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Chromosomes and Gametes

Aims:
Gamete formation and migration
The advantages of sexual reproduction
Meiosis and mitosis in gametes
Structure of gametes related to function
Gamete signalling
Research into oocyte and sperm structure

Reproduction
Reproduction in mammals involves sex, which results in fertilisation. During fertilisation, the
opposite gametes known as Spermatozoa and oocyte fuse and form a zygote. During sexual
reproduction, a new individual is formed, which receives its chromosomes in two portions- half
carried in the spermatozoa and the other half in the oocyte.
How are Gametes made?
Sexually reproducing species put aside germ cells, which travel to the specific gonads and
differentiate into the specific gametes (spermatozoa or oogonia).In these gonads, these gametes
undergo a reduction division known as meiosis, which allows the reduction of the chromosomes by
half and allow genetic diversity.
Definitions Box:
1. Reproduction: is a biological process by which a new offspring individual organism are made from their
parents
2. Sexual reproduction: involves the formation of a new individual, which receives its chromosomes from
gametes; half from the spermatozoa and half from the oocyte.
3. Asexual reproduction: reproduction without the fusion of gametes
4. Zygote: A diploid cell resulting from the fusion of two haploid gametes; a fertilised ovum.
5. Haploid set of chromosomes: a set of 23 chromosomes
6. Diploid set: a cell with a complete set of chromosomes= 46 chromosomes
7. Gamete: A mature haploid male or female germ cell that is able to unite with another of the opposite sex in
sexual reproduction to form a zygote.
8. Gonads: an organ that produces gametes: testis and ovary
9. Oogenesis: the process by which an ovum is formed
10. Homogametic sex : when both the sex chromosomes are X and all the oocytes are similar to one another
(only females)
11. Heterogametic sex: the pair sex chromosomes that contain both the X and Y chromosomes, so producing
two distinct populations of spermatozoa, one bearing an X and the other a Y chromosome.
12. Spermatogenesis: the production of the mature spermatozoa
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Mitosis
Each human cell contains 23 pairs of homologous chromosomes resulting in 46 chromosomes in
total. The aim of mitosis is to transfer the parents genome into two identical daughter cells.
Mitosis (non-sexual reproduction):
1. Interphase: before division, cell grows and duplicates both its centrioles and the DNA in
each of its chromosomes.
2. Mitotic prophase: nuclear membrane breaks down
3. Mitotic metaphase: microtubules from a mitotic spindle between the two centrioles and the
chromosomes lie on its equator.
4. Mitotic anaphase: centromere of each chromosomes splits and the two chromatids in each
chromosomes migrate to opposite poles on a spindle.
5. Mitotic telophase: reformation of nuclear membrane and nucleoli; division of the cytoplasm
into two daughters; breakdown of the spindle and decondensation of the chromosomes (no
longer visible under the light microscope). Two genetically identical daughter cells exists
now
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Meiosis
Meiosis is the random distribution of male and female homologous chromosomes, where
chromosomes crossing-over occurs
Meiosis involves two divisions
Main difference between meiosis and mitosis is the crossing-over stage, which only occurs in
meiosis allowing more variation.
Meiosis occurs before birth in females and in males it occurs at puberty.
Purpose: reduction division, which means it reduces the number of chromosomes by half,
resulting in haploid daughter cells. It also aims to allow genetic recombination.
Meiosis has the same phases as mitosis but with the addition of the following:
Interphase: Before division, DNA is replicated resulting in 92 chromosomes.
Prophase 1: homologous chromosome become pairs and crossing-over occurs between
these chromosomes pairs.
Metaphase 1: Independent assortment occurs, which is the random alignment pattern of the
homologous pairs.
Prophase 1 and Metaphase 1: Genetic recombination occurs here.
Anaphase 1: Homologous chromosome pairs separate and each chromosome migrates to
the opposite pole on a spindle.
Telophase 1: Each daughter cell is diploid. Chromosome no: 46
Telophase 2: Each daughter cell is haloid (23 chromosomes)


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Gametes
Gametes are highly specialised cells which are produced in processes known as oogenesis and
spermatogenesis. Gametes originate from germ cells and are made in the male and female gonads:
testis and ovary. Each gonad makes a specific sex steroid hormone, which modifies the body tissues
to generate specific male or female phenotypes.
How are gametes produced?
Females: Oogonia
All Female gametes are called Oogonia and their production occurs during foetal life.
Oogenesis:
Oogenesis is one of the most highly specialised and regulated cellular biological process
known.
At 4-6 weeks of embryonic life, about 100 germ cells, which originate extragonadally,
migrate to the gonadal ridges (genital ridges), where the germ cells will be sexually
differentiated into oogonia.
Within the gonads, the oogonia will go through many cycles of mitotic division over several
months in order to proliferate their numbers.
By 7 months, there will be about 7 million oogonia. Each oogonia is then surrounded by a
few layers of cells which are the precursor of the follicle. The oogonia is now called an
oocyte.
Cessation of Mitosis so there will be no more opportunities for the oogonia to increase their
numbers. When meiosis begins, the oogonia is called an oocyte.
When the baby is born, the oocyte cease in the first stage of meiosis until it does or is
ovulated.
During ovulation, the oocyte grows and we can observe the re-imitation of meiosis. During
meiotic maturation we observe the 1
st
meiotic reduction and ovulation of the egg.
When the sperm penetrates the oocytes, the 2
nd
meiotic reduction takes place.
What is the follicle and how does it protect the oocyte?
The follicle is a cellular structure surrounding and protecting the oocyte.
A large proportion of the 7 million eggs die, so that at birth the ovary contains less than l
million eggs and by 7 years of age, there are 3-4 x 10
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, each within a follicle (Figure 5). Of
these only 0.1% (approximately 400) will ovulate.
Most follicles become atretic and die in the pre-antral stage (see later). It should be made
clear that the follicles and ova that die off are in not defective and if induced to mature by
excess exogenous hormones most will be capable of normal fertilization, as for instance, in
IVF. The large wastage is probably evolutionary, ie survival of the fittest.
The oocytes then begin the first meiotic prophase and remain at this stage until ovulation.
The oocyte is embedded in 2 or 3 layers of granulosa cells. These are the cells that produce
oestradiol.
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These structures (egg+ cells) are called primordial or resting follicles
After puberty some follicles increase their size and the number of granulosa cells, which
secrete glycoprotein around the oocyte forming the zona pellucida. These are primary
follicles.
Stromal cells of the ovary condense around the follicle forming the theca which becomes
vascularised.
The theca is separated from the granulosa cells by a basal lamina through which all
nutrients, hormones, etc must diffuse.
The granulosa cells remain avascular, to protect the oocyte. The signals initiating primordial
follicles growth are largely unknown.
Once the theca forms, gonadotrophins are definitely required for further growth and at this
stage FSH causes the follicle to grow through the primary to the secondary to the pre-antral
phase

Figure 5. Numbers of oogonia and then follicles throughout female lifespan


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As the numbers of granulosa cells increases the follicle expands and areas of fluid appear in
the centre. These coalesce and are called the antrum..hence antral follicles. The oocyte is
displaced to one side and protrudes into the antrum on a stalk of granulosa cells. The follicle
undergoes maxiumum growth by cell dividion and antrum expansion; it reaches 20mm in
diameter by ovulation.
The oocyte is surrounded by a ring of granulosa cells some of which send processes through
the zona pellucida and via these supply nutrients to the egg. A substance is also passed
which is believed to keep the oocyte in the 1
st
phase of meiosis.
There is a small rise in FSH between the end of one cycle and beginning of the next (see
notes on menstrual cycle) and at this time a group of 4 or 5 antral follicles will be present in
the ovary. One follicle will be better placed to respond to the FSH rise than the others. It will
rapidly produce oestrogen which re-exerts negative feedback and causes the FSH to drop
again. This prevents development of any further follicles. Those follicles not selected will
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become atretic. The selected or dominant follicle, in the meantime, grows significantly and
pushes up to the surface of the ovary and protrudes from it.
Essentially, LH stimulates androstenedione and testosterone production from theca.
FSH acts to induce aromatisation of androgens to oestrogen in granulosa cells. The enzymes
required to produce androgens are only in theca and that for oestrogen only in granulosa. At
this stage FSH receptors are only in granulosa and LH only on theca. Thus there is a two cell-
2 gonadotrophin steroidogenic system;
Dominant Follicle.
Only one follicle undergoes the changes described above, becoming the dominant follicle. In
the late follicular phase, FSH induces LH receptors on granulosa cells in the dominant follicle.
LH can now act on granulosa cells, stimulating aromatase and hence oestrogen. This allows
the dominant follicle to survive the fall in FSH. It is the dominant follicle that is solely
responsible for the huge increase in production of oestradiol in the first half of the
menstrual cycle. The granulosa cells divide to give approximately 60 million which is how an
exponential rise in serum E
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is achieved. The dominant follicle reaches 20mm in diameter
prior to ovulation, ie grows from 5-20mm in 2 weeks.






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Oocytes Structure/Function/Characteristics
Oocyte to an egg:
Egg is the only cell capable of giving rise to a new individual
Roles: provides nutrition, mitochondria and spindle.
The zona Pellucida and basal lamina protects the egg.
Susceptible to damage because it has a long life and remains in the first phase of meiosis for
a long period of time, which means DNA can get damaged.
Contains nuclear DNA, 46 chromosomes and the mitochondrial DNA is present outside the
nucleus.
Oocyte growth:
Egg is the largest cell---100m
Can increase 1000 fold in mass from when the follicle starts to grow
Egg in selected follicle grows approximately 300x during meiotic arrest
Oocyte characteristics:
Germinal vesicle:
This is the nucleus of the immature oocyte arrested in meiosis
Pale, plane appearance with single nucleolus
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Nucleus to cytoplasmic volume greatly reduced with growth
Nucleolus changes from fibrillogranular to fibrillar, increasing in density
Cortical granules:
Small spherical membrane bound organelles (lysosomes) probably derived from golgi
Fuse with the oolemma at fertilisation, release contents
Alters ZP, acts as a second block to polyspermy
Heterogeneity in the contents

Zona Pellucida:
Thick extra-cellular coat increases in thickness with growth
Permeable to large macromolecules
First sign of oocyte growth in ovarian cortex
Created from fine filaments that push between follicle cells
Filaments later create mesh
Follicle cell extensions continue through it
Required for acrosome reaction
Golgi ribosomes and mitochondria specialities?
CAMP holds the oocyte in the meiotic arrest because it inhibits the cell from completing
meiosis. However other factors are also involved such as other cells/ovarian environment,
steroids/calcium and the integrity of the follicle
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Meiotic maturation involves:
Germinal vesicle breakdown
visible undulations in surface
nuclear envelope breaks appear
nuclear envelope disperses into the EpR
nucleolus disperses into the cytoplasm
Chromosome condensation
Spindle formation
Separation of homologous chromosomes
Polar Body emission



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Males
Gamete: Spermatozoa
The functions male gonads:
To produce mature germ cells (spermatogenesis)
To produce hormones that maintain the reproductive tract and reproductive processes.
90% of the testes consist of seminiferous tubules which are tightly coiled and make up the fabric of
the gonad. They lead to an area on one side of the testis called the ret. The walls of the tubules, the
seminiferous epithelium, are made up of tall columnar endothelial cells called Sertoli cells and
between these, lying on the basement membrane made of peritubular cells are the primary germ
cells called spermatogonia. The spaces between the tubules, the interstitial spaces, are filled with
Leydig cells and also macrophages, blood and lympathetic vessels; all are bathed in interstitial fluid.
The Sertoli cells secrete agents that maintain spermatogenesis and the Leydig cells are the main site
of production of testosterone. The passage of hormones and nutrients necessary for the energy-
demanding process of spermatogenesis all occurs via the interstitial fluid since the seminiferous
tubules are avascular. The volume of interstitial fluid within the testes is very high; such a volume
would be considered oedemic in any other tissue. The testes lie in the scrotum outside the
abdominal cavity at a temperature 1.5-2.5
o
C below core body temperature. This is essential for
sperm production. Testicular structure is outlined in figs 1 and 2
Spermatogonia lie on the basement membrane of the seminiferous epithelium. Every 16 days, a
wave of them start to divide mitotically into primary spermatocytes or daughter spermatogonia. The
daughter spermatogonia keep the pool replenished whereas the primary spermatocyte is committed
to undergo development towards being a sperm. The primary spermatocytes possess a diploid
number of chromosomes i.e. 46; these cells divide meiotically to form secondary spermatocytes
possessing 23 chromosomes (see figure 3 and 4 for mitotic and meiotic division).
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Figure 2. Cartoon of section through a seminferous tubule. Sperm progress from the basement
membrane towards the lumen as they mature and divide. The Leydig cells are in contact with
small capilliaries and produce androgens, the Sertoli cells are avascular and convert androgen to
oestrogen.
No further divisions occur as the secondary spermatocytes differentiate into spermatids. The
differentiation changes the shape of the sperm and is called spermiogenesis (figure 5). All these
processes occur within the seminiferous epithelium, in the spaces between the Sertoli cells (figure 2
& 3). After each division the germ cells move toward the lumen of the tubule, so that eventually the
spermatids are embedded in the apical region of the Sertoli cells (figures 1 and 2) from where they
are extruded into the lumen. As the spermatozoa move into the lumen, they lose part of their
cytoplasm. This cyclical process is completed in 64 + 4 days. In addition, there are waves of
development occurring every 16 days, longitudinally along the tubule, so that at a given site there
are more mature stages in front and less mature stages behind (figure 4).
At each stage the germ cells need different agents for their maintenance and development. These
factors are all produced by the Sertoli cells.
Blood-Testes Barrier
Sertoli cells are irregular in shape and adjacent cells are joined by tight junctions immediately above
the spermatogonia. Just before meiosis, the tight junction parts and allows the germ cell to move
from the basal (outside) to the adluminal (inside) compartment: it then closes again. These tight
junctions prevent molecules passing into the intercellular spaces. This means that all molecules
passing to the spermatocytes etc. must pass through the blood-testis barrier, or be secreted into it
by the Sertoli cells(similar to oocyte). Note that once meiosis has taken place the haploid germ cells
are "foreign" protein so the blood-testis barrier isolates the germ cells from the rest of the body
(including circulation) and protects them from the immune system. In addition, it allows the haploid
germ cell to exist within the adluminal compartment in a different and unique ionic
microenvironment with especially high levels of certain amino acids etc which may be required for
germ cell production.
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THE DUCTUS EPIDIDYMUS
The sperm, having reached the lumen of the seminiferous tubules, pass into a common area (the
ret), where all the tubules open out. From there they pass into the ductus epididymus. The
epididymus is a tube of 5 metres in length and highly coiled. Passage of sperm through the
epididymus takes 8-14 days. It can be divided into three parts; caput, corpus and cauda (Latin for
head, body and tail): in the epididymus fluid is reduced, motility gained and maturation advanced.
The spermatozoa becomes "progressively" motile; this means it can progress forward (Figures 7 and
8 show the structure of the mature sperm).


Fig 3. Drawings comparing spermatogenesis and oogenesis. Oogonia are not shown in this figure
because all oogonia differentiate into primary oocytes before birth. The chromosome complement
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of the germ cells is shown at each stage. The number designates the total number of chromosomes,
including the sex chromosome(s) shown after the comma. Note that (1) following the two meiotic
divisions, the diploid number of chromosomes, 46 is reduced to the haploid number, 23; (2) four
sperms form from one primary spermatocyte, whereas only one mature ovum results from
maturation of a primary oocyte; and (3) the cytoplasm is conserved during oogenesis to form one
large cell, the mature oocyte of ovum. The difference is because a large number of sperm are
required which only have to function as transporters of DNA whereas the large egg cytoplasm must
support multiple cellular divisions following fertilization. Each cell is highly specialised to perform its
function.



Figure 5. During spermiogenesis the rounded spermatid gradually reduces its cellular contents and
becomes elongated. The mitochondria move behind the nucleus, the tail develops and the head
develops an envelope containing the acrosomal enzymes.
The sperm are mainly stored in the epididymus, but also in the ampulla of the vas deferens (figure
1). The spermatozoa exist in epididymal fluid, but at ejaculation secretions from the accessory sex
glands are added to form seminal fluid and make up a large percentage of the total volume. These
secretory glands are : the seminal vesicles, the prostate, the bulbo-urethral gland.
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Figure 6 and 7. Structure of a mature spermatoan (plural spermatozoa) showing the streamlined
shape of the head encompassing the acrosome, the power pack of mitochondria in the middle
piece and the comparative length of the tail



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Sperm develop in a syncytial arrangement:
As sperm pass through stages of development and division occurs, the progeny of a single
spermatogonium remain connected by cytoplasmic bridges.
This is advantageous because sperm has completed meiosis prior to most of the
maturational stages.
Each sperm only has haploid chromosomes and some are X and some are Y.
How are sperm adapted to their job?
Motile
Very small
Aerodynamic (has a shape that reduces drag from air); important as it moves through the
uterine fluid
Acrosome
No cytoplasm and mitochondria
How will our understanding of oocyte and sperm function be useful in clinical practice?
Contraception
Assisted reproduction
Freezing eggs or part of ovary for cancer patients before chemotherapy/radiotherapy


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