First Semester Comparative Vertebrate Embryology - Laboratory

Exercise 1: Gametogenesis  Egg

o Increase in size
Two type of Cell o Accumulation of yolk
 Germ Cells
o Gives rise to the gametes (Egg and SPERMATOGENESIS
the sperm cells)  Formation of male gametes
 Somatic cells  Occurs in the seminiferous tubule
o Other cells in the Bod  In association of specialized somatic cells

Primordial Germ Cells Histology of Grasshopper’s Testis

 The initial germ cells that form  Development happens in the male gonad
 Must journey to the gonads and divide  Testis divide to several lobes
mitotically. o Arranged with their blunt ends
o Spermatogonia – males facing the surface of the organ.
o Oogonia – Females o The more pointed leads to the vas
deferens
Stem Cells  Inside the lobe
 Undifferentiated cells that divide mitotically o Compartments – Cysts
producing two types of cells. o Similar cells undergoing the same
 Some will remain undifferentiated to stage of development
maintain cell line alive. o Separated by septa – connective
tissue
Gametogenesis o The more immature cysts of the
 First phase of sexual reproduction periphery
 Formation of sex cells for male and female o Shows progressive maturation
gametes
o Spermatogenesis Spermatogonia
o Oogenesis  Small, crowded, at the blunt ends of the
testis
Major Processes of Gametogenesis  Nuclei – beaded apperance of chromatin
 Cell Proliferation material
o Primitive germ layer undergo  Have large nuclei and conspicuous
repetitive mitotic division into chromatin
identical cells
 Cell Growth Primary Spermatocyte
o Cessation of mitosis in favour of  Spermatogonium undergone growth, ready
acquisition of raw materials for to divide.
synthesis of protoplasm  Located near the spermatogonia
 Cell Maturation  Very large nuclei
o Two successive cell division  Stages of Prophase 1
o Accompanied by chromosomal o Interphase – Fine granular, show
division that will lead to reduction of single pair vascular tissue
chromosome number to half. o Leptotene – Chromatin crust finely
beaded filaments, filamentous,
Changes in appearance chromosome uncoiled- completely
 Sperm fill the cell
o Elongation that is more pointed in o Zygotene – have thickened region-
one part synapsis, synaptonemal complex,
o Losses cytoplasm, leaving minimal paired chromosomes takes the
amount shape of a “bouquet”, double
o Flagellation

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 First Semester Comparative Vertebrate Embryology - Laboratory
thickness of chromosomes means
advance pairing.
o Pachytene – Complete pairing and
eventual exchange of chromosomal
arm, the cells seems to be haploid,
chromosomes shorten and thicker.
o Diplotene – Chromosomes partly
split, relaxation of pairing, presence
of “chiama”, configuration of “x”,
cross, or loop
o Diakinesis – progressive
condensation by means of coiling,
chromosomes are in their shortest
and thickest appearance.
 The nucleus goes through the metaphase,
anaphase, and telophase – first maturation
division.
Secondary Spermatocyte
 Smaller than the primaries
 Chromosomes are not that evident
Spermatids
 Half the size of the secondary spermatocyte
 Possess haploid set of chromosomes but
not capable of functioning as male gamete
 SPERMIOGENESIS – transformation of
spermatids to spermatozoa.
o May be observed in the inner region
of the lobe
o Major nuclear and cytoplasmic
changes that will eventually
terminate in the formation of
functional gamete.
o Elongation of spermatid – cytoplasm
sloughed off.
o Tail Sheath – surrounding the tail
filament, grows back from
centrosome
o Head – Condensation and
elongation of chromatin mass
o Middle Piece – contains the
centrosome and mitochondria
o Tail
Spermatozoa
 Exhibit long, thin, darkly staining head with
distinct tails.
 Tail measures 7-8 times longer than the
head.
 Abundant at the distal region of the testis.
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Histology of Frog’s Testis
Mesorchium
 Portion of the peritoneum that suspends the
testis to the dorsal body wall.
Tunica Albuginea
 Outer connective tissue that covers the
testis
 Known as septula or intertubular connective
tissue
Seminiferous tubule
 Much-coiled tubule containing the clusters
of spermatogenic cells undergoing the same
stage of development and the sertoli cells
 Connected to the vas deferens
 Cells going through same stage are found in
a cluster or group within the tubule.
Spermatocysts
 Cluster of similar cells undergoing the same
stage of development
 Surrounded by follicle cells
 Formed from primary spermatogonium and
its associated follicle
Spermatogonia
 Small, closely packed cells with granular oval
nucleus.
o Primary Spermatozoa – fastened to
the walls of the seminiferous tubule
o Residual Spermatozoid – Source of
germ cell proliferation
 Active Spermatogenesis – short season in
summer, begins with a rapid succession of
mitotic division
 The millions of sperm came from
comparatively large number of gonia by
repeated mitotic division, maturation, and
spermatogenesis.
 The seminiferous tubules do not contain
permanent population of sperm cells.
Primary Spermatocyte
 Very large cell, spherical, with large
vesicular nuclei
 Characteristic meiotic prophase
configurations of chromosomes can be seen.
Secondary Spermatocyte
 Result of the first meiotic division
 Half of the size of the primaries and lie
towards the lumen of the tubule.
 First Semester Comparative Vertebrate Embryology - Laboratory

 Have darkly staining nucleus, and clear and  Spherical structures filled with
visible cytoplasm. spermatogenic cells.
 In mammalian, convoluted tubules and
Spermatids made up of specialized seminiferous
 Even smaller than the preceeding cells epithelium
 Possess a condensed nucleus of irregular o Sertoli cells
shape. o Male Sex cells undergoing
 During spermiogenesis, the inner of the two spermatogenesis and
spermatid centrioles passes into the nucleus spermiogenesis
while the other one give rise to the  Contains the cells with conspicuous nuclei
flagellum. call the Interstitial or Leydig cells
Spermatozoa o Responsible for the production of
 Average length of 0.03mm. testosterone.
 Has an elongated solid staining head with an
anterior chromosomes Spermatogenic cycle
 The tail appears like gray filamentous  Last a total of 48 days, successive cycles start
extension every 12 days
 The maturation depends upon the external  Total of 4 overlapping cycles at every site of
sources of nutrition, grouped into 25-40 cells the tubule.
converging into the relative columnar type
basal cell – sertoli cells. Sertoli Cells
Sertoli Cells  Located near the spermatogonia
 Supplying nutrient to the cluster of sperm  Has distinct outline and large, oval, often
until they are ejaculated. indented nuclei with compound nucleoli
 Associated with the primary spermatogonia  Surrounded by spermatozoa
and spermatocyte as follicle cells.  Two types of sertoli cells
 In frogs, sperms remained attached o Active- fulfilling the normal function,
throughout the winter, as long as six pyramidal in shape, nucleus
months. perpendicular to the tubule wall
Interstitial Cells o Inactive (resting)- polygonal in
 Small clusters of cells found interspersed shape, nucleus parallel to the dorsal
with the intertubular connective tissue body wall.
Ova SPERMATOGENIC CELLS:
 Egg cells seen between the seminiferous Spermatogonia
tubule.  Most primitive of the male sex cells
 Making the organ “ovotestis”  Three criteria:
o Rests on the basement membrane
HISTOLOGY OF MOUSE TESTIS and interrupted at intervals of
 Stained in haematoxylin and eosin Sertoli cells
o Nucleus – blue (basic, stained by o Appear as thin layer of cells with
haematoxylin) compact deeply staining nuclei
o Cytoplasm – pinkish orange (acidic o Nuclei tend to be quite small,
by eosin) comparable with the size of the
Tunica Albuginea spermatids.
 Thick layer of fibrous connective tissue with  Three types
a covering of mesothelial cells o Type A- chromatin particles appear
 Subdivides the testis into a number of fine and evenly dispersed, in
lobules each containing seminiferous eccentric position, usually in the
tubules germ cell line
o Type B- chromatin is identified with
Seminiferous tubule the presence of coarse granules or

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 First Semester Comparative Vertebrate Embryology - Laboratory
flakes, most common gonia, last
stage before primary spermatocyte
o Intermediate- chromophilic
chromatin in the form of delicate
crustor flake, has prominent nuclei
appearing in the center.
Primary Spermatocyte
 Significantly large in size
 Nuclei larger than the spermatogonia
 The chromosomes are very much visible and
the nuclear membrane is disintegrated
already.
 Visible stages:
o Interphase – Similar to gonia B,
finely granular nucleus with pale
nucleolus, exist for about 6 hours.
o Prophase I
o Metaphase I
o Anaphase I
o Telophase I
Secondary Spermatocyte
 Result of first meiotic division of the primary
spermatocyte
 Pale stained granular chromatin
 Twice the diameter of spermatids.
Spermatids
 Result of second meiotic division
 Lie next inward beyond the spermatocyte
layer in the seminiferous tubule
 Nuclei quite round, small size (2/3 of primary
oocyte), pale staining properties, and
position above the level of primary
spermatocyte – IMPORTANT CLUES
 DNA undergoes progressive condensation
 Decrease of nuclear size due to the
elimination of nuclear sap.
Spermatozoa
 Deeply stained when matured
 The heads are buried at the Sertoli cells
 Matured spermatozoa becomes free of the
nurse cells and pass into the ducts
*The histology of Chick’s Testis is also the same as
mouse’s testis
OOGENESIS
 Process of producing egg
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 Products of meiotic division which is
responsible for haploid condition
 Formation of Polar bodies
o Produced after first meiotic division
o Smaller than egg
o Due to unequal cytokinesis
o Provides the ovum a much greater
amount of cytoplasm and store
food.
o Non functional
 After meiosis, the ootid quickly change into
the mature egg cell or ovum
 It takes place within the follicles of the ovary
o The ovary is covered by continuous
sheet of epithelium called the
germinal epithelium
Mesovarium
 The mesentery that attached the ovary to
the body wall
 Route for blood vessels to and from the
ovary.
Ovary
 Divided into two parts
o Central Medulla – consist largely of
connective tissue and blood vessels,
appear more or less continuous with
the mesovarium
o Peripheral cortex – business part of
the ovary
HISTOLOGY OF FROG OVARY
 A large sac of yolk with heavier and larger
granules concentrated toward the animal
hemisphere
 Surrounded by a coat necessary for retaining
the shape, aids in cleavage and gastrulation
Mesorvarium
 Double-layered extension of the peritoneum
 Attachment to the dorsal body wall
Theca externa
 Outermost connective tissue surrounding
the ovary
 Continuous with the mesovarium
 Ruptures during ovulation
Theca interna
 Inner connective tissue surrounding the
ovary and each egg
 First Semester Comparative Vertebrate Embryology - Laboratory

 Another layer of connective tissue below the

Follicle Cells Tunical albuginea
 Derived from oogonia  Made up of connective tissue fibers and
 Contains oval and granular nuclei spindle shaped cells.
 Surrounds the developing oocyte
Oogonium
Ovarian Follicle  Potential ovum with vesicular nucleus with a
 Collective term for theca externa, theca prominent nucleolus
interna, follicle cells  Embedded at the stroma and in clusters.

Oogonia Primary Follicle Stage

 Located at the periphery of the ovary and  Surrounded with single layer of flattened
much smaller than the oocyte follicular cells.
 No pigment and contains a little yolk
 Each cluster of oogonia represents a future Structures of Follicle Stage Cells
ovarian unit.  Follicles
 The majority of oogonia never mature into o Single layer of cells that surrounds
ova. the developing germ cell
o Have dark stained nuclei and maybe
Oocytes squamosal or low cuboidal.
 Bigger than the oogonia  Oogonium
 Lightly stained at the center is the germinal o Potential ovum surrounded by single
vesicle or immature nucleus. layer of follicle cells
 Distinction to primary and secondary oocyte o Spheroid and has a vesicular nucleus
is not possible with a prominent nucleolus

Vitelline Membrane Growing Follicle Stage

 Non-cellular transparent membrane  Immediately succeeds the primary follicle
between follicle cells and each mature egg stage
cells.  Proliferated by mitotically dividing cells and
 Derived from ovum and follicle cells are now several layers thick.
 Developed during maturation process.  Increased in size and complexity.

Perivitelline Space Structures

 Fluid-filled space formed between the egg  Follicle Cells
and the vitelline membrane after o Several layers thick, low columnar or
fertilization cuboidal cells enclosing the oocyte
 Shows compensatory shrinkage.  Antrum
o Follicular cavity
HISTOLOGY OF CAT OVARY o Formed by the union of spaces
between the follicle cells
Germinal epithelium o Precipitation of proteins in the fluid
 Forms the outer surface may give the cavity a granular
 Epithelial lining of the ovary appearance.
 Cuboidal cells  Liquor Folliculi
o Viscous follicular fluid secreted by
Tunica albuginea follicle cells.
 Layer of fibrous connective tissue below the  Oogonium or Primary Oocyte
germinal epithelium o The probable stage of development
Stroma of the cell in the growing follicle

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 First Semester Comparative Vertebrate Embryology - Laboratory

o Hardly distinguishable from each o Exhibited by the presence of

other collapsed zona pellucida
 Zona Pellucida o Corpus atretica – mass of tissue
o Deeply staining membrane regressing corpus luteum
surrounding the egg cell and  Corpus lutea
separating it from the follicle cells o Masses of granular tissue
o Result from the ovulation of egg
Graafian follicle cells from follicles.
 Large follicle that bulges on the free surface o Responsible for the production of
of the ovary progesterone
o Lightly stained, follicles converging
Structures at the middle portion
 Antrum o Lutein Cells, Capillaries, Capsule
o Fluid-filled cavity, now it is bigger.  Corpus Albicans
o Lined by stratum granulosum, thick o Found in the central portion of the
layer of follicle cells ovary.
 Cumulus Oophorus o Non-functional corpus luteum when
o Eccentric mound of follicle cells fertilization does not take place
surrounding egg cells o Pale stain in its central portion
o Includes the oocyte, zona pellucida,  Periovarial Membrane
amd surrounding granulose cells o Peritoneum of the ovary which
 Primary oocyte continues into the Oviduct
o Stage of development at this point  Periovarial Space
o Has a heavy nuclear envelope o Space between the periovarial
 Stratum Granulosum membrane and the ovary
o Layers of follicle celss boredering the  Oviduct
Graafian, excluding the cumulus o Convoluted structure that is found
oophorus alongside of the ovary
 Corona Radiata o Attached to the periovarial
o Layer of follicle cells of the cumulus membrane
oophorus that lie nearest to the
oocyte
 Zona Pellucida
 Liquour Folliculi
 Theca folliculi
o The stroma immediately adjacent to
the follicle
 Theca interna
o Inner layer between the theca
interna and the granulose cells
o Richly vascularized
o Cells are secretory
 Theca Externa
o Outer layer of the capsule composed
of spindled shaped dense connective
tissue fibers
 Atretic Follicles
o Unsuccessful follicles undergoing
degenerative process
o Shows antrum scattered follicle cells