Exercise 7 Studies on Chick Embryo Using Prepared Slides

EXERCISE 7
STUDIES ON CHICK EMBRYO
USING PREPARED SLIDES

Structure
7.1 Introduction Study of Chick Embryo at
18 Hours
Objectives
Study of Chick Embryo at
7.2 Material Required
24 Hours
7.3 Study Guide
Study of Chick Embryo at
7.4 Study of Developmental 33 Hours
Stages of Chick
Study of Chick Embryo at 72
Study of Chick Embryo at Hours of Incubation
4 Hours
Study of Chick Embryo at 96
Study of Chick Embryo at Hours of Incubation
16 Hours
7.5 Terminal Questions

7.1 INTRODUCTION
In exercise 6 you have already studied the developmental stages of frog with
the help of prepared permanent slides under the compound microscope. The
prepared permanent slides of the whole mounts and sections of embryos of
chicks can be used for the study of chick embryology and to follow the course
of development of chicks. In this exercise you will observe under the
microscope the developmental stages of chick embryo, starting from 4 hours
after incubation and following its development while the organs and organ
systems are being formed upto 96 hours of incubation.

Objectives
After completing this exercise you will be able to:

identify the various stages of development in chick embryos from

prepared permanent slides,

75
BZYCL-134 Laboratory of Comparative Anatomy and Development Biology of Vertebrates

draw well labeled diagrams of the various stages of the developing
embryos that have been studied in this lab exercise, and

trace the progressive development of organ and organ system of the

chick embryo during its development.

7.2 MATERIAL REQUIRED

1. Permanent, prepared slides of chick embryo at the following stages of
incubation:

• 4 hours after incubation (whole mount),

• 16 hours after incubation (whole mount and longitudinal section (L.S.)),
• 18 hours after incubation (whole mount and longitudinal sections
through head and mid body regions),
• 24 hours after incubation (whole mount and transverse section (T.S.)),
• 33 hours after incubation (whole mount and transverse section),
• 72 hours after incubation (whole mount), and
• 96 hours after incubation (whole mount).

2. Laboratory manual of BZYCL-134.

3. Practical note book.

4. Pen, Pencil, Eraser and Ruler.

5. Dissecting microscope.

6. Compound microscope.

7.3 STUDY GUIDE

In order to complete your laboratory exercise satisfactorily, you must read in
advance, Unit 15, entitled, “Development of Chick” given in Volume 2 of the
theory Core Zoology Course-BZYCT-133 (Comparative Anatomy and
Developmental Biology of Vertebrates) which is part of the B.Sc. General
Course (CBCS).

In the present exercise, you will study with the help of prepared slides, the
salient developmental stages of chick observed by you under the compound
microscope. You are expected to consult the present laboratory exercise since
the descriptions and labeled figures given in it will help you to study the
developmental stages of chick properly.

Make careful observations of the slides showing different stages of

development of chick embryos. Match your observations with the descriptions
we have provided here for each of the stages and draw neat and labelled
diagrams in your record note book. The study of embryology of chick would
enable you to understand the differentiation of tissues, organs and organ
systems from early embryonic cells. It is worth to note that such a study with
help of permanent slides is possible only upto a certain stage of development.
76
Exercise 7 Studies on Chick Embryo Using Prepared Slides

7.4 STUDY OF DEVELOPMENTAL STAGES OF

CHICK
We will begin the study of chick embryo from 4 hours of incubation at which
stage the differentiation of the blastodisc occurs. After this we will follow the
development of different stages of development till the chick embryo is at 96
hours of incubation.

7.4 .1 Study of Chick Embryo at 4 Hours

Features of whole mount chick embryo at 4 hours.

i. At 4 hours of incubation the differentiation of the blastodisc into area

pellucida and area opaca can be observed in the chick embryo.

ii. One quadrant of the area pellucida appears thickened which is the
region of the future caudal end of embryo (Fig. 7.1). After 7 to 8 hours,
the thickening will become more elongated and then represents the start
of primitive streak.

Fig. 7.1: Chick embryo 4 hours after incubation.

7.4 .2 Study of Chick Embryo at 16 Hours

I. Features of whole mount of chick embryo at 16 hours of incubation.

i) In the whole mount of a 16 hour chick embryo (Fig. 7.2) you will
observe the distinct primitive streak. The embryo at this stage is
characterized as being in the primitive streak stage.

ii) In the whole mount, the embryo is seen to have a central furrow,
called the primitive groove which appears lined by thickened
primitive ridges.

iii) At the cephalic end (head end) of the embryo, closely packed cells
form a thickened area, called as Hensen's node.

iv) Part of area pellucida adjacent to the primitive streak shows

increased thickness and forms the embryonic area or embryonic
shield. 77
BZYCL-134 Laboratory of Comparative Anatomy and Development Biology of Vertebrates

v) The area of pellucida at this stage assumes an elliptical shape.

vi) An elongated primitive streak is visible which represents the long

axis of the future embryonic body.

vii) The end diametrically opposite to the Hensen's node is the caudal
end of the embryo.

Fig. 7.2: Whole mount of chick embryo, 16 hours after incubation.

II. Features of chick embryo in longitudinal section (LS) at 16 hours

after incubation

i) The LS through the 16 hour chick embryo (Fig. 7.3) represents the
embryonic chick stage shortly after primitive streak formation. It
also marks the beginning of morphogenetic movement of cells to
form the notochord.

ii) The LS shows the ectoderm, Hensen's node, primitive pit, primitive
groove, notochord and primitive gut. The mesoderm extends on
either side between ectoderm and endoderm.

78 Fig. 7.3: LS. of 16 hour old Chick embryo after incubation.

Exercise 7 Studies on Chick Embryo Using Prepared Slides

7.4 .3 Study of Chick Embryo at 18 Hours

I. Features of whole mount of chick embryo at 18 hours of incubation.

i) In the 18 hour whole embryo you will observe that the notochord has
become markedly elongated to form a conspicuous structure (Fig
7.4).

ii) Notochord is seen to extend from the Hensen's node towards the
cephalic region present in the middle.

iii) Embryo at this stage of development is said to be in the head

process stage.

iv) Neural plate develops around the notochord.

v) The dark peripheral area opaca, the inner translucent area

pellucida and the central embryonal area are clearly visible.

vi) In the anterior region a small and more translucent portion of area
pellucida, known as proamnion can be observed.

vii) The primitive streak lies in the middle of the area pellucida in the
posterior half.

viii) The neural plate and primitive streak can be seen to be separated
by Hensen's node.

Fig. 7.4: Whole mount of chick embryo, 18 hours after incuhation.

II. Features of chick embryo in longitudinal section (LS) at 18 hours

after incubation

i) The LS of the 18 hours incubated chick embryo (Fig. 7.5) shows

advanced inner structures of the germ layers. 79
BZYCL-134 Laboratory of Comparative Anatomy and Development Biology of Vertebrates

ii) Ectoderm is seen to have vertical cells while the cells of the
mesoderm are represented by heavy angular dots. Endoderm is
represented by stippling backed by a single line.

iii) Yolk, ectoderm of neural plate, notochord, mesodem, ectoderm

and endoderm of blastoderm can be observed in the section of the
slide (Fig. 7.5).

iv) You will also observe the primitive pit, primitive ridge, and primitive
gut.

Fig. 7.5: Two longitudinal sections of chick embryo at 18 hours after incubation
in which section is from two regions (A). The two LS. of chick embryo
at 18 hrs (B1 and B2).

7.4 .4 Study of Chick Embryo at 24 Hours

I. Features of whole mount of chick embryo at 24 hours of incubation.

i) In the whole mount of a 24 hour old embryo (Fig 7.6) the cephalic
region is prominently visible because of rapid growth in this region.

ii) The cephalic region extends anteriorly and overhangs the

proamnion region. The cephalic region which projects free from the
blastoderm can now be properly termed as the head of embryo.

iii) A space between the head and the blastoderm is formed and is
visible and is called the sub- cephalic pocket.

iv) In the mid-line of the embryo the notochord is visible. It is larger at

the caudal area near its point of origin than at the cephalic area.

v) The neural plate is much more clearly visible. The neural folds
appear as a pair of dark bands.

vi) At its cephalic end, the neural groove is deeper. The neural folds
are correspondingly more prominent at the cephalic end than they
are caudally.

vii) Four pairs of somites are seen in the mid-line.

viii) Primitive streak is seen to gradually decrease in size.

80 ix) Foregut is also formed and is visible.

Exercise 7 Studies on Chick Embryo Using Prepared Slides

x) The part of the gut, caudal to the foregut is termed the midgut and
the opening of the midgut into the foregut is called the anterior
intestinal portal.

xi) In addition to the structures, mentioned above the area opaca

vitellina, area pellucida, proamnion, Hensen's node, area
vasculosa, blood islands and unsegmented mesoderm are also
seen (Fig. 7.6).

Fig. 7.6: Whole mount of Chick embryo at 24 hrs of incubation.

II. Features of chick embryo in transverse section (T.S) at 24 hours

after incubation

i) The transverse section passing through head region (Fig. 7.7a) shows
the folded neural plate forming a complete tube.

ii) The notochord can be seen beneath the neural fold.

iii) Mesenchyme, foregut, ectoderm of head, mesoderm and

endoderm can be seen in this section.

iv) The transverse section passing through mid-body of the chick

embryo (Fig. 7.7b) shows formation of somites and changes in the
mesoderm.

v) Mesoderm is seen to be differentiated into: i) dorsal mesoderm, ii)

intermediate mesoderm and iii) lateral mesoderm.

vi) Other structures seen in the slide are ectododerm, endoderm,

lateral margin of anterior intestinal portal, midgut and pericardial
coelom. 81
BZYCL-134 Laboratory of Comparative Anatomy and Development Biology of Vertebrates

Fig. 7.7: T.S. passing through a) head region; and b) mid body in chick of
embryo 24 hours after incubation.

7.4.5 Study of Chick Embryo at 33 Hours

I. Features of whole mount of chick embryo at 33 hours of incubation

i) The 33 hour old embryo of chick (Fig. 7.8) shows some of the
fundamental structures which will be involved in the formation of
central nervous system and circulatory system.

ii) You will notice remarkable changes in the development of the

brain. Observe that the brain is differentiated into prosencephalon
(fore-brain), mesencephalon (mid-brain), and rhombencephalon
(hind- brain).

iii) The optic vesicles have been established and can be seen in the
whole mount as paired lateral outgrowths of the prosencephalon.
The optic vesicles will later extend to occupy the full width of the
head. Infundibulum is seen to have formed in the floor of the
prosencephalon.

iv) Mid-region of the heart is considerably dilated and is bent to the

right.

v) Twelve pairs of somites can be observed.

vi) Anterior omphalomesentric veins have developed and can be

seen.

vii) The primitive streak becomes shorter because of the lengthening

of the neural tube.

viii) Proamnion, neural tube, notochord, sinus rhomboidalis and sinus

82 terminalis are also present and can be seen.
Exercise 7 Studies on Chick Embryo Using Prepared Slides

Fig. 7.8: Whole Mount of Chick embryo at 33 hours of incubation.

II. Features of chick embryo in transverse section (T.S) at 33 hours

after incubation

i) The transverse section of the 33 hour old embryo chick (Fig. 7.9)
shows the following structures ectoderm, prosocoel, opticoel,
mesenchyme, somatic mesoderm, splanchnic mesoderm and
endoderm.

ii) The section shows mid- structures namely, mesocoel, anterior

cardinal vein, dorsal aortic root, somatopleure, extraembryonic
coelom, splanchnopleure, foregut, notochord and ventral aortic
root.

Fig. 7.9: T.S. of 33 hour chick embryo after Incubation.

7.4.6 Study of Whole Mount of Chick Embryo at 72

Hours of Incubation
I. Features of whole mount of chick embryo at 72 hours of incubation

i) The 72 hours old chick embryo (Fig. 7.10) as observed in the whole
mount is seen to be affected throughout by torsion and the entire
body is seen to have turned through 90". Torsion is complete in the
chick embryo posterior to the level of heart, but the caudal portion
of embryo is not turned on its side. 83
BZYCL-134 Laboratory of Comparative Anatomy and Development Biology of Vertebrates

ii) The long axis of the embryo due to the presence of cranial and
cervical flexures, shows nearly right angled bends in the mid brain
and neck region. The mid body appears concave.
iii) Visceral arches have developed and can be seen.
iv) Mandibular arch is visible and has formed the caudal boundary of
oral depression and become more distinct.
v) Nasal pits appear as shallow depressions.
vi) Cephalization can be seen to be in process. Telencephalon has
developed and is visible.
vii) In the eye, the lens, sensory and pigmented layers can be seen to
have differentiated.
viii) Number of somites have increased to 36 pairs.
ix) Vitelline arteries and vitelline veins also make their appearance
and are visible.

Fig. 7.10: Whole mount of Chick embryo at 72 hours of incubation.

7.4 .7 Study of Whole Mount of Chick Embryo at 96

Hours of Incubation
I. Features of whole mount of chick embryo at 72 hours of incubation

i) At 96 hours of incubation the entire body of the chick embryo has been
turned through 90" and the embryo lies with its left side on the yolk
(Fig. 7.11).

ii) At the end of 96 hours the body folds have undercut the embryo so
that the embryo is seen to be attached to the yolk only by a
84 slender stalk.
Exercise 7 Studies on Chick Embryo Using Prepared Slides

iii) The yolk stalk soon becomes elongated, allowing the embryo to
become first straight in the mid-dorsal region and then in the dorsal
region.

iv) The progressive increase in the cranial, cervical, dorsal and caudal
flexures results in the bending of the embryo on itself so that its
originally straight long axis becomes C shaped and its head and
tail lie close together.

v) Paired optic cups show more developed lens.

vi) Endolymphatic ducts arise from the paired auditory vesicle.

vii) Visceral arches appear to have thickened.

viii) Appendage buds appear increased in size and can be seen to

have become elongated.

ix) The number of somites are seen to have increased to 41 pairs.

x) Allantois has also made an appearance and is visible.

xi) Omphalomesentric artery and omphalomesentric vein have also

developed and can be observed.

Fig. 7.11: Whole mount of Chick embryo at 96 hours of incubation.

7.5 TERMINAL QUESTIONS

1. With the help of labeled diagrams differentiate between the whole mount
of chick embryo at 16 hours and 33 hours of development.

2. At which stage of development does the chick embryo show

fundamental features for the formation of central nervous system and
circulatory system? 85