Fertilisation Flashcards
how do sperm gain their fertilising capacity?
Through multiple interactions with the female reproductive tract
When does sperm fertilising capacity begin?
It begins when sperm is deposited in the vagina, through the sperms journey (through the cervix, uterus, oviduct to the site of fertilisation) the sperm undergoes a series of changes and series of interactions and events that allow it to fertilise the oocyte. This process is known as sperm capacitation, it is not one process, it is a series of changes that occurs:
Define Sperm Capacitation
Sperm Capacitation - further and final maturation process which confers the following functional characteristics on to sperm and ultimately renders them capable of fertilisation:
1) A change to a state of hyperactivated motility.
whiplash flagellar movements by the sperm crucial for the penetration of the oocyte vestments.
2) The ability to bind to the oocyte’s zona pellucida and afterwards undergo the acrosome reaction.
Change in membrane properties of the sperm
3) The capacity to fuse with the oocyte
Female tract interaction mechanisms
Chemotaxis (has the most supportive data)
Is a chemical guiding mechanism.
Response to chemical gradient stimuli
Sperm responding to a gradient of chemoattractant e.g. steroid hormones from the cumulus oocyte complex such as progesterone.
Molecular and behavioural mechanisms yet to be fully understood.
Thermotaxis
Temperature guiding mechanism.
Female tract consists of different areas with marginal differences in temperature.
Sperm responding to changes in the extracellular temperature gradient.
Rheotaxis
Ability to respond to fluid currents in the female tract environment.
Response to direction of fluid currents
Boundary-following navigation
Takes advantage of sperms Ability to turn corners in response to surface boundaries
Human sperm with preference to follow boundaries on the left or right hand side have been shown to possess higher DNA integrity than straight swimming sperm (Huang et al., 2014; Eamer et al., 2016).
A lot of the stimuli resulting in the mechanisms:
Chemotaxis
Thermotaxis
Rheotaxis
Boundary-following navigation
Are mediated via the CatSper channel
What are the Cat-SPer channels?
CatSper=Cation channel of sperm
This is the principal sperm calcium channel that is located in the flagella area.
It is responsible for calcium influx, and this calcium influx is what triggers any downstream signalling and any interaction mechanisms at the molecular level.
The CatSper is characterised to respond to a large range of stimuli:
Changes in membrane potential
Changes in pH
Progesterone
Prostaglandins
Other small organic molecules.
One of the key events during sperm capacitation is the loss of cholesterol from the cell membrane.
What does this do?
This alters the phospholipid : cholesterol ratio. This will affect membrane fluidity and the availability of certain soluble proteins in the cell.
One way to characterise cholesterol loss in a lab is to have sperm cell media and supplement the media with a cholesterol acceptor, you will see that an increased proportion of sperm cells in that culture will undergo capacitation (eg switch to hyperactivated motility). An example of an acceptor is serum albumin.
The change in membrane fluidity due to cholesterol loss affects the availability of soluble membrane proteins such as soluble adenylyl cyclase.
CatSper Channel
There is a natural inhibitor to the CatSper channel called 2AG
In the absence of any activation 2AG is responsible for keeping the CatSper channel inactive.
When cholesterol binds to ABHD2, this complex will breakdown 2AG -> AA + Glycerol. And as it breaks down the 2AG it releases the CatSper channel and allows for calcium influx from the extracellular environment.
Soluble adenylyl cyclase (SACY) is a
membrane protein
In order for it to be activated we need calcium influx and bicarbonate influx
Once SACY is activated
It converts ATP into cAMP
cAMP then will activate PKA
PKA then facilitates the tyrosine phosphorylation of specific proteins involved in the changes associated with sperm capacitation such as hyperactivated motility.
SO… Removal of cholesterol is likely to alter membrane fluidity via change in cholesterol/phospholipid ratios. This in turn could affect the activity of surface membrane proteins like SACY.
Sperm-oocyte interaction
Once a few capacitated sperm make it to the site of fertilisation, they come in contact with the cumulus-oocyte complex (COC) (in the oviducts) and a much higher concentration of follicular fluid (FF).
FF in addition to secretions from the COC e.g. progesterone further modulate the spermatozoa to initiate the process of sperm-oocyte interaction and subsequent fertilisation.
Penetration of the cumulus oophorus
Sperm-zona binding – Where sperm binds to ZP
Acrosome reaction
Sperm-oocyte fusion and oocyte activation
List of chronological sperm-oocyte interaction
- Penetration of the cumulus-oophorus
- Spem-zona binding
- Acrosome reaction
- Sperm oocyte fusion and oocyte activation
Penetration of the cumulus-oophorus
At this point sperm cells are capacitated with whiplash tail movements giving them the momentum required to get through the cumulus cell mass
A visible feature of sperm capacitation is the switch from progressive motility to hyperactivated motility, characterised by whiplash tail movements. This is necessary for penetrating the oocyte vestments.
Once sperm come in contact with the cumulus-oophorus cell mass surrounding the oocyte, the enzyme hyaluronidase present on the surface of the sperm head dissolves hyaluronic acid (hyaluronan) – the major (binding) cementing factor between cells that constitute the cumulus oophorus cell mass.
Sperm Zona binding
Sperm cell is now at the zona pellucida – only one makes it past the cumulus cells
Four zona pellucida (ZP) glycoproteins expressed – ZP1, ZP2, ZP3, ZP4.
Studies across mammalian species have shown that sperm bind to ZP2 and ZP3 via receptors present on the sperm’s plasma membrane.
Recent studies by Yauger et al. (2011) and Baibakov et al. (2012) show that human sperm bind to ZP2.
Acrosome Reaction
The release of the exocytosis enzymes that are present in the sperm acrosome region.
Triggered in response to sperm-oocyte interaction or in response to signals in the female reproductive tract.
There is fusion of the sperm’s plasma membrane with it’s outer acrosomal membrane.
This results in perforations in the membrane resulting in the release of acrosomal enzymes:
Results in the release of hydrolytic and proteolytic enzymes e.g. hyaluronidase and acrosin.
Different sub populations of sperm undergo the acrosome reaction at different points, some in the COC some in ZP. BUT the ones that make it to the oocyte are the ones that undergo the acrosome reaction at the right place at the right time.
Sperm oocyte fusion & oocyte activation
Recent studies have shown that there is a protein binding that occurs in order for sperm oocyte fusion to be successful called Juno-Izumo binding.
When sperm successfully fuses with the oocyte there is an intracellular release of calcium from the intracellular stores in the oocyte which is usually the endoplasmic reticulum.
This triggers a cortical reaction, where there is release of cortical granules which prevents further sperm from entering.
This intracellular calcium spike triggers oocyte activation:
Release from meiotic arrest and completion of meiosis II.
At the point of ovulation the oocyte is at metaphase 2.
Incorporation of sperm DNA and pronuclear formation.
Start of embryo cleavage
The completion of the second meiotic division is characterised by the extrusion of the 2nd polar body.
Oocyte activation
How does the sperm trigger oocyte activation?
We know that sperm oocyte fusion triggers activation
The sperm introduces Phospholipase C Zeta (PLC ζ) into the oocyte to trigger activation
It is a soluble sperm protein that is introduced in the oocyte to trigger the events of oocyte activation
Phospholipase C Zeta (PLC ζ) Acts by activating inositol trisphosphate (IP3) production
It breaks down PIP2 in the oocyte -> IP3 + DAG
IP3 will bind to the IP3 receptors on the endoplasmic reticulum leading to repetitive calcium release (oscillations) from the oocyte’s calcium stores.
