Addis Ababa University

College of Veterinary Medicine and Agriculture

Department of Clinical Studies

Course Title: Theriogenology I

(Vetm4161)

Tefera Yilma (PhD)

March, 2021

4/4/2021
 Sub-topics:
Functional anatomy of female genital system
Origin and early development - sexual differentiation
 Functional anatomy of female reproductive system

Season and estrous cycles

Reproductive hormones
Physiology of fertilization/gestation
Pregnancy diagnosis
Pregnancy diagnosis

4/4/2021
Anatomy of female genital system
 Origin and early development of the reproductive organs,
sexual differentiation
…developed from the outer part of intermediate mesoderm of
the germ layers
Early embryonic development:
primordial germ cells migrate into the Urogenital ridge -
common precursors of the urinary and genital systems and of the adrenal
cortex

Fetal reproductive system consists of:

 two sexually non-differentiated gonads - bipotential system or
sexual indifferent stage - apparently identical
 two pairs of ducts (Muellerian and Wolffian), urogenital sinus and a
genital tubercle
… embryonic bisexuality
4/4/2021
  Origin and early development … (cont’d)

differentiates into a male and female system (2 months after

fertilization) -  internal and external genitalia
Gonads:  either ovary/testis
Ducts:
Muellerian (paramesonephric) duct develops into
•oviducts,
•uterine horns, uterus
•cervix and vagina
 Wolffian (mesonephric) duct develops into
•epididymis,
•vas deferens, seminal vesicles and the ejaculatory ducts
Urogenital sinus and genital tubercle  the external
genitals in both sexes
4/4/2021
  Sexual differentiation (cont’d)

4/4/2021
 Sexual differentiation (cont’d)

The sex of the fetus depends on

•inherited genes (chromosomes)
•the formation and maturation of accessory reproductive ducts
Genetic sex
 determined by the presence or absence of the Y-
chromosome
•the Y- chromosome determines the development of testes
and maleness (SRY, Sex determining Region of Y -
Chromosome)
• … independent of the ovum
 Ovum fertilized by an X-spermatozoon, the offspring is XX, a female
 Ovum fertilized by a Y-spermatozoon, the offspring is XY, a male

4/4/2021
 Sexual differentiation (cont’d)

Gonadal sex - the presence of normal ovaries or testes

…Male phenotype sexual differentiation is directed by the
function of the fetal testis

Testis determining gene (TDG)

 H-Y Ag (= Histocompatibility of Y antigen) /
 SRY gene (= Sex Determining Region of Y Chromosome)

… as the testes grow leydig cells produce testosterone

(Testosterone and 5-dihydrotestosterone stimulate the growth and
differentiation of the Wolffian ducts in the male)

…the Wolffian ducts develop into the male reproductive tract

4/4/2021
  Sexual differentiation (cont’d)

… The müllerian ducts regress by the anti-müllerian

hormone (Müllerian–inhibition factor, MIF) from the
Sertoli cells

 In the absence of the testis:

•female differentiation ensues
•no need of extra stimulatory or inhibitory mechanism

 …In the female fetus

•there is a developing ovary
•no anti-müllerian hormone

4/4/2021
 Sexual differentiation (cont’d)
•The müllerian ducts develop into the female reproductive tract
• the fallopian tube (oviduct)
• the uterine horns
• the uterus
• the cervix and
• the upper (cranial portion) vagina

•…the Wolffian ducts degenerate (b/c the ovary does not secrete
testosterone)
•When a normal female fetus is exposed to androgens during the
period of differentiation of the external genitalia, an apparent male
can result

4/4/2021
 Sexual Differentiation

4/4/2021
Sexual Differentiation

5α-R/DHT

4/4/2021
 Anatomy and function of the female reproductive system
Knowledge is useful in :
1) identifying and correcting situations leading to poor reproductive
efficiency/infertility
 early detection of problems
 re-synchronization of non-pregnant cows
for breeding
…→success of a reproductive management

2) the application of reproductive

biotechnologies to enhance the
reproductive performances of farm
animals- genetic improvement
Genetic improvement for:
 improved meat quality or yield
 increased milk production and composition
 reduced susceptibility to disease, environmental stress etc.
4/4/2021
  Anatomy and function…(cont’d)

The female reproductive

tract consists of:
Vulva, vestibule, vagina
Cervix, uterus, uterine horns,
The oviducts
Ovaries

Except for the vulva, all parts are located beneath the rectum
… rectal palpation (easy)

4/4/2021
 Anatomy and function…(cont’d)

Four basic functions of cow's reproductive system:

Produces ova (eggs) which provides half of the genetic
makeup of the offspring
Provides an environment and conditions for the fertilization
of the ova
Provides a place for the nourishment and fetal development
Provides a mechanism for the birth of the calf

4/4/2021
Anatomy and function…(cont’d)

The vulva
External part (“Entranceway”) of the female reproductive tract,
consists of:
(a) the labia (lips)
•labia minora, inner folds or lips of the vulva
•labia majora, outer folds or lips of the vulva
(b) the clitoris, homologous to the glans penis (same embryonic
origin as the penis)
Main functions(vulva): Labia
•passage of urine Clitoris
•opening for mating
•part of the birth canal

4/4/2021
  Anatomy and function…(cont’d)

Vulva: Sensitive to  estrogen concentration - swelling,

redness due to blood flow …estrus detection when coupled
with other signs
Swollen Vulva in Sow, Cow, & Bitch

 Clitoris: homologous to the glans penis (same

embryonic origin as the penis)
4/4/2021
 Anatomy and function…(cont’d)

The vestibule
•Common to the reproductive and urinary systems
•Joins the vagina at the external urethral orifice
The vagina
extends from the urethral opening
to the cervix
vaginal wall consists of epithelium, muscular layer and serosa
 surface epithelium is composed of glandless stratified,
squamous epithelial cells, except in the cow: some mucus
producing cells in the cranial part
4/4/2021
 Anatomy and function…(cont’d)

 Vagina:- Muscularis is well supplied with

•blood vessels
•nerve bundles
•dense connective tissue

Functions (vagina):
•A copulatory organ
•Site of semen deposition during natural mating (service):
anterior portion of the vagina
•Serves as reservoir for semen until transported to the uterus
through the cervix
4/4/2021
 Anatomy and function…(cont’d)

 Cervix (Cervix uteri)

The muscular “valve” or “control gate” between the uterus and

the vagina (5 to 10 cm long with diameter of 2 to 7 cm)
Characterized by a thick /dense wall
and constricted lumen
landmark when AI

Made of muscular folds that traps

foreign (invading) materials
Closed except during estrus and
parturition (calving)

4/4/2021
 Anatomy and function…(cont’d)
 …Cervix – peculiar arrangement:

In ruminants:
•the cervix is made of transverse/spirally interlocked ridges
known as annular rings
•helps seal the uterus from contaminants (external environment)
In the sow:
•a corkscrew arrangement of the rings, adapted to the spiral
twisting of the tip of the boar’s penis
In the mare: characterized by prominent folds in the mucosa,
projecting folds into the vagina
•the mucosa of the cervix is composed of secretory epithelial
cells in all species of animals
…the cervix opens anteriorly into the uterine body
4/4/2021
  Anatomy and function…(cont’d)

Functions (cervix):

 Semen deposition (anterior cervix)

during AI
 acts as sperm reservoir
 facilitates sperm transport
through the cervical mucus to
the uterine lumen

 Selection of viable sperm, preventing the transport of

nonviable and defective sperm

4/4/2021
  Anatomy and function…(cont’d)

 The uterus and uterine horns

 The uterus consists of

• two uterine horns (cornua)
•a body and
• a cervix (neck)
U- horns
 the extensions on either side of the uterus
that lead to the oviducts - Curl - like ram horns
 the proportion, the shape and arrangement
of the horns vary according to species

4/4/2021
  Anatomy and function…(cont’d)

Two basic types of uteri in farm animals:

1) the bicornuate uterus in the sow, cow, doe and ewe

 the body of the uterus is short, long uterine
horns
 the sow has long folded/convoluted (as long as
4 to 5 feet) uterine horns

2) the bipartite type (uterus bipartitus) in mare

 prominent uterine body is appreciable

4/4/2021
 Reproductive System of the Sow - Top View
Uterine Body

Uterine Horn

Uterus

Ovary
Infundibulum
Oviduct
Cervix

Bladder
Opening
of Urethra

Vagina
Vulva

4/4/2021
 Anatomy and function…(cont’d)

•Both sides of the uterus are attached to the pelvic and abdominal
walls by the uterine ligament (broad ligament, mesometrium) and
suspended within pelvic cavity (non-pregnant)
4/4/2021
Anatomy and function…(cont’d)

Functions of the uterus

The endometrium and its fluid* play major role in the reproductive
process:
1) Major site of semen deposition during AI
2) Sperm transport from the site of ejaculation to the site of
fertilization
3) Muscle of the uterus aids in expulsion of the calf at birth

• *As spermatozoa are transported through the uterine lumen to the oviducts,
they undergo capacitation in endometrial secretion

4/4/2021
Function of the uterus (cont’d)

4) Regulation of the function of the CL:

The uterus produces luteolytic
substance PGF2 that selectively
transferred from uterine vein
to the closely adherent ovarian
artery and reaches the ovary in
much greater concentrations than
that in peripheral blood Luteolysis
5) Implantation, pregnancy, and parturition

4/4/2021
The Oviducts (fallopian tubes)
Lie between each ovary and tip of the adjacent uterine horn
Tissue layers: serosa, muscularis and mucosa

Functional segments of the oviduct:

 the finger-like fimbriae (serves to
capture the ovum after ovulation)

 the infundibulum (funnel-shaped

opening near the ovary), receives the
ovum
dilated ampulla, joins the isthmus
at ampullary-isthmus junction
(site of fertilization)
the isthmus, narrow portion of the oviduct connecting the oviduct with the
uterine horn

4/4/2021
Anatomy and function…(cont’d)

the isthmus (function)

•filtering out dead sperm
•transport of motile sperm to the site of fertilization
•joins the tip of the uterine horn at the utero-tubal junction

Function of the oviduct

 conveying the eggs and spermatozoa
 Site of fertilization and early cleavage of fertilized eggs
 …  the oviduct and oviductal fluid provides an optimal
environment for union of the gametes and for early embryo
development

4/4/2021
  Anatomy and function…(cont’d)

The oviduct is endowed

with mucosal epithelial cells
and ciliated cells that beat toward
the uterus
The activity of ciliated cells,
coupled with oviductal
contraction, keeps ovulated eggs
in constant rotations:
• bringing egg and sperm
together (fertilization)
• preventing oviductal
implantation
• facilitating embryo transport

4/4/2021
  Anatomy and function…(cont’d)

 The Ovary
•the primary reproductive organ
in the female

•suspended from the broad ligament

(mesovarium)
•lie near the tips of the curved uterine
horns, remains in the abdominal cavity
(unlike the testis)

Functions:
• Egg release
•Endocrine (steroidogenesis- estrogen and progesterone) 
hormones involve in the regulation of the estrus cycle and
pregnancy
4/4/2021
 Anatomy and function…(cont’d)
Functions: Gametogenesis/Oogenesis Production of ova

Estrogens

Ovary
Steroid H. Progestins

Hormonogenesis Androgens

Nonsteroidal H.
(Relaxin)

4/4/2021
 Anatomy and function…(cont’d)

•Ovaries composed of
an inner medulla and Cortex
cortex (the predominant
tissue of the ovary)
Medulla
Cortex
•ovum production Cortex

•hormone production
Cumulus-oocyte
complex
Corpus
Medulla consists of haemorrhagicum

•connective tissue
•nervous and vascular systems
→reach the ovary through4/4/2021
the hilus
Follicles – produce estrogen
• Primary follicles – with centrally
located oocytes and a single layer
of granulosa cells
•Secondary follicles- with increased
number of cuboidal granulosa cells
• Tertiary (vesicular) follicles
with multilayered granulosa cells
that secrete a fluid, the liquor
folliculi (causes formation of a large
fluid-filled cavity-the antrum)
•Graafian follicles- follicular cells
increase in size; the antrum is
increasingly filled with follicular fluid

• Preovulatory follicle- a blister-like

structure protruding from the ovarian
surface due to rapid accumulation of liquor folliculi and thinning of the granulose layer
4/4/2021
Corpus luteum
 produces progesterone, relaxin,
oxytocin
 develops after the collapse of the
follicle at ovulation
 hypertrophy and luteinization of the
granulosa cells after ovulation – effect of
LH
•CL weight (size) increases between 3
and 7 days of the estrous cycle
•Size maintained constant until day 12 (by
the effect of basal level of LH)
•Regression begins day16/17 due to the
action of PGF2
4/4/2021
Season and growth - estrous cycle
The onset of puberty:
“The onset of such changes, in which
young female animal becomes sexually
mature and able to reproduce (shows
evidence of recurring or
cyclical periods of sexual receptivity)”
•Sexual maturity
•Release of gametes
•Manifestation of sexual behavior sequence
Maturity of reproductive endocrine system
(Hypothalamic-Pituitary-Gonad Axis)
… Development of the hypothalamic
neuron- release of GnRH
4/4/2021
... Maturation of regulation system

 Hypothalamus (GnRH)

 Hypophysis/Pituitary
(FSH, LH)

 Gonad (Estrogen,
progesteron)

 Endometrium (PG)
 Maturation of hormone feedback-system
= Pos
Hypothalamus
= NEG!

GnRH

Progesterone
estradiol-17ß

Hypopysis
FSH/LH Inhibin

Ovary

Oxytocin PGF2

Uterus

4/4/2021
Season & cycles (cont’d)
Maturation of GnRH release centers of the hypothalamus:

1. Tonic release center (continuous, basal release center)

 pulsatility of GnRH and gonadotropins- LH and FSH
• in amplitude
• in frequency
Sensitive to negative feedback of steroid hormones (P4 & E2)

2. Surge center (controls the short-lived massive

release of gonadotropins) estrus period!)
 in sensitivity to estradiol, sensitive to
Positive Feedback of increased level of estradiol
GnRH, LH, FSH
•in follicular development, maturation steroidogenesis (estradiol)
•Estradiol reach threshold concentration to cause positive feedback
•…  LH surge…ovulation… CL formation
4/4/2021
4/4/2021
Season & cycles (cont’d)
The changes at puberty depend upon the activity of the ovaries
(ovarian cycle):
•production of the female gametes (ova)
•synthesis of hormones (E2 & P4)

Initiation of puberty is a function of the

animal’s age and maturity

 The age at first estrus/ puberty is affected by nutrition and

season of birth
•… at puberty the genital organs increase in size (growth
rate is accelerated)
4/4/2021
 Female domestic species reaches the age of puberty at the
following times:

Species Average Puberty age

Mare 14 mo 1-2 years

Cow 11 mo 7-18 months

Ewe 7 mo 6-15 months

Doe female goat 7 mo 4-8months

Sow 7 mo 6-8 months

Bitch 7 mo 6-12 months

Queen cat 7 mo 8-9 months

4/4/2021
Season & cycles (cont’d)

…The time of onset of puberty is determined by the

individual’s genotype (genetics), inherent timing/ “biological
clock”:

E.g.: Breed differences

 Smaller breeds of animals tending to be slightly more
precocious
•Jersey earlier than Brown Swiss
•Angus earlier than Charolais
•Ponies earlier than draft horses
•Dairy cattle earlier than beef cattle
4/4/2021
Season & cycles (cont’d)

…The inherent timing (“biological clock”) is influenced by

external factors:
1. Nutrition/ Body weight: the age of puberty is closely related to
body weight (30-50% of adult weight)
E.g.- Dairy heifers reach puberty at 30-40% of adult weight
-Beef heifers reach puberty at 45-55% of adult weight
-Sheep reach puberty at 40-50% of adult weight

Threshold body size is more important than age:

• Well fed with good growth rate (earlier)
• Poorly fed with slow growth rate (later)
Overfeed earlier than underfeed flushing stimulate puberty
Poor nutrition delays puberty
4/4/2021
 Onset of puberty in relation to breed, age and
weight
Breed Age (days) Weight
(kg)
Jersey 322 219

Charolais 398 303

Holstein 306 288

Hereford 408 274

Angus 360 255

4/4/2021
2. Season of the year:
In mare and ewe, seasonal breeders, the age at which puberty
occurs influenced by the effect of season of the year
•E.g.: Ewes born latter in the spring reach puberty faster than earlier born
ewes

3. Climate/Temperature:
 Extreme temperatures extend onset of puberty
4. Disease:
 interfere with feeding and utilization of nutrients
 influence growth rate  delay the onset of puberty
5. Social factors:
Exposure to opposite sex generally hastens puberty
•E.g.: Boar exposed gilts reach faster than alone -
Pheromone effect!
Group gilts reach faster
4/4/2021
Breeding seasons, seasonal breeders
Mating only during certain times of the year
…to allow for the births/ progeny a time optimal for the
survival in terms of temperature, food and water

Mating is regulated by length of daylight

(photoperiod) and thus season

Photoperiod affects seasonal breeders

through changes in melatonin secretion
by the pineal gland (epiphysis) that alters
GnRH release by the hypothalamus
…  Pineal gland controls reproduction and timing of puberty
by influencing the release of GnRH, FSH, LH
4/4/2021
Seasonal breeders (cont’d)

Two groups:

1. Short-day breeders - Sheep

 Enter into the breeding season during shorter photoperiods
 Cycle when the length of daylight shortens (fall/autumn and
winter)
 … anestrus in spring and summer

 Short day breeders: Sheep, goat, fox, deer

4/4/2021
 Seasonal breeders (cont’d)
Photoperiods stimulate
gonadal activity
 Stimulation of the retina,
by the amount of light
present
Sheep - secretion of
melatonin stimulates GnRH
release

…Melatonin is released during the

night, as days shorten and nights
lengthen in the autumn, the
duration of the nocturnal
melatonin peak is increased
4/4/2021
In sheep when light is abundant, an
excitatory pathway is active inhibiting
melatonin release

•An intact pineal gland for a normal

photoperiodic response to altered daylight
patterns

Rhythmic administration of melatonin to

adult ewes exerts a similar effect to
increased hours of darkness by inducing
the onset of the breeding season and
cause changes in prolactin concentrations
in the plasma that are similar in those
following exposure to short days

Sheep and goats in tropical climates may

breed throughout the year
4/4/2021
Seasonal breeders (cont’d)

2. Long day breeders - Mare

 The mare is a seasonal breeder, switched-on by

increasing day length
 Cycle when days get longer (spring/summer) and
are anestrus in fall/autumn and winter

 Long day breeders: Horse, hamster, mink

4/4/2021
Seasonal breeders (cont’d)

 In mares melatonin levels increase during hours of

darkness
 The effect of melatonin on reproductive systems of long
day breeders is anti-gonadotropic (GnRH, FSH, LH)

 …Melatonin inhibits the secretion of the

gonadotropic hormones LH & FSH, due to inhibition
of Gonadotropin-Releasing Hormone from the
hypothalamus
4/4/2021
Reproductive pattern (mono-, di-, polyestrous and
seasonal polyestrous)
The reproductive cycle: a series of events occur in a definite order
over a period of days
 E.g. Cow: 21 days in averages (ranges 18-24d)
Cattle are polyestrous (polycyclic) throughout the year
•…the reproductive tract is prepared for estrus or heat and
ovulation (egg release)
•In polycyclic animals (cow, sow), the recurring cyclic activity is
interrupted by pregnancy, lactation and pathological conditions
In seasonally polycyclic species (mare, ewe, cat) or monocyclic
(bitch) there are periods of sexual quiescence or anoestrus

4/4/2021
 Classification of animal species in relation with
reproductive patterns
Species type of the estrous cycle

Cow polycyclic throughout the year

Sow polycyclic throughout the year
Ewe seasonal polycyclic
Mare seasonal polycyclic
Cat seasonal polycyclic
Bitch mono/di-cyclic
Camel seasonal polycyclic

4/4/2021
THE ESTROUS CYCLE – reproductive pattern – cattle
After puberty- regularly recurring changes
in the reproductive system
Ultimate goals:
•producing fertile egg(s)
•creating favorable conditions for
further development of the zygote
in the oviduct and uterus
The estrous cycle: the period of time
from the beginning of one estrus period
to the beginning of the next estrus period
Estrous cycle length: 18 to 24 days,
average 21 days for cows
The reproductive tract is prepared for
estrus or heat (the period of sexual receptivity) and ovulation (egg release)

4/4/2021
Based on internal changes estrous cycle is divided into two:
1.Ovarian cycle, cyclic changes in the ovary:
•Follicular phase (coincides with pre-estrus and estrus period) - Estrogen
•Luteal phase (coincides with met- and di-estrus period) – Progesterone

 The ovarian cycle is based on the interactions of the CNS, hypothalamus with
adenohypophysis, ovary and uterus.
The mediators of these interactions:
- Neurotransmitters (CNS)
- Releasing /Inhibiting hormones (Hypothalamus)
- Gonadotropins (Anterior pituitary)
- Gonadal steroids (Ovary/Testis)
- Prostaglandin (Uterus)
The end effect the ovarian cycle is creating a condition of
“standing heat/estrus” associated with development and
release of fertile egg (s)
4/4/2021
2. Cycle of the genital tract (cyclic changes in the mucus membrane
of the genital tract): controlled by the ovarian cycle !
The cycle of genital tract refers to the overall estrous cycle related
alterations (changes) in the
 oviduct, uterine horns
 particularly uterus (uterine cycle)
 cervix,
 vagina and vulva
Two phases of the uterine cycle:
 the proliferative phase (coincides with follicular phase and
time of estrogen dominance)
 the secretion phase (coincides with the luteal phase and time
of progesterone dominance)

4/4/2021
Regulation system
 Hypothalamus (GnRH)

 Hypophysis
(FSH, LH)

 Gonad (Estrogen,
progesteron

 Endometrium (PG)
4/4/2021
Based on easily recognizable sexual behavioral changes (four
phases)
•Pro-estrus/Pre-estrus
•Estrus period
•Metestrus/postestrus
•Diestrus

1) Proestrus, Pre-estrus (days 18/19-20)

•the period just prior to estrus (immediately
preceding estrus)
•extends from the time of commencement
of sexual behavioral changes to the time when
the cow accepts the bull to be mated for the
first time
•the secretory activity of the CL declines
•the genital organs are dominated by the
action of estrogen, produced by the developing
follicles
4/4/2021
Pre-estrus (cont’d)
Estrogen effect
•stimulate blood supply to the genital tract
•changes in the reproductive system
•changes in psychosexual behavior
Symptoms of pre-estrus are:
•ridding (mounting) of animals of the herd (group)
•increased desire of communication/seek of contact with other animals
•slight swelling of the vulva
•hyperaemia of the vagina and vulva
•increased amount of secretions from the mucus membrane (cervix and anterior
vagina)
•collection of mucus in the anterior vagina
•relaxation of the cervix
•increased contraction of the uterus
•follicular growth, regression of the CL of the previous cycle
•enlargement of the uterus
•congested and oedematous endometrium

4/4/2021
2. Estrus (Day 0)
 Female animal stands to be ridden (mounted) by another cow or is receptive
to be mated by the bull “standing heat”
•the only readily identifiable phase of the estrus cycle
•the onset and end are accurately measurable points in the estrus cycle  used
as baseline for determining cycle length
 Estrus lasts 18 hours (average ), may vary from 2 to 30 hours depending on:
a) Exogenous factors
•season
•temperature (shortening of the estrus period)
•number of cows that are in estrus at the same time
•insufficient feeding and inappropriate light condition
•presence of a bull in a herd intensify the external manifestation of estrus
and the estrus period will also be short
b) Endogenous factors
•breed (Bos tarsus, Bos indicus x Bos tarsus = 16 to 18 hours, Bos indicus
(Zebu) = 5 hours, mainly in the night
•age (shorter in heifers), milk yield
4/4/2021
 …Estrus manifestation is more pronounced (marked) in heifers than cows
Symptoms:
•increased uterine, cervical and vaginal glands secretion, discharge of clear
and transparent mucus (frequent genital discharge of transparent mucus, elasticity
causes to hang in complete clear strands from the vulva to the ground, adheres to the
tail and flanks )

•hyperemic vaginal epithelium and endometrium

•the cervix is relaxed
•the female animal seeks out the male and “stands” for mating (standing
heat/estrus) = primary sign!
•restlessness
•reduction in the time spent eating, ruminating
•reduced milk-yield
•after service the cow stands with raised tail and arched back (this posture
indicates that mating has occurred)

4/4/2021
 Ovulation occurs near the end of
estrus in all domestic species except Graafian follicles
in the cow (about 10 to 12 hours after
the end of estrus)
 Ovulation is spontaneous process
in all domestic species except in the
cat, camel and rabbit
 Pro-estrus and estrus referred to as
the follicular phase of the ovarian
cycle, coincides with the proliferative
phase of the cycle of the genital mucus
membrane
During follicular phase :
 there is follicular growth,
development and maturation
 estrogen dominance
 functional CL is absent

4/4/2021
3. Metoestrus, Post-oestrus (days 1-2/3)

 immediately following estrus and ovulation,

succeeding estrus

 from the refusal of the male until the

cease of internal and external estrus
symptoms (contraction of the uterus,
hyperaemia, increased secretions
in the cervix, vagina and vestibules,
closing of the cervix)

 formation (proliferation) of CL:

granulosa cells change and become
the lutein cells of the corpus luteum,
by the action of LH

4/4/2021
Metoestrus …(cont’d)

 Reduced secretion from the uterine,

cervical and vaginal glands

 Metoestrus may be associated with

post-estural bleeding, small amount
of bloody mucus usually found on
the underside of the tail or near the
pins area about 2 days after the end
of estrus (50 to 60 % of the cows)
 This discharge is a good indication
that the cow was in heat about 2
days before, has no relationship to
success or failure of conception!

 Metestrus will be transformed into sexually indifferent period (dioestrus/

interestrus) or occasionally into anestrus

4/4/2021
4. Diestrus (days 3-17/18)

 the lengthiest period of the estrous cycle

 the period of the corpus luteum, between

post-estrus and pre-estrus

 Met- and diestrus coincides with the

luteal phase of the ovarian cycle and
secretion phase of the cycle of the genital
mucus membrane

CL reaches its maximum growth and

function about day 12, secretes the
hormone progesterone, which inhibits
(blocks) LH release by the pituitary gland
via negative feedback mechanism

4/4/2021
Diestrus (cont’d)

 the uterine glands undergo hyperplasia

and hypertrophy
 the cervix becomes constricted
 besides CL, follicles can be palpated
(2 to 3 follicle waves)
 Di-estrus ends with the regression of
CL induced by prostaglandin F2
secreted by the endometrium
(During diestrus, the ovaries are relatively inactive except for the functional
CL. Progesterone and some estrogens enter the circulation to affect mammary
development and uterine growth, endometrium prepares for implantation, CL
continues throughout pregnancy)
If implantation does not occur, the CL degenerates after approximately 16/17
days by the action of PGF2 new cycle initiated
4/4/2021
THE HORMONES OF REPRODUCTION

Primary hormones of reproduction: directly involve in the

regulation of reproductive activities

1. Gonadotropin-releasing hormone (GnRH)

produced by the hypothalamus

transported via the hypothalamic-hypophysal portal vessels in
to the anterior pituitary
controls the release of gonadotropins (FSH, LH) from the
anterior pituitary

4/4/2021
 HORMONES (cont’d)

4/4/2021
HORMONES (cont’d)

GnRH
a ten amino acid peptide
synthesized and secreted from
hypothalamic neurons
binds to receptors on gonadotrophs
regulates LH and FSH secretion

GnRH stimulates pulsatile secretion of

FSH & LH,  in turn stimulate gonadal
secretion of the sex steroids (estrogen,
progesterone)

Sex steroids in turn inhibit (negative

feedback) secretion of GnRH, also have
direct negative effects on gonadotropins
(FSH, LH)

4/4/2021
Female reproductive hormones (cont’d)
Commercially available (common
names of) GnRH products:
•Gonadorelin
•Buserelin
•Cystorelin
•Receptal
•Fertagyl

Pharmacological action:
stimulates the short surge of FSH and
LH after a single injection
Indication:
•follicular cysts
•delayed ovulation
•acyclicity,
•synchronization of estrus &ovulation
4/4/2021
Female reproductive hormones (cont’d)

2. Hypophyseal gonadotropins (FSH and LH)

2.1 Follicle stimulating hormone (FSH)
In Female:
Biological action:
•stimulates the growth and maturation of the
ovarian follicle
•stimulates estrogen production from ovary
or testis in the presence of LH
Indication: Superovulation in donor cows for embryo transfer
In Male: FSH acts on the germinal cells in the somniferous tubules of the testis,
responsible for spermatogenesis up to the secondary spermatocytes androgens
are responsible for final stages of spermatogenesis
4/4/2021
Female reproductive hormones (cont’d)

2.2 Luteinizing hormone (LH)

Tonic or basal levels of LH
In Female:
•induce estrogen secretion (together with FSH) from the
developing ovarian follicle –in the ovary
•responsible for maintenance of CL (di-estrus)
In Male: stimulates interstitial cells (leyding cells, testis)
produce androgens –testosterone
Pre-ovulatory LH-surge/peak controlled by GnRH from the
surge center
• Oocyte maturation
• Follicle rupture, ovulation
• Corpus luteum formation and function
4/4/2021
 Hormone …(cont’d)
•The number of pulses of GnRH and LH varies from a few per
day (diestrus) to one or more per hour (estrus)
•In females, pulse frequency is related to stage of the cycle

4/4/2021
 Female reproductive hormones (cont’d)

Tonic LH and FSH release:

 LH and FSH are released in a tonic or basal fashion in

both sexes

 Controlled by negative feedback from the gonads

(progesterone & estrogen)

…Functions in Male (LH)

 Interstitial cell stimulating factor (ICSF)
 Stimulates the Leydig cells, Testosterone synthesis &
release …Maintains spermatogeneis
4/4/2021
Hormone …(cont’d)

Pre-ovulatory LH and FSH release:

evident in the female before ovulation
initiated by an increase in the circulating
estrogen concentration
•Positive feedback effect of estrogen
on the hypothalamus pre-ovulatory
surge of FSH and LH  Ovulation
Ovulation: 24-30 hours after the initial
maximal LH-surge (cattle), last from 6 to 12 hours

(Human chronic gonadotropin (hCG) is used in place of pituitary LH to

induce ovulation, primarily in cows with cystic ovaries)

4/4/2021
 Hormones of reproduction
Hormone Origin Target Functions

Gonadotropin Releasing Hypothalamus Anterior pituitary Release of FH & FSH

Hormone (GnRH)
Luteinizing Hormone Anterior pituitary Ovary Ovulation,
(LH) Testis CL formation,
Testosterone
Follicle stimulating Anterior pituitary Ovary Development of follicle,
hormone-FSH gland Testis Sperm production

Estrogen (several types) Follicle Uterus, Mating behavior,

Placenta Hypothalamus Secondary Sex
Characteristic
Progesterone Corpus luteum Uterus, Maintains pregnancy,
(CL) Hypothalamus Inhibits GnRH release

Prostaglandin Uterus Corpus luteum Regression of CL

F2 (endometrium)
Testosterone Testis Skeletal muscle, Sperm production, male
testis, libido,
Secondary sex character

4/4/2021
3. Extra hypophyeal gonadotropins – placental
hormones

3.1 PMSG or equine chorionic gonadotropin (eCG):

a glycoprotein hormone produced by the endometrial cups of

the uterus of the mare from about 50 to 120 days of pregnancy

eCG stimulates development of ovarian follicles

•Some of the follicles ovulate
•most form a luteinized follicle, due to the LH-like action of
the eCG; forming accessory corpora lutea, produce
progesterone, necessary for the maintenance of pregnancy
in the mare

4/4/2021
 Female reproductive hormones (cont’d)

PMSG/eCG has both FSH and LH biological actions,

with the FSH actions being dominant
isolated from the blood of pregnant mares (not found
in urine)
•one of the first commercially available
gonadotropins
•used to induce superovulation in Embryo
Transfer (ET)
Indication: superovulation of donor cows for ET
4/4/2021
Female reproductive hormones (cont’d)

3.2 Human chorionic Gonadotropin (hCG):

 a glycoprotien hormone
 synthesized by the syncytiotrophoblastic cells of the
placenta of primates

 found both in the blood and urine

 extracted from urine of pregnant women

•primarily an LH-like effect, used as substitute
for the more expensive LH
…hCG has a longer half-life than LH
4/4/2021
Female reproductive hormones (cont’d)
LH-like action: for treatment of cystic ovaries in cattle, (the follicle either
ovulates and forms a CL or more often luteinize)

Commercially available products:

•“chorionic gonadotropin”
•“chorulon”, “cystovet”

Pharmacological action of hCG:

•stimulates follicular maturation and ovulation
•CL formation and maintenance
•stimulates androgen production by the theca cells
of the ovary and leydig cells of the testis

Indication:
•delayed ovulation or anovulation
•ovarian cysts (follicular type)
• to improve chances of pregnancy in cyclic non-pregnant cows (repeat-
breeder cows)
•to improve libido in bull
4/4/2021
Female reproductive hormones (cont’d)

4. Prolactin (Luteotrophic hormone, LTH), Polypeptide

hormone
produced in anterior pituitary (adenohypophysis)
 gonadotrophic hormone/ luteotropic properties,
maintenance of CL in rodents (mouse, rat), stimulates
the secretion of progesterone

secretion of prolactin near the end of pregnancy and lactation

•Mammogenesis:the development of mammary gland during pregnancy
•Lactogenesis:milk production together with progesterone and
estrogen, necessary for the maintenance of lactation (secretion)

Negative feedback on gonadotropins (lactation anestrous, particularly in swine)

CNS: Prolactin induces maternal behavior (nest building, nursing behavior)

4/4/2021
Hormone …(cont’d)
5. Oxytocin (Neurohypophyseal hormone), a peptide hormone

Synthesized together with carrier protein (neurophysins) in the

hypothalamus

Transported to and stored in

Neurohypophysis (posterior pituitary)
•stored at the nerve endings next
to the capillary beds until its
release into the circulation
also produced in the Corpus luleum,
has two sites of origin: the ovary and
the hypothalamus.
4/4/2021
Hormone …(cont’d)
Functions (oxytocin):
Contraction of smooth muscle of the
female reproductive tract …transport of
both female and male gametes in the oviduct

Milk let-down:
Suckling or milking induces the release of
oxytocin into the circulation, to act on the
myoepithelial cells (smooth muscle cells)
that surround the alveoli in the mammary
gland resulting in milk-letdown

 Myometral contractions during

parturition and postpartum
Estrogen enhances responsiveness
of smooth muscle to oxytocin (Ovarian oxytocin is involved in luteal function by
acting on endometrium to induce prostaglandin F2 release  luteolytic action
(regression of the corpus luteum)
4/4/2021
Female reproductive hormones (cont’d)
Indications (oxytocin)
Cattle
 milk-letdown , expulsion of retained placenta
 hastens uterine involution following dystocia, caeserean operation, after
replacement of uterine prolapse
Horse
 induces foaling
 expulsion of retained placenta
 induces milk-letdown
Release of oxytocin is dependent on a signal
from the nervous system:

 Reflexly induced by the fetus in the birth canal

 Physiological stimuli or teat stimulation for milk letdown
 Genital tract stimulation (copulation) causes oxytocin release to aid in
propulsion of semen through the genital tract
4/4/2021
Hormone …(cont’d)

6. Gonadal steroid hormones

Synthesized from cholesterol (C-27)
Secreted by the ovary, testis, placenta and
adrenal cortex

Basic or common nucleus

“cylopentanoperhydrophenanthrene
nucleus”

Products of cleavage are sex steroids with

 a 21 carbon steroid- gestagen activity
 a 19 carbon steroid- androgen activity
 a18 carbon steroid- estrogen activity

•Testis synthesizes primarily androgens

•Ovaries: estrogen and progesterone

4/4/2021
Hormone …(cont’d)

6.1 Estrogens (E2)

Biologically active estrogen are:
Estradiol, estrone and estriol
Estrogens secreted by the ovary, placenta
and adrenal gland, have androgenic precursors
•Estrogens carried by binding proteins
in the circulation (like androgens)

Function:
CNS: induce behavioral estrus in the female

Uterus:
•the mass of both the endometrium and myometrium due to cell
hyperplasia and hypertrophy (proliferative phase),
• contraction of genital tract by potentiating the effect of oxytocin and
PGF2.
4/4/2021
Hormone …(cont’d)

Estrogens:
Attributes to physical development of female secondary sex characteristics
Stimulates mammary gland development
…  duct growth
Have both a negative (on the tonic center) and a positive feedback (pre-
ovulatory/ surge center) control through Hypothalamus on LH and FSH release

Estrogens used to abort cow and sheep …  a luteolytic action (regression

of CL), caused by synergistic effect with PGF2

In the sow estrogens have a luteotrophic action, thus used to synchronize
estrus to hold sows in luteal phase until treatment withdrawal followed by a
PGF2 injection causing regression of the CL

4/4/2021
Hormone …(cont’d)

6.2 Progesterone (P4)

secreted by luteal cell of the CL,
by the placenta and adrenal gland

Secretion is stimulated by LH
P4 transported in blood by a binding
globlin
Function:
 prepares the endometrium for implantation and maintenance of pregnancy by
• increasing secretory glands (secretion phase) in the endometrium
• inhibiting the motility of the myometrium – “the hormone of pregnancy”
• develops the secretory tissue of the mammary glands
4/4/2021
 Female reproductive hormones (cont’d)
  P4 inhibit estrus and the preovulatory surge of LH, ( negative
feedback) …hormonal regulation of the estrous cycle

 Indications (P4):
 Progestogens are given to prevent abortion
in females prone to abortion as a result of
insufficient secretion of endogenous P4
 Used in birth control pills for women to
prevent LH surge and subsequent ovulation
 Synchronization of estrous cycle of cows and pig, inhibits LH surge
preventing estrus and ovulation but allows development of ovarian
follicle
4/4/2021
Hormone …(cont’d)
Function (P4)
Synchronization of estrus in cows and heifers:
A device inserted into the vagina and left in situ, subcutaneous implants or
feeding for 7- 12 days, with PGF2 administration 24 hours before removal,
estrus occurs in 2-5 days after withdrawal
Controlling the estrous cycle by extending the luteal
phase of the cycle with subsequent regression of the
CL during the time of progestin administration
Synthetic progesterone - estrus synchronization
 Progesterone releasing intra-vaginal device (PRID)
 Controlled Internal Drug Release (CIDR)
 Melengesteral acetete (MGA) - feeding
 Fluorogestone acetate intra-vginal sponges & Medroxyprogesterone acetate
intra-vaginal sponges (used to synchronize ewes and female goats i.e. single sponge is inserted
into anterior vagina, remain 12-14 days (estrus occurs 48-72 hours after withdrawal)
 Progestin implants are also available
 Medroxyprogesterone acetate tablets: to postpone estrus in bitch
 Proligestone injection interrupt & postpone estrus (bitch)
4/4/2021
 Hormone …(cont’d)

Diagram of the PRID

Picture of CIDR with its applicator and during inserting in to the

vagina of a cow (Perry et al., 2004)

4/4/2021
Hormone …(cont’d)
6.3 Androgens/Testosterone
Male: Interstitial cells (leydig cells) of the testes,
limited amount by adrenal cortex
Testestrone ---> Dihydrotestosterone (biologically active),
binds to the nuclear receptor
 stimulate late stage of spermatogenesis and prolong the life span of epididymal
sperm
 promote growth, development and secretory activity of the accessory sex
organs of the male: prostate, vesicular glands, bulbouretheral gland, vas
deferens and the external genitalia (penis and scrotum)
 maintenance of secondary sex characteristics and sexual behavior or libido
 Naturally occurring or synthetic analogues, have limited application in animal
reproduction  Testosterone propionate is administered to cows or steers
used as teaser animals for detection of estrus
4/4/2021
Hormone …(cont’d)

7. Relaxin, peptide hormone

Relaxin is produced/secreted primarily by the corpuls luteum, the

placenta (sheep, horse)
Produced and released during the late pregnancy (shortly before
parturition)
Biological action:
 Dilatation of the cervix and vagina before parturition
•Relaxation of the ligaments (lig. Sacrotuberale) and joints
(sacroiliac joint, pelvic symphysis)
4/4/2021
Hormone …(cont’d)

8. The prostaglandins
derived from arachidonic acid,
first isolated from accessory
sex gland fluids, the prostate gland
secreted by almost all body tissues

PGF2 is the natural luteolytic agent, terminates the luteal phase …PGF2
controls the life span of the CL, that regulates the length of the cycle

The CL of pregnancy is resistant to the luteolytic effect of PGF2, because of

the production of Interferon tau(τ) which reduces the receptor for oxytocin (no
PGF2 synthesis by the endometerium)

PGF2 is particularly potent in terminating early pregnancy

4/4/2021
4/4/2021
4/4/2021
Hormone …(cont’d)

Function (PGF2)
Contraction of smooth muscles of reproductive tract in conjunction
with oxytocin and estrogen (estrogen and oxytocin stimulates
PGF2 synthesis and release)

An increase in estrogen, stimulates

PGF2 synthesis and release by the
endometrium

 erection, ejaculation, sperm transport

 ovulation
 parturition and milk-ejection

4/4/2021
Hormone …(cont’d)

During the first 5 days(ewe, cow) of luteal

development (in sow until day 12 of the
estrous cycle) and during natural CL
regression (after day 16/17 of the estrous
cycle) the CL is not responsive to PG,
“refractory”

PGF2 passes directly through the walls

of the utero-ovarian vein into the ovarian
artery
… to the corpus luteum (ovarian artery
is tightly adherent to the uterine vein,
“countercurrent mechanism”)

4/4/2021
Hormone …(cont’d)

 Trade names (PGF2):

Estrumate
Lutalyse
Prostamate
Indications:
CL persistent (pyometra)
Treatment of endometritis-initiation of new estrous cycle estrogen 
defense mechanism
Treatment of ovarian cyst (luteal cyst)
Treatment of postpartum disorder-RFM, Mucometra…)
Induction of parturition

Synchronization of estrus
• Single injection - Luteal (CL) phase
• Double injection - (unknown reproductive phase)
4/4/2021
Hormone …(cont’d)
2. Secondary hormones of reproduction
Hormones primarily involved in various functions (activities) of the
body and indirectly influence reproduction
A) Adrenal corticoids (corticosteroids, regulated by ACTH)
The common corticosteroids are
•Dexamethasone
•Flumethasone
•Betametasone
•Cortisone
Indication: Induction of parturition in ruminants- to reduce
dystocia cases and thus mortality
B) The thyroid hormones derived from the amino acid tyrosine contain
iodine and produced by the thyroid cells of the thyroid gland
 Combined with plasma proteins for transport in the blood
4/4/2021
Hormone …(cont’d)

 Regulation and discharge of thyroxin is controlled by thyrotrophic

hormone (TSH)
 Secreted by the adenohypophysis in response to the hypothalamic
releasing hormone (T-RH) - Influenced in a feedback
mechanism by the thyroid hormones (Thyroxin (T4) and tri-
iodothyronine (T3)

 After absorption thyroxin causes an increase in metabolic rate

with:
• utilization of carbohydrate (energy)
• combustion of fat with metabolization of fat from all depots
Deficiency:
•Clinical manifestation takes the form of goiter and are most apparent
in young animals (be born dead, hairless calves and piglets)
4/4/2021
 Physiology of fertilization/gestation
Outline
 Ovulation and fertilization
 Ovulation and ovum transport
 Spermatozoa transport, capacitation
 Entry of the sperm into the ovum, pronucleus formation
 Syngamy, Embryo cleavage, - transport and uterine accommodation

 Physiology of gestation

 Maternal recognition of pregnancy

 Implantaion
 Placentation
 Methods of pregnancy diagnosis

4/4/2021
Ovulation and fertilization
Introduction
 Luteal (CL) phase:
• progesterone concentration, negative
feedback on pituitary and hypothalamus,
↓ GnRH ↓FSH, LH, no estrus & Ovulation
• 2 or 3 waves of follicle development,
induced by FSH, but no ovulation (undergo atresia, atretic)

 Follicular phase (up on regression of the CL):

•↓P4 level in the blood
• development of dominant follicle, estrogen
synthesis by the developing follicles
• Positive feedback of estrogen,  secretion of GnRH  release of FSH,
LH
• Pre-ovulatory LH surge  Ovulation… CL formation
4/4/2021
 Ovulation and ovum transport:
Ovulation: a process in which the matured graafian follicle ruptures →release of
the egg (ovum)

 bulging or protrusion of follicle from the

surface of the ovary
 forming of cone like projection … → rupture!

→ A thin follicular fluid, the egg and the cell mass

pass out detaching from the degenerating granulosa cells

 Cow and ewe: follicle ruptures at any point on the ovaries except at the hilus

 Mare: follicles develops in the centrally

located cortex, migrates into the area of
ovulation fossa …→ ovulation!
…b/c the tough tunica albuginea prevents ovulation
through the surface of the ovary

4/4/2021
  Ovulation …(cont’d):

Species Type of ovulation

Cow spontaneous
Sow spontaneous
Ewe spontaneous
Mare spontaneous
Cat induced ovulation
Bitch spontaneous
Camel induced ovulation

4/4/2021
 Ovulation … (cont’d)

 Pre-ovulatory changes of the Graafian follicles:

 Cytoplasmic and nuclear maturation of the oocyte
 after few ours of LH surge meiosis I (reduction division) resumes and ends
resulting in:
 secondary oocyte: chromosome numbers change
from the diploid (2n) to the haploid (n) state
 the first polar body: trapped between the vitelline
membrane and the zona pellcida in the perivitelline space

Disruption of cumulus cell cohesiveness among the

cells of the granulosa layer
- Dissociation of cumulus cells from the granulosa layer  frees the oocyte
 Thinning and rupture of the external follicular wall  ovum release

4/4/2021
  OVUM TRANSPORT
 Fimbria of the oviduct “captures” the released egg

 Infendibulum acts as a funnel to receive the ovum:

ovum and surrounding cells free in the infundibular
portion of the oviduct

 …Oviduct rhythmically propel (drive) the egg

down the oviduct (ampulla)

Folliclar fluid serves as a medium for the

free-floating ovum
 Further ovum transport depends on:
 the normal function of the cilia lining the oviduct
 oviductal smooth muscle activity
 functions of the ampullary-isthmic junction and uterotubal junction

4/4/2021
 OVUM TRANSPORT (cont’d)
 … Ovum transport is controlled mainly by the ovarian steroid
hormones’ balance after ovulation

- Estrogen (“tube locking”)

- Progesterone (“tube unlocking”)
and
- Prostaglandin (locally produced PG of the F series impedes and of
the E series accelerates)
The ovum(a) arrested at the ampullary-isthmic junction
for some times: ready for fertilization!
 At ovulation the egg is surrounded by layers of granulosa cells:
 the comulus oophorus
 the corona radiata
 zona pellucida
 vitelline membrane

4/4/2021
 Ejaculation, spermatozoa transport in the female genital
tract
 Ruminants:
 small volume of semen is ejaculated into the cranial end of the vagina and
into the cervix (Bull: 7 ml, Ram: 1.5 ml )
 Horse and swine:
 voluminous ejaculate is deposited into the uterus (Stallion: 80 ml, Boar: 200
ml)
 Spermatozoa transported through various luminal fluids of different physiologic
and biochemical characteristics:
 Testicular fluid, Epididymal fluid, Seminal plasma
 Vaginal fluid
 Cervical mucus
 Uterine fluid
 Oviductal fluid

 Three stages are recognized in sperm transport in the female reproductive tract:
 Short, rapid sperm transport
 Colonization of reservoirs and
 Slow, prolonged release
 Rapid Transport:
 Spermatozoa penetrate the cervical mucus, quickly transported through the
cervical canal and some sperms reach the site of fertilization

 Colonization of sperm reservoirs:

 Massive numbers of spermatozoa are trapped in the mucosal folds of the
cervical crypts (fewer leucocytes, less phagocytosis of spermatozoa in the
cervix than in vagina and uterus)

 Spermatozoa leave the cervix:

• actively by means of their own motility or
• passively by cervical and uterine contraction

 Slow release and transport:

 After establishment of sperm reservoirs, the spermatozoa are released slowly
for a prolonged period, ensuring continued availability of spermatozoa for
entry to the oviduct to effect fertilization of the egg
 Sperm Capacitation
Capacitation: gaining of the ability of sperm cell to be able to fertilize the ovum,
as it traverse the genital tract secretions
Changes in the sperm making capable of fertilization, takes some 4 to 6
hours (in the female genital tract)
Modifying or removal of sperm surface
components by genital tract secretions
 destabilizing of phospholipids layer
 acrosomal activation → subacrosomal
enzymes release
• Acrosin
• Hyaluronidase
• Neuroaminidase
Subacrosomal enzymes digest bonds between
the cells of surrounding layers of the egg
Sperm capacitation starts in the uterus; major site is the oviduct,
especially the isthmic region

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 Entry of sperm into ovum
The process of stripping away of the surrounding layers of cells is called
“denudation of the ovum”:

1. Penetration of the cumulus mass: hyaluronidase enzyme & acrosin dissolves

the hyaluronic acid matrix

2. Penetration of the zona pellucida: facilitated by Neuroaminidase (proteolytic

enzymes) of the inner acrosome membrane

 “Zona reaction”: chemical reactions between the products of the sperm

and the zona pellucida ( Sealing: preventing penetration by other
sperms)
3. Acrosin (protease) stripes the vetelline membrane
After the sperm head denuded the acrosome, the exposed perforatorium
penetrates the vitelline membrane (by attaching of the sperm head to the
surface of the vitellus)
 “Vitelline block”: sealing of the vitelline membrane to prevent other
sperm from penetrating
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FERTILIZATION
Fertilization: the fusion of two haploid cells
to form a single cell, the zygote
It involves:
Activation of the ovum by the sperm: with fertilization,
the second meiotic division completes, products are the zygote
(fertilized egg) and the second polar body

…Introduction of hereditary material from the male

into the ovum

The site of fertilization is the lower portion of the

ampulla in all farm animals

Egg enters the ampulla with its mucoprotein coat

(zona pellucida) surrounded by a cluster of granulosa
cells called cumulus cells

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Pronucleus formation
After penetration of the sperm into the ovum, the perforatorium
and the tail drop off leaving the sperm nucleus, followed by:

•…Formation of the male pronucleus: a number of nucleoli

appear within the sperm nucleus, then a nuclear membrane
develops

•…the female pronucleus is formed soon after sperm entry

(the female pronucleus resembles the male pronucleus in the appearance and
the formation of a nuclear membrane)

→ → the two pronuclei develop synchronously

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SYNGAMY
 At stage of maximum development, the pronuclei migrate
toward one another and fuse (come into contact)

 the nucleoli and nuclear membrane disappear, the two

pronuclei can no longer be seen

 the maternal and paternal chromosomes unite to form a

single group representing the prophase of the first cleavage
mitosis - fertilization complete !

  The new nucleus and its surrounding cytoplasm form a

zygote, ready for mitotic cell division to form a new life

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 CLEAVAGE
The zygote, or one-cell stage, is quite large having a low
nuclear/cytoplasmic ratio

Embryo undergo rapid cell cycles, mitotic divisions

 Cell divisions are without an increase in cell mass, producing

a cluster of cells the same size as the original zygote - the
process is referred to as cleavage

 Mammalian embryos contain little egg yolk

 They relay on oviductal and uterine secretions (histotroph) during early

pregnancy
 The pole of the egg with the highest concentration of yolk is the vegetal
pole while the opposite is the animal pole

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 CLEAVAGE (cont’d)

 Cleavage of the zygote is by vertical division from animal to

vegetal pole

 The resulting daughter cells up to the blastula stage are called

blastomeres

 Upon Cleavage/ mitotic divisions: each daughter cell

(blastomere) receives the full assortment of chromosomes
 Blastomeres are encased within the zona pellucida, they must
accommodate themselves to this limited area

Embryo with 8 to 16 blastomeres (in some cases more) is referred

to as a morula, mulberry like
… Cell division & cell orientation: morula becomes hallow →
blastocyst

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 Early development of the fertilized ovum to the blastocyst stage

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Embryo transport and uterine accommodation
Embryo at 8 to 16-cell stage transported into the uterus, where it
continues to develop or proliferate

Movement through the oviduct to the uterus controlled by:

 estrogen (“tube locking”)
 progesterone (“tube unlocking” and
 prostaglandin (locally produced PG of the F series impedes
and of the E series accelerates)
 activity of the tubal muscle & cilia
 the muscular constriction at the ampullary- isthmus or at the
uterotubal junction

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Embryo transport …cont’d

Up to the time of shading of the zona pellucida at the ninth day
there is little absolute growth of the mammalian zygote from its
original dimension of 0.14 mm

Follwing shading of zona pellucida:

Ruminants (cow, sheep): Blastocyst elongates after the 9th day of
gestation
•amniotic sac filled by amniotic fluid
•allantochorionic sac filled by allantoic fluid,
•gravid horn distends
•chorion extends into the non-gravid horn
4/4/2021
 Bovine conceptus at 60 days: fetus contained within the amniotic sac, an elongated
allantochorion and evidence of the formation of cotyledons

4/4/2021
 Embryo transport …cont’d
Equine:
•Migration of the conceptus, the early embryo does not
elongate
•E.g. at 35 days the blastocyst is oval rather than
cylindrical, more distended by the allantoic fluid

Early embryonic attachment to the uterus occurs at

12 days of fertilization in cow
14 “ in sow
15 “ in ewe
25-30 days in mare
13-17 days in dog & cat

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Physiology of gestation
 Maternal recognition of pregnancy:

 “Maternal recognition of pregnancy”: the period when the

conceptus signal its presence so that pregnancy can be
established
 mediated through factors of embryonic origin, directly or
indirectly prevents regression of the corpus luteum
 associated with the maternal endocrine response

 … Establishment and maintenance of pregnancy requires:

 prolonged luteal phase: presence of a single corpus luteum or
corpora lutea
 …elevated progesterone concentrations

P4 exerts a negative feedback on the anterior pituitary and

hence inhibition of follicular maturation and ovulation

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 Maternal recognition of pregnancy (cont’d)

 The developing embryo (conceptus) exerts its effect through either:

 an antiluteolytic action: preventing the synthesis, release or action of

prostaglandin F2α and or
 a luteotrophic effect, overrides the effect of luteolycin (PG F2α )

 The conceptus produce (ruminants):

- trophoblastic proteins, glycoprotein (intact embryo synthesizes
interferon like substance (ovine/bovine trophoblast interferon,
oIFN/bIFN-tau, ) to signal its presence to the maternal system)

 oIFN-  & bIFN-  inhibit endometrial oxytocin receptor synthesis, ↓PGF2α

synthesis
…  CL maintained , production of P4 comparable to di-estrus period
… Continuing of endometerial development and secretory activity

4/4/2021
 Maternal recognition of pregnancy (cont’d)

 In the cow: bIFN secreted by the conceptus between days 16 to

17 of gestation inhibit PGF2 production by uterine
endometrium

 In the ewe: MRP occurs between days 12 to 21 of gestation

through oIFN
 In pig:
Estrogens produced by the conceptuses provide the signal for
MRP between day 11 and 12 of gestation
 Presence of conceptuses in both uterine horns, at least two
piglets per uterine horn
 … PGF2 produced by the endometrium re-routed into the
uterine lumen (estrogen action) and metabolized, not drained
by the uterine veins

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 Maternal recognition of pregnancy (cont’d)
 In mare:
 the presence /migration of the conceptus prevents
luteolysis by inhibiting the production of PGF2 by uterine
endometrium

 the conceptus must migrate within uterus b/n 12 to 14 times

per day during days 12, 13 and 14 of pregnancy to inhibit
PGF2 production (the early embryo does not elongate)

 Estrogens and /or conceptus secretory proteins may also

provide MRP (not clearly determined!)

4/4/2021
The time of maternal recognition of pregnancy
Species Days of maternal Substance responsible Days of definite
recognition of for maternal attachment
pregnancy recognition of
pregnancy

16 –17 (bTP-I (bovine Trophoblast protein 18 - 22

Cow Trophoblast protein, synthesized by the
bIFN- blastocyst
12 – 13 Trophoblast protein 16
Ewe
(oTP-I, oIFN-)
synthesized by the
blastocyst

13 – 14 Conseptus migration, 36 – 38
Mare Proteins/estrogens???
synthesized by the
blastocyst
12 Estrogens synthesized 18
Sow by the blastocyst

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Pregnancy …(cont’d)
IMPLANTATION
Implantation: Embedding of the embryo around
the wall of the uterus
•Superficial and noninvasive- in domestic animals
•Invasive: in rodents and primates implantation
Phases involve:
•trophoblast–uterine cell contact
•apposition
•adhesion
Attachment: through interdigitation of the
uterine crypts and trophoblastic microvilli
Ruminant: uterine caruncles interdigitate
with the microvilli from the cotyledon
forming placentomes
• Cow: 80 –120 , Sheep: 80 Bovine conceptus at 115 days- blood supply
• Goat: up to 160 cotyledons arranged in to and from the developing cotyledons
four rows along each of the uterine horns
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Pregnancy …(cont’d)

PLACENTATION
Placentation: development of extra embryonic fetal membranes
(amnion, allantois and chorion
Farm animals: chorioallantoic - fusion of the outer layer of the allantois
to the chorion (fetal vessels in the allantois come into close apposition to the
umbilical arteries and veins located in the connective tissue between the allantois
and chorion)

Placenta classified according to:

 Morphology- distribution pattern of the microvili

 Microscopic characteristics of the maternal-fetal barrier
 Loss of maternal tissue at birth
4/4/2021
 Classification of chorioallantoic placentas
Species Chorionic microvili Maternal –Fetal Loss of maternal
distribution/pattern Barrier tissue at birth
Ruminants Cotyledonary Epitheliochorial Nondeciduate
(semiplaccenta)
Mare Semiplacenta diffusa Epitheliochorial Nondeciduate
completa (semiplacenta)

Pig Semiplacenta diffusa Epitheliochlorial Nondeciduate

incompleta (Semiplacenta)

Dog, cat Zonary Endotheliochorial Deciduate (placenta

vera)

Primate Disicoid Hemochorial Deciduate (placenta

(human, vera)
monkey)
4/4/2021
Pregnancy …(cont’d)

Functions of placentas:

•Synthesis of hormones (Progesterone & estrogen)

•Respiratory organ (gas exchange)

•Organ of nutrition (permits the transport of sugars, amino

acids, vitamins and minerals to the fetus as substrates for fetal
growth based on net flux)
•Organ of protection (placental barrier)

4/4/2021
Pregnancy …(cont’d)

Fetal fluids and membranes during pregnancy

 fetal fluid consist of amniotic and allantoic fluids

 total quantity of fetal fluid increases progressively:
•5 liters at 5 months
•20 liters at term

Origins of fetal fluids are:

•the respiratory, urinary, digestive system & the fetal skin
• Other sources are: secretion of fetal salvery glands, buccal
mucosa
4/4/2021
Pregnancy …(cont’d)

Volume of fetal fluid:

Fluctuation in relative volume of amniotic and allantoic fluids: allantoic fluid is

relatively higher than amniotic fluid during pregnancy

Sharp rises in the total fluid:

•first 3rd of pregnancy, 40 and 60 days of gestation (more allantoic fluid )
•second third pregnancy, 3 and 6 months (more amniotic fluid)
•final third of pregnancy, 6½ and 7½ months (allantoitic fluid is in excess)

•At birth the allantoic sac forms the first and the amnion the second “water
bag”
•The allontochorion is thicker and tougher than the transparent amnion sac

4/4/2021
Pregnancy …(cont’d)
Fetal growth: Increase in size and weight, many changes in form
Cattle: Over one half of the increase in fetal weight last 2 months of gestation

Factors affecting fetal growth

Genetic factors: at birth Holstein fetuses weigh about 35% more than Jersey
calves
Environmental factors: parity, nutrition of the mother, litter size (in pig),
climatic factors (stress); high ambient temperature during pregnancy
(Twin fetuses in monotocos species (cattle) are smaller than single fetuses,
length of gestation reduced)

Fetal hormones: Fetal hormones influence fetal growth (growth hormone

stimulate fetal growth)
Insulin-like growth factors (IGF-I and–II) or sommatomedins are
polypeptide hormones similar to insulin, occur in fetal and placental tissues
4/4/2021
Pregnancy …(cont’d)

Age determination of bovine fetuses

Long bone length (radius and tibia) is a reliable indicator of fetal

age from 50 days of gestation to term, obtained ultrasonically
Length of long bones at various stages of conception

Days after conception Length of radius Length of tibia

(mm) (mm)
50 4.8 50
60 10 12
100 36 47
120 56 76
150 79 107

4/4/2021
Pregnancy …(cont’d)

Formula for calculating the age of the fetus from its crown – anus length in
centimeters (Richardson, 1980)
Pig X = 3 (Y + 21)
Calf X = 2.5 (Y + 21)
Lamb X = 2.1 (Y + 17)
Where X = developmental age in days
Y = Crown-anus length in cm More accurate determination of the
age of the embryo/fetus by means of
Fetal body length in cows at various
ultrasound
stages of pregnancy (Crown-anus)

•the heartbeat (day 22)

Pregnancy Fetal body length (cm)
•the spinal cord (day 28)
(month)
1 0.5 •the placentomes (day 35)
2 6 •split hooves (day 44)
5 40 • the ribs (day 52)
7 70
9 90
4/4/2021
PREGNANCY DIAGNOSIS (Cow)
A tool to measure the success of a reproductive management:
•for early detection of problems
•… for re-synchronization of non-pregnant cows
Methods:
1. Non-return to estrus, failure to return to estrus
Non pregnant cow:
• the CL regresses (days 16/17 of the estrous cycle)
• ↓plasma P4 concentrations, follicle development
• signs of estrus 20-23 days after AI non-pregnant
Pregnant cow:
• no regression of the CL
• plasma P4 concentrations, a possibility of pregnancy
…  non- return to estrus
4/4/2021
Pregnancy diagnosis…(cont’d)
2. Milk or plasma P4 concentration at about 21 days
after AI
•CL regression (days16/17 of the estrous cycle)
•↓ milk or plasma progesterone concentrations
detection of non-pregnant cows as early as 21 days
after AI

•at least two times samples collection at 21 and 24 days of AI: not
pregnant, if one of the samples has low progesterone concentrations

•… around 80% reliability of progesterone tests to detect pregnant

cows, even if both samples have high progesterone concentrations

…→a good diagnosis tool to detect non-pregnant cows!

4/4/2021
Pregnancy diagnosis…(cont’d)
Reasons 20% failure:
a) Cows do not show estrus exactly every 21 days:
 a non-pregnant cow may show estrus, ovulate and form a CL less than
21 days after AI
 P4 concentrations
…  non-pregnant cow may be erroneously diagnosed as pregnant!
b) Cows with reproductive problems:
 ovarian cysts or uterine infections -  two consecutive high P4
concentrations
  non-pregnant cow may be diagnosed as pregnant!
c) Pregnant cows at 21 days after AI may lose that pregnancy in the next 30
to 40 days (pregnancy losses in that period may reach up to 30%)
d) Cows inseminated when not really in estrus will often be in the luteal
phase at 21 days after AI:
•P4 concentrations, even if not pregnant!
…  non-pregnant cows diagnosed as pregnant!
Pregnancy diagnosis…(cont’d)
3. Ultrsonography
Doppler Effect
Ultrasonic fetal pulse detector based on Doppler Effect:
• the fetal heart, from 6 – 7 weeks
• blood flowing in arteries
audible sound produced

B-or brightness mode (real time

utrasonography)
•with 5.0 MHz transducer as early as 12 to 14 days
•with 7.5 MHz transducer as early as 9 days
Visualization of a fluid filled cavity
containing a viable fetus within one of
the uterine horns
4/4/2021
Pregnancy diagnosis…(cont’d)

4. Rectal Palpation
Changes in the size of uterus and uterine horns offers strong
evidence of pregnancy!
•PD as early as 35 days after AI
•… Palpation of the uterine contents and CL rectally, common !
•CL of pregnancy persists at its maximum size throughout the
period of gestation, though it is impossible to differentiate b/n CL
of pregnancy and that of diestrus

•False pregnancy diagnosis!: pyometera, mucometra,

incomplete uterine involution

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4.1 Asymmetry of the uterine horns
 Difference in size of the
pregnant uterine horn,
accumulation of fetal fluids,
particularly the allantoic fluid,
fluctuating feel with good tone

…a definite diagnosis can

be made based on these signs
Asymmetry of the horns and CL on ovary
alone ipsilateral to the pregnancy

4/4/2021
Pregnancy diagnosis…(cont’d)
4.2 Palpation of the allantochorion (membrane slip)

 Feeling of the slipping of the allantochorion membrane (fetal

membrane) along the greater curvature
within the uterus

Steps:
•identify the bifurcation of the uterine horns
•pick up the enlarged, gravid horn between
thumb and finger just cranial to the bifurcation
•gently squeeze the whole thickness of the horn
(…The allantochorion slips b/n the thumb and finger before
the uterine and rectal walls are lost from the grasp)

4/4/2021
Pregnancy diagnosis…(cont’d)

4.3 Palpation of placentomes

 at about 90-100 days, 8-10 cm in front of
and over the pelvic brim
impossible to palpate b/n five and seven
months: the uterus has sunk into the abdomen

4.4 Palpation of the fetus

 between 120-160 days of gestation, the extremity will lie within reach in
front of and below the pelvic brim
 the fetus is less detected b/n 5½ and 7½ months
 7½ months to the end of gestation in the majority cases (head and/or flexed
limbs readily palpated)

4/4/2021
Pregnancy diagnosis…(cont’d)

Other methods
Abdominal ballottement
fairly vigorous pummeling of the ventral abdomen and
flank with clenched fists
… as early as seven months of gestation in some small
breeds (Jersey)
Vaginal examination - manual or visual (using illuminated
speculum)
dryness and blanching (pale) of the mucus membrane as in case of
diestrus
Plug sealing of the cervical canal (gelatinous and tough mucus), adhesive
tenacious secretion on gently pressing of the cervical os

•…toward the end of pregnancy: edematous vaginal mucosa,

edematous and congested vulva
4/4/2021
Cervix:
Tension on the cervix, pulled forwards and downwards over the
pelvic brim in advanced pregnancy (non-pregnant or early
pregnant cervix is freely moveable from side to side)

Mammary glands (best observed in primigravida):

 Mammary glands and the teats of the gravid heifer commence to
enlarge about the fourth month
 enlarged and oedematous gland as parturition approaches
 umbilicus region may become swollen by oedema
 a honey-like secretion from the teats of pregnant heifers after the fourth
month
 External indications:
 increasing size of the abdomen
 enlargement of the udder
 behavioral changes (sexual quiescent)

4/4/2021
Uterine position and diameter, as well as structures felt at palpation according to
stage of pregnancy

Stage of Uterine Uterine Palpable Structures

pregnancy (days position diameter
of gestation)
35-40 Pelvic floor Slightly Uterine asymmetry
enlarged
45-50 Pelvic floor 5.0 - 6.5 cm Uterine asymmetry

60 Pelvis/Abdo 6.5 – 7.0 cm Membrane slip

men
90 Abdomen 8.0 - 10 cm Small placentomes/fetus (10-15
cm long)
120 - 150 Abdomen 12 cm - 18 cm Placentomes/fetus (25-30 cm
long)/fremitus
Placentomes/fetus (35-40 cm
long)/fremitus

4/4/2021
References
 Reproduction in farm animals (1993). 6th edition. E.S.E. Hafez. Lea &
Febiger, Philadelphia.

 Veterinary reproduction and obstetrics (1989). 6th edition. Geoffrey H.

Arthur; David
Veterinary reproduction and obstetrics (2001). 6th edition. Geoffrey H.
Arthur; David E. Noakes; Harold Pearson. Bailliere Tindall.
London -Philadelphia - Toronto – Sydney – Toky
animal reproduction (2000). 5th edition. H. Joe Bearden; John W.
Fuquay. Prentice-Hall, Inc. Upper Saddle River. USA.

4/4/2021
 Thank
you

4/4/2021