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Introduction QuizQuiz 09

Spermatogenesis is initiated in the male testis with the beginning of

puberty. This comprises the entire development of the spermatogonia

(former primordial germ cells) up to sperm cells. The gonadal cordsthat are solid up till then in the juvenile testis develop a lumen with the

start of puberty. They then gradually transform themselves into

spermatic canals that eventually reach a length of roughly 50-60 cm.

They are termed convoluted seminiferous tubules (Tubuli seminiferi

contorti) and are so numerous and thin that in an adult male testicle

their collective length can be 300 to 350 meters. They are coated by a

germinal epithelium that exhibits two differing cell populations: some

are sustentacular cells (= Sertoli's cells) and the great majority are the

germ cells in various stages of division and differentiation .

Quiz

Quiz 10

Fig. 8 -

Convoluted seminiferoustubules

Fig. 9 -

Convoluted seminiferoustubules

Legend

In certain

forms of

impotence and

for an

assisted

fertilization

the testicle

is

"biopsied":

from a piece

of testicularparenchyma

the contorted

seminiferous

tubules are

extracted

with forceps

and unwound.

Fig. 8

Testicular

biopsy:

Pulling with

the forcepsmakes the

testicular

canals

visible.

Fig. 9

The tubules

are clearly

visible using

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a binocular

magnifying

glass.

(Video, 390

kB)

Commentary

For an optimal sperm cell production a certain milieu is needed. By

transferring the testicles into the scrotum a testicular temperature 2-3

C lower than body temperature is attained. In addition, a slightly

elevated pressure from the surroundings is necessary. This is why when

the taut tunica albuginea is slit open, the testicular parenchyma bulges

out by itself. Evidently, both elevated pressure and lowered

temperature are necessary for producing sperm cells.

Fig. 10 - Convoluted seminiferous tubules Legend

1

2

3

4

56

7

8

Basal lamina (membrane) (not recognizable)

Myofibroblast

Fibrocyte

Sertoli's cell

SpermatogoniaVarious stages of the germ cells during

spermatogenesis

Spermatozoon

Lumen

Fig.10

Histological

preparation of a

section through

a convoluted

seminiferous

tubule in an

adult. Outside

its basal lamina

a layer of

myofibroblasts

and fibrocytes

surround the

tubule. The

germinal

epithelium lies

on the tubule

wall. One can

recognize the

spermatogonia

sitting on the

basal lamina.

The nuclei of

the Sertoli'ssustentacular

cells have a

rarified

chromatin and

the nuclei with

clear nucleolus

that are often

oriented

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perpendicular to

the basal

lamina. The

overall picture,

though, is

dominated by the

cells occupiedwith

spermatogenesis.

The development of the germ cells begins with the spermatogonia at

the periphery of the seminal canal and advances towards the lumen over

spermatocytes I (primary spermatocytes), spermatocytes II

(secondary spermatocytes), spermatids and finally to mature sperm

cells.

Structure of the germinal epithelium QuizQuiz 08

The epithelium consists of Sertoli's sustentacular cells and the

spermatogenic cells. The Sertoli's cells form a single-layered lamina andextend from the basal lamina to the tubule lumen. With their

labyrinthine cellular processes they surround the individual types ofgerm cells more or less completely. Spermatogenesis is thus

accomplished in close contact with the Sertoli's cells, which not onlyhave supportive and nourishing functions, but also secrete hormones

and phagocytize cell fragments. Somewhat above the basal lamina theyare bound to each other through complicated occluding junctional

complexes (tight junctions), so that 2 separated compartments are

present in the epithelium: a basal one, in which the spermatogonia are

lined up, and a luminal one, in which all the other stages of

spermatogenesis are found.

Quiz

Quiz 11

Fig. 11 - Germinal epithelium Legend

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1

2

3

4

5

6

7a

7b

8

9

1011

A

B

Peritubular cells

Basal membrane

Spermatogonia

Tight junction

Spermatocyte I

Spermatocyte II

Spermatids

Spermatids

Acrosome

Residual bodies

SpermatozoasCell nucleus of sustentacular cells (Sertoli)

Basal zone

Adluminal zone

Fig.11

Schema of the

germinal

epithelium: The

supportive

(Sertoli) cells

sit on the basalmembrane.

Towards the

lumen of the

spermatogonia

(lowest row of

cells) the

Sertoli cells

are connected

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with each other

by the occluding

junctional

complexes (tight

junctions). This

seal gives rise

to the blood-testicle

barrier. The

cytoplasm of

these supportive

cells gets

formed into

complicated

processes

because they

surround all of

the cells

involved withspermatogenesis.

Commentary

Through the occluding junctional complexes of the Sertoli's cells a"blood/testicle" barrier is created in the tubule. This means that outside

this barrier, in the tubular periphery, cells, substances and hormonesfrom the blood have unhindered access.

On the other hand, the inner compartment of the tubule is protected bythe barrier, which is selectively permeable and serves as an entry

check. This is of practical importance because haploid cells in theinner part of the tubule exhibit surface antigenic properties, different

from all other body cells. They must thus be kept secluded from the

immune system of the organism by the "blood/testicle" barrier.

Developmental stages of spermatogenesis QuizQuiz 12

In the course of spermatogenesis the germ cells move towards the

lumen as they mature. The following developmental stages are therebypassed through:

Quiz

Quiz 13

Quiz

Quiz 14

y A-spermatogoniumy B-spermatogonium

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y Primary spermatocyte (= spermatocyte order I)y Secondary spermatocyte (= spermatocyte order II)y Spermatidy Sperm cell (= spermatozoon)

The spermatogenesis can be subdivided into two successive sections:

y The first comprises the cells from the spermatogonium up to andincluding the secondary spermatocyte and is termedspermatocytogenesis.

y The second one comprises the differentiation/maturation of thesperm cell, starting with the spermatid phase and is termed

spermiogenesis (or spermiohistogenesis).

The temporal course of spermatogenesis

The approximate 64 day cycle of the spermatogenesis can besubdivided into four phases that last differing lengths of time:

Mitosis of the

spermatogonia

16 days Up to the primary spermatocytes

First meiosis 24 days For the division of the primary

spermatocytes to form secondary

spermatocytes

Second meiosis A fewhours

For engendering the spermatids

Spermiogenesis 24 days Up to the completed sperm cells

Total ~64

days

Fig. 12 - The spermatogenesis generations Legend

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Fig.12

The stem cell

population of

the germinal

cells lies on

the basal

lamina of theconvoluted

seminiferous

tubules.

These are Type

A

spermatogonia.

These cells

undergo

mitosis: one

of the

daughter cells

renew thestock of type

A

spermatogonia,

the other

becomes a type

B

spermatogonia.

These divide

and their

daughter cells

migrate

towards thelumen.In

roughly 64

days they

differentiate

themselves

thereby into

sperm cells up

to the outer

surface of the

epithelium

(one should

note that inthese cellular

divisions, the

separation of

the cytoplasm

is not

complete.

Whole networks

of connected

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cells arise.

So, for

example in the

last

generation,

the

spermatids,far more cells

are bound to

each other

than as shown

here).

Spermatocytogenesis

Among the spermatogonia (all in all, over 1 billion in both testicles)

that form the basal layer of the germinal epithelium, several types

can be distinguished: certain type A cells are seen as spermatogonia

that divide mitotically and reproduce themselves (homonymous

division), whereby the spermatogonia population is maintained.

The beginning of spermatogenesis is introduced through the so-

called heteronymous division, in which the daughter cells (second

group of type A cells) remain bound together by thin bridges of

cytoplasm. Through the preservation of these cytoplasmic

connections, spermatogonia are inducted into the spermatogenesis

process.

After a further mitotic division type B spermatogonia are

engendered that also divide themselves mitotically into primaryspermatocytes (I).

The freshly created primary spermatocytes (I) now enter into the

first meiosis. They then go immediately into the S phase (that is, intothe preleptotene meiosis), double their internal DNA, leave the basal

compartment and reach the special milieu of the luminal

compartment. Following the S phase, these cells attain the complex

stage of the prophase of the meiosis and become thereby noticeably

visible with a light microscope.

This prophase, which lasts 24 days, can be divided into five sections:

y Leptoteney Zygoteney Pachyteney Diploteney Diakinesis

Commentary

In theheteronymous

division the

cytoplasmic

division is

not completed;

the daughter

cells stay

bound together

through thin

cytoplasmic

bridges.

Also in thesubsequent

meiosis the

cytoplasmic

division is

incomplete, so

that from one

spermatogonium

a network of

daughter cells

arises that

doubles in

size in eachgeneration.

The forming of

such networks

assures that

all of the

processes in

each

generation

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occur in step

witheach

other.

In the prophase in every germ cell a new combination of maternal and

paternal genetic material occurs. After the long prophase follow the

metaphase, anaphase and telophase that take much less time. One

primary spermatocyte yields two secondary spermatocytes.

Commentary

about

meiosis

The secondary spermatocytes go directly into the second meiosis, out of which the

spermatids emerge. Since in the secondary spermatocytes neither DNA reduplication nor arecombination of the genetic material occurs, the second meiosis can take place quickly. It

lasts only around five hours and for that reason secondary spermatocytes are rather seldomseen in a histological section. Through the division of the chromatids of a secondary

spermatocyte, two haploid spermatids arise that contain only half the original DNA content.

Besides the sperm cells the spermatids are the smallest cells of the germinal epithelium. In a

process lasting several weeks (so-called spermiogenesis or spermiohistogenesis) they are

transformed into sperm cells with the active assistance of the Sertoli's cells.

Local course of spermatogenesis - the

spermatogenesis wave

In examining a cross-section of a

convoluted seminiferous tubuleone notices that cells appear in

groups having the samematuration stages. However, not

all the spermatogenesis stages are

found in a cross-section.

Fig. 13 - Developmental stages Legend

1

2

3

Leptotene/zygotene of

the spermatocytes typ

I

Pachytene of the

spermatocytes

Fig.13

Various

developmental

stages in a

light

microscope

cross-

section

through a

convoluted

seminiferous

tubule.

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4

5

6

typ I

Young spermatids

Older spermatids

(sperm cells heads

can be recognized)

Sertoli's cells

Spermatogonia

On the one hand, the reason for this appearance lies in the fact that the

daughter cells, generated by each meiotic step, remain bound together

by thin cytoplasmic bridges. Thus with each meiotic step the following

generation is twice as large, until the cells have formed a relatively

complex network. The result is that cells of the same development

stages are seen there in groups. On the other hand, in addition, other

spermatogenesis generations are wound around each other in spirals

along the seminiferous tubule. This is why one meets with groupings of

various generations in a tubule cross-section. Thus, it is highlyimprobable that all of the development stages will be seen in a single

section at the same time.

Fig. 14 - Spermatogenesis wave Legend

Fig.14

This picture

shows three

waves of

spermatogenesis

generations.

As in the diagram, spermatogenesis waves move in spirals - like a

corkscrew - towards the inner part of the lumen. Outside, on the edge of

the tubule and at the beginning of the spiral, lie the spermatogonia; and,at the end of the spiral, the fully developed sperm cells are in the lumen.

From the diagram, it can be seen that several differing generations canbe found in a tubule cross-section. As time goes on, the wave of

spermatogenesis is shifted towards the right (as seen here) in order toalways newly begin again.

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Spermiogenesis (spermatohistogenesis) and

structure of the sperm cellQuiz

Quiz 15

The differentiation of the spermatids into sperm cells is called

spermiogenesis. It corresponds to the final part of spermatogenesis and

comprises the following individual processes that partially proceed atthe same time:

y Nuclear condensation: thickening and reduction of the nuclearsize, condensation of the nuclear contents into the smallest

space.

y Acrosome formation: Forming a cap (acrosome) containingenzymes that play an important role in the penetration through

the pellucid zone of the oocyte.

y Flagellum formation: generation of the sperm cell tail.y Cytoplasma reduction: elimination of all unnecessary

cytoplasm.

Quiz

Quiz 16

Quiz

Quiz 17

Quiz

Quiz 18

Fig. 15 - Spermiogenesis Legend

1

2

3

45

6

7

8

9

10

Axonemal structure, first flagellar primordium

Golgi complex

Acrosomal vesicle

Pair of centrioles (distal and proximal)Mitochondrion

Nucleus

Flagellar primordium

Microtubules

Sperm cells tail

Acrosomal cap

Fig.15

Three differing

stages of

spermiogenesis:

on the left a

fresh

spermatid, on

the right an

immature sperm

cell, and in

the middle an

in-between

stage. A

rotation of the

nucleus causes

a repositioning

of the

acrosomal

vesicle to

occur. This

inverts itself

like a cap over

the nucleus

that continues

to be condensed

(dotted line).

The cytoplasm

cell components

that are no

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longer needed

are discarded

and

phagocytized by

Sertoli's

cells. The

mitochondriaare packed

thickly

(tightly)

together around

the beginning

part of the

flagellum (mid-

piece). As a

sign of its

immaturity, the

sperm cell (on

the right) thathas issued into

the lumen still

has a bit of

cytoplasm

around its neck

(compare with

fig. 16 below).

Nuclear condensation

The nucleus becomes smaller, denser and takes on a characteristic,

flattened form. Seen from above, the nucleus is oval and, from thenarrow side, is pear-shaped. The acrosome lies over the tip. Nucleus andacrosome form the sperm cell's head that is bound to the mid-piece by a

short neck.

Acrosome formationThe Golgi complex engender the vesicles, which then merge into a

larger formation that settles close to the cell nucleus and finally inverts

itself like a cap over the largest part of the nucleus. The acrosome

corresponds functionally to a lysosome and thus contains lysosomal

enzymes (hyaluronidase among others).

Development of the flagellumThe future axonemal structure grows out of one centriole (distal). This

consists of a bundle of nine peripheral double microtubules and twosingle ones in the center. During its development, through the rotation

of the nucleus and acrosomal vesicle, the flagellum primordium comesto lie on the opposite side of the acrosome.

Four parts of the finished flagellum can be distinguished:

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y The neckcontains the two centrioles (proximal and distal)among other things.

y The mid piece consists of a sheath of ring-shaped mitochondriagrouped around the axoneme to provide the energy for the

flagellar movement.

y The principle piece has a sheath of ring fibers around theaxoneme.

y The tail consists of only the 9+2 structure of the axonemeThe mature sperm cell is approximately 60 Qm long and completely

enveloped by the plasma membrane.

Fig. 16 - The mature sperm cell Legend

12

3

4

5

6

7

8

9

10

11

12

A

B

C

D

E

Plasma membraneOuter acrosomal membrane

Acrosome

Inner acrosomal membrane

Nucleus

Proximal centriole

Rest of the distal centriole

Thick outer longitudinal fibers

Mitochondrion

Axoneme

Anulus

Ring fibers

Head

Neck

Mid piece

Principal piece

Endpiece

Fig.16

The mature

sperm cell

is slender;

in themiddle part,

the

mitochondria

are thick

and ring-

shaped. The

DNA in the

nucleus is

maximally

condensed.

Moreinfo

More

information

about this

illustration

Cytoplasmic reduction

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The cytoplasm of the spermatids that is no longer needed is phagocytized by Sertoli's cellsor is disposed of in the lumen of the tubules. A clump of cytoplasm, though, can remain

hanging on the neck and mid piece of the sperm cell for a little while.

During sperm cell production considerable individual variations exist that are also

partially influenced by psychological factors. Per day roughly 100 million sperm cells are

produced. It is said that in each ejaculate an average number of 50-200 million sperm cellsare present (WHO standard value: over 40 million).

Leydig's interstitial cells and hormonal

regulation

Between the seminal canals lie Leydig's interstitial cells. These are

endocrine cells that mainly produce testosterone, the male sexual

hormone, and release it into the blood and into the neighboring tissues.

An initial active stage of these cells occurs during the embryonicdevelopment of the testis. Later in juvenile life, due to the influence of

the LH (luteinizing hormone) secreted by the anterior hypophysis

(pituitary gland), Leydig's interstitial cells enter a second, long lasting

stage of activity. Together with the hormones secreted by the adrenal

cortex, testosterone initiates puberty and thus the maturation of the

sperm cells.

Fig. 17 - Leydig's interstitial cells Legend

1

2Leydig's interstitial cells

Crystalloids of Reinke

Fig.17

Group of large

cells in theinterstice between

tubules. Leydig's

interstitial cells

characteristically

contain large

protein crystals

(crystalloids of

Reinke), the

importance of

which is unknown.

The crystals are

uncolored andstand out as light

structures against

the red cytoplasm

of Leydig's

interstitial

cells.

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Commentary

Testosterone production is directed byLH (luteinizing hormone),

secreted by the anterior lobe of the hypophysis. Pronounced cycles inhormone production, as are present in women, do not exist.

The second hormone secreted by the anterior hypophysis, FSH(follicle-stimulating hormone) affects Sertoli's cells, in that it triggers

the formation of a testosterone-binding protein. Thereby testosteronecan be transported by Sertoli's cells into the luminal compartment and

there be concentrated. Testosterone is decisive for spermatogenesis.Testosterone is also carried away via blood and lymph fluid.

Testosterone has effects on all tissues, especially also on the brain

during development as well as on the sexual organs.