fertilization

Sperm pathways

• follow a path from the site of deposition (vagina or cervix/uterus depending on species) toward the oviduct

• Barriers: sperm must navigate the cervix, the uterus, and the utero-tubal junction

The fate of most sperm

• vast majority of sperm (more than 99%) do not reach the egg

• Most are lost via retrograde loss (draining out of the tract), killed by the acidic environment of the vagina, or attacked by the female’s immune system (neutrophils)

Changes during capacitation

The final maturation process sperm undergo in the female tract (specifically the oviduct) to become fertile. Key changes include:

• hyperactivation

• Cholesterol removal

• Surface modification

Hyperactivation

Sperm develop a frantic, high-amplitude whip-like flagellar beat to provide the force needed to penetrate the egg coats

Cholesterol removal

Cholesterol is stripped from the sperm plasma membrane, destabilizing it to prepare for the acrosome reaction

Surface modification

Coating proteins and seminal plasma proteins are removed or relocated to expose zona pellucida (ZP) receptors

Primary sperm storage site

• The isthmus of the oviduct acts as the primary reservoir

• Sperm bind to the oviductal epithelium (cilia), which prolongs their lifespan and prevents them from rushing to the ampulla all at once

• They are released in small batches toward the site of fertilization

Evidence of the reacted acrosome

• experiments using fluorescently tagged sperm (often green fluorescent protein) demonstrated that sperm initiate the acrosome reaction while navigating the cumulus matrix, before they reach the zona pellucida

• It was shown that those successfully binding to and penetrating the ZP had already begin or completed the reaction

Importance of the acrosome reaction

The acrosome reaction is a chemical reaction”key” for the egg’s “lock”:

• enzyme release: it releases enzymes (like acrosin) that may help create a penetration slit in the zona pellucida

• Membrane exposure: it exposed the inner acrosomal membrane (IAM), which contains the specific proteins necessary for the sperm to bind to and fuse with the oocyte’s plasma membrane

IVF (In Vitro Fertilization)

• Hundreds of thousands of capacitated sperm are placed in a dish with an oocyte

• The sperm must naturally penetrate the cumulus and ZP on their own

ICSI (Intracytoplasmic sperm injection)

• a single sperm is picked up with a needle and injected directly into the cytoplasm of the oocyte

• This bypasses the need for sperm motility, the acrosome reaction, and ZP penetration

Steps of sperm fusion and oocyte activation

1. Sperm factor release: the sperm introduces an enzyme (PLC-zeta) that causes calcium (Ca2+) spikes in the oocyte

2. Cortical reaction: high calcium triggers the release of cortical granules

3. Meiotic completion: the oocyte finally completes meiosis II and expels the second polar body

Zona block (slow block)

• cortical granules release enzymes (like ovastacin) into the perivitelline space

• These enzymes “clip” ZP2 and inactivate ZP3, making the zona pellucida hard and incapable of binding more sperm

Membrane block

The oocyte plasma membrane changes its electrical potential or molecular structure to prevent further sperm fusion

Ovastacin

The specific protease in cortical granules that cleaves ZP2 to prevent polyspermy

Mutation outcome of ovastacin

• if mutated or inactivated, ZP2 is not cleaved after fertilization

• Consequently, sperm will continue to bind to the zona pellucida even after an embryo has formed, significantly increasing the risk of polyspermy (lethal for the embryo)

Formation of male pronuclei

After the sperm enters, the sperm head swells, and its DNA decondenses to form the pronucleus

Formation of female pronuclei

Simultaneously, the egg completes meiosis and its remaining chromosomes are enclosed in the pronucleus

Fate of male/female pronuclei

• the two pronuclei move toward each other (guided by microtubules)

• Their membranes breakdown and the chromosomes align on a single mitotic spindle (process called syngamy) forming a diploid zygote

First differentiation

• occurs at the blastocyst stage

• This is when cells first commit to different “fates” (becoming either part of the embryo or the placenta)

Embryo growth

• the embryo does not grow in total size during early cleavage (it is physically restricted by the zona pellucida)

• It only begins to grow in size after hatching from the zona pellucida (roughly 7-10 days post-fertilization)

Inner cell mass (ICM)

The internal clump of cells that will eventually form the fetus and all body tissues

Trophoblast

The outer layer of cells that will form the placenta and extra-embryonic membranes