF ICTALURUS NEBULOSUS

28

28

29

1

30

Nx

32

31

Copyright © 1962 by Syracuse University Press, Syracuse, New York

PLATE VI - STAGES IN THE DEVELOPMENT OF ICTALURUS NEBULOSUS

35

Copyright © 1962 by Syracuse University Press, Syracuse, New York

C"

%$»

*

PLATE VII - STAGES IN THE DEVELOPMENT OF ICTALURUS NEBULOSUS

37

.""

37

f

'

38

38

39

39

Copyright © 1962 by Syracuse University Press, Syracuse, New York

% '

PLATE VIII - STAGES IN THE DEVELOPMENT OF ICTALURUS NEBULOSUS

40

0
451

40

7'

f% .." #

41

41

\

Copyright © 1962 by Syracuse University Press, Syracuse, New York

PLATE IX - STAGES IN THE DEVELOPMENT OF ICTALURUS NEBULOSUS

43

'

w* *'■' \

Copyright © 1962 by Syracuse University Press, Syracuse, New York

*

c

PLATE X - STAGES IN THE DEVELOPMENT OF ICTALURUS NEBULOSUS

45

m

.. -- -,. ?... -. -

it

Copyright © 1962 by Syracuse University Press, Syracuse, New York

^a

<N

PLATE XI - STAGES IN THE DEVELOPMENT OF ICTALURUS NEBULOSUS

Copyright © 1962 by Syracuse University Press, Syracuse, New York

*.

*

m

PLATE XII - STAGES IN THE DEVELOPMENT OF ICTALURUS NEBULOSUS

■ Y

ft

1

W

48

48

Copyright © 1962 by Syracuse University Press, Syracuse, New York

(^

PLATE XIII - STAGES IN THE DEVELOPMENT OF ICTALURUS NEBULOSUS

50

50

Copyright © 1962 by Syracuse University Press, Syracuse, New York

9j

PLATE XIV - STAGES IN THE DEVELOPMENT OF ICTALURUS NEBULOSUS

51

f

}

Copyright © 1962 by Syracuse University Press, Syracuse, New York

($$&*

4jM**

PLATE XV - STAGES IN THE DEVELOPMENT OF ICTALURUS NEBULOSUS

52

Copyright © 1962 by Syracuse University Press, Syracuse, New York

PLATE XVI - STAGES IN THE DEVELOPMENT OF ICTALURUS NEBULOSUS

HXi>

53

■ \ s-.

j

'' " ■■ » « ' '1 S ,

'

Copyright © 1962 by Syracuse University Press, Syracuse, New York