Hypothalamic Pituitary Gonadal Axis

Question 1

Describe the HPG axis

Answer 1

It begins with a positive drive from the hypothalamus. The hypothalamus synthesises and secretes GnRH to the anterior pituitary.
The gonadotroph cells in the anterior pituitary synthesise and secrete FSH and LH to the gonads.
The cellular compartments of the gonads synthesise and secrete steroid hormones (progesterone, estrogen and testosterone).
These feedback at different points on the anterior pituitary and the hypothalamus.
In males the feedback is ALWAYS negative
In females the feedback is primarily negative EXCEPT from the LH surge required for ovulation.

Question 2

What are two regions in the hypothalamus that express GnRH neurons?

Answer 2

GnRH neurons in the hypothalamus are expressed in two regions:
- The arcuate nucleus (infundibular nucleus)
- Medial preoptic nucleus

These neurons synthesise and secrete GnRH

Question 3

Hypothalamus–> Pituitary (anterior)

Answer 3

GnRH will make its way through the medial eminence to the anterior pituitary.
It does this through the hypophyseal portal system (hypophyseal portal circulation).
The hypophyseal portal system is a network of blood vessels that connect the hypothalamus and the anterior pituitary, allowing GnRH to make its way to the gonadotroph cells of the anterior pituitary.

Question 4

What is adenohypophysis?

Answer 4

Adenohypophysis = Anterior pituitary (gonadotroph cells)

Question 5

Kisspeptin History

Answer 5

Kisspeptin was originally called metastin because of it’s ability to suppress metastatic spread of human melanomas and breast carcinomas.

Kiss1 gene was discovered in 1996
Peptide products = Kisspeptins discovered in 2001
Kisspeptin receptor = GPR54

Question 6

What is kisspeptin (KISS1/Kiss1)?

Answer 6

Kisspeptin has a role as the gatekeeper of puberty
It controls the synthesis and secretion of GnRH in GnRH neurons
It functions upstream of GnRH
It is expressed in the hypothalamus, both in the arcuate nucleus (ARC) and the anteroventral periventricular nucleus (AVPV)
The expression of kisspeptin is regulated by gonadal steroids
There are regions in kisspeptin neurons that express the estrogen receptor (ER) and the progesterone receptor (PR).

Question 7

Describe the kisspeptin structure?

Answer 7

It is initially synthesised as a pre pro protein, it then undergoes proteolytic cleavage into different kisspeptins.
Following proteolytic cleavage of the prepro protein, you have kisspeptin-54, which is a 54 peptide sequence.
Kisspeptin-54 then undergoes further cleavage into Kp-14 -> Kp-13 -> Kp-10
Kp-54 is mainly used in in vivo studies as it is able to cross the BBB.
Kp-10 is used in in vitro studies as Kp-10 is a very active ligand of the GPR54 receptor. It is also cheaper to make a 10 peptide sequence than a 54.

Question 8

Kisspeptin receptor (GPR54/KISS1R)

Answer 8

GPR54 is expressed on GnRH neurons
The kisspeptin/KISS1R system is a component trigger if puberty.
Kisspeptin binding to its receptor has to occur in order to trigger puberty.
We know this as inactivating mutations of KISS1R and kisspeptin result in failure to undergo spontaneous puberty
Conversely activating KISS1R mutations (activate receptor without kisspeptin binding) can lead to precocious puberty.

Question 9

How does KISS1 regulate GnRH secretion?

Answer 9

Kisspeptin neurons upstream send projections of Kisspeptin to GnRH neurons, and this binds to the KISS1R expressed on the GnRH neurons.

A bolus of kisspeptin correlates with a peak of LH secretion.
GnRH activity is measured by measuring LH, as LH secretion is GnRH driven.
We measure LH to it being practical and the ethical considerations.
In order to take GnRH samples, you will have to access hypophyseal portal circulation in the brain or CSF samples which is impractical.

There are sexual dimorphic differences between males and females in how kisspeptin regulates GnRH activity
Cyclical differences

Question 10

Kisspeptin-mediated regulation of GnRH

Answer 10

The sex steroids are produced by the gonads downstream of the hypothalamus and pituitary.

There is negative feedback whereby the estrogen, progesterone and testosterone mediate feedback via Kisspeptin neurons located on the arcuate nucleus (ARC).
In turn GnRH is down regulated and so is the expression of FSH and LH.

BUT when the estrogen level threshold has been reached and there is a LH surge, there is a switch from negative to positive feedback.
This feedback is mediated via the kisspeptin neurons on the anteroventral periventricular nucleus (AVPV).
This in turn results in the upregulation of GnRH and therefore the LH surge.

Question 11

GnRH

Answer 11

GnRH is synthesised and secreted from specialised neurons of hypothalamus- GnRH neurons.
It is secreted in a pulsatile fashion - pulse generator is responsible for this activity.
The pulse is crucial
The pulse generation is a series of specialised neurons that modulate the pulsatile fashion of GnRH

GnRH stimulates the synthesis and secretion of pituitary gonadotrophin hormones - LH and FSH.

Question 12

GnRH structure

Answer 12

Like kisspeptin, it is synthesised as a prepro protein before it undergoes proteolytic cleavage into GnRH which is a decapeptide (a 10 peptide sequence), another product that is expressed alongside it is GAP (GnRH Associated Protein).

Question 13

GnRH pulsatility

Answer 13

GnRH is secreted in pulses from hypothalamus every 30-120min

A GnRH pulse stimulates a pulse of LH and FSH secretion from the anterior pituitary

Pulsatile GnRH secretion is vital for stimulation of synthesis and secretion of LH/FSH
Slow frequency pulse favours FSHb transcription (gene) and FSH release
Rapid pulse frequency favours LHb transcription (gene) and LH release
Continuous release results in cessation of response

Question 14

Briefly describe the role of GnRH pulsatility on FSH/LH secretion

Answer 14

Slow GnRH pulse = FSH
Rapid GnRH pulse = LH

Question 15

What is the GnRH Pulse Generator?

Answer 15

A population of neurons in the arcuate nucleus (ARC) mediates the pulsatility with possible input from other areas of the brain.

A new study found: A sub-population of Arcuate Nucleus Kisspeptin Neurons (ARNKISS)

ARNKISS were all found to exhibit intermittent periods of synchronized electrical activity which had a near perfect correlation with pulsatile LH secretion.
Inhibition of ARNKISS in the mid-caudal region of the arcuate nucleus suppressed pulsatile LH secretion

Question 16

Mode of GnRH action

Answer 16

GnRH binds to GnRHR
This triggers signal transduction
This results in the expression of the FHb and LHb genes (gonadotroph gene expression)
Then gonadotroph synthesis
Then gonadotroph secretion (FSH and LH)

Question 17

Gonadotrophin hormones: LH, FSH (and hCG)

Answer 17

They are glycoprotein hormones
A protein that has a sugar residue attached to it (post translational modification -glycosylation).

They are heterodimeric
They consist of two sub units that are different

LH, FSH and hCG all share a common alpha subunit.
It is the beta subunit that gives the distinct properties / functions.
The beta subunit is also crucial for the pulsatility
The beta subunits are synthesised and expressed in response to GnRH pulastility. The alpha subunit is produced constitutively.

Question 18

Structures of Gonadotrophins (FSH and LH)

Answer 18

You have a heterodimeric configuration that is made of an alpha and a beta subunit.
The alpha subunits are the same.
The alpha subunits are held together by disulphide bonds (N)
(y) = the attached sugar residues

What gives the beta subunit its uniqueness:
Length of the sequence
Type of and location where glycosylation that occurs

Question 19

Structure of the Gonadotrophin Receptors

Answer 19

They have a very large extracellular region where FSH and LH bind to.
This is also the region where there can be inactivating/activating mutations
Overall they have: Extra-cellular, trans-membrane and intra-cellular region.

Question 20

Pituitary–> Gonads: What is the function of gonadotrophins (FSH)?

Answer 20

Follicle-stimulating hormone (acts via FSHR)

Testis in males:
Regulation of Sertoli cell metabolism

Ovary in females :
Follicular maturation
Granulosa cell estrogen synthesis

Question 21

What is the function of gonadotrophins (LH)?

Answer 21

Luteinising hormone (acts via LHR)
Testis:
Stimulation of Leydig cell androgen synthesis
Ovary:
Theca cell androgen synthesis (estrogens -> androgens)
Ovulation – LH surge
Progesterone production of corpus luteum
The CL via the LHR is crucial for progesterone production in the luteal phase of the menstrual cycle.

Question 22

Briefly summarise the function of gonadotrophins

Answer 22

To summarise: They support germ cell growth, development and maturation to produce oocyte and sperm competent for fertilisation.

Question 23

List some steroid hormones

Answer 23

Progesterone
Oestrogens
Oestradiol and also oestrone
Androgens
Androstenedione, testosterone, dihydrotestosterone

Question 24

Describe the male gonadal steroid production

Answer 24

Cross section of semifenerious tubules.
In the interstitial region between the tubules you have the Leydig cells that express:
Leydig cells express the LH receptor. Via the LHR, LH modulates the production of androgens:
Androstenedione, DHEA, Testosterone production
Inside the tubules are the Sertoli cells, they nurture the developing germ cells.
Sertoli cells express the FSH receptor. FHS modulates Sertoli cell metabolism, which is crucial for spermatogenesis. It is also responsible for the conversion of testosterone into DHT.

Question 25

Describe the female gonadal steroid production

Answer 25

Mature antral follicle, can see oocyte with follicular fluid inside
The outside lining is where the theca cells are located.
The theca cells express the LH receptor and produce testosterone and androstenedione.
Further inwards, the cells that surround the oocyte are called granulosa cells.
Granulosa cells express the FSH receptor and modulate the conversion of the androgens produced by the theca cells into estrogen (Oestradiol and Oestrone) via the enzyme aromatase.

Corpus luteum in the luteal phase
Corpus luteum expresses both the LHR and the FSHR.
As a result produces both progesterone and estrogens in the luteal phase

Question 26

Inhibin

Answer 26

Peptide hormones
Heterodimers
Two subunits, alpha and beta
2 isoforms of inhibin: A and B.
Inhibin is produced by the gonads and inhibits FSH secretion via direct negative feedback to anterior pituitary.
Anterior pituitary has inhibin receptors, through which inhibin mediates direct negative feedback.

Males = Inhibin B isoform
Females = Depends on when in the cyclical dependent. But there is usually a switch from inhibin B in the follicular phase to inhibin A in the luteal phase.

Question 27

Female HPG axis

Answer 27

Starts in the hypothalamus where the kisspeptin neurons send projections of kisspeptin to the GnRH neurons.
In response to this, the GnRH neurons synthesise and secrete GnRH.
GnRH acts on the gonadotroph cells of the anterior pituitary
In turn this secretes FSH and LH
LH acts on the theca cells to produce androgens.
FSH acts on the granulosa cells and converts the androgens to estrogens.
Estrogen and inhibin feedback negatively on the anterior pituitary and on the hypothalamus. Progesterone also feedbacks negatively.
BUT in ovulation, estrogen feedbacks positively