Fertilization - Last Minute Revision Highlights

1. Site & Timing

• Of 200-300 million sperm deposited, only 300-500 reach the ampulla. Only one fertilizes the egg.
• Sperm are guided by chemotaxis - the oocyte and cumulus cells secrete attractants.

2. Capacitation

• Duration: ~7 hours in the female reproductive tract
• Process: removal of glycoprotein coat and seminal plasma proteins from the sperm plasma membrane overlying the acrosomal region
• Location: mainly in the uterine tube (epithelial interactions with mucosal surface)
• Key point: Only capacitated sperm can pass through the corona radiata and undergo the acrosome reaction. Sperm that rush to the ampulla too fast are NOT yet capable of fertilizing.
• Capacitation can also occur in vitro (basis of IVF) and via GIFT or intraperitoneal insemination - it is NOT organ specific.

3. Phases of Fertilization (3 Phases)

Phase 1: Penetration of the Corona Radiata

• Capacitated sperm pass freely through follicular cells
• Enzyme hyaluronidase released from the acrosome disperses corona cells
• Mechanical tail movements also assist

Phase 2: Penetration of the Zona Pellucida

• Zona is a glycoprotein shell composed of ZP1, ZP2, ZP3 (and ZP4 in humans)
  • ZP1 cross-links ZP2 and ZP3 into a latticework
  • Sperm receptors bind to ZP3 → triggers acrosome reaction
  • ZP2 binds sperm pre-acrosin after acrosome reaction
• Acrosome reaction: after ZP3 binding, Ca²⁺ influx → outer acrosomal membrane fuses with sperm plasma membrane → exocytosis of acrosomal enzymes
  • Key enzymes: acrosin (serine protease - most important), hyaluronidase (spermatogenic cell-specific isozyme), neuraminidase, esterase
• After one sperm penetrates: zona reaction - cortical granules release lysosomal enzymes into perivitelline space → alters ZP2 and ZP3 → zona becomes impermeable to other sperm (block to polyspermy)

Phase 3: Fusion of Oocyte and Sperm Cell Membranes

• Initial adhesion: integrins on oocyte + disintegrins on sperm
• Fusion proteins on sperm: IZUMO, ADAMs 1/2/3, CRISP1
• Receptor on oocyte: Juno, integrins, CD9, CD81
• The plasma membrane over the acrosomal cap has disappeared (used in acrosome reaction), so fusion occurs at the posterior region of sperm head
• Both the sperm head AND tail enter oocyte cytoplasm; sperm plasma membrane and mitochondria remain outside

4. Oocyte Responses After Sperm Entry (3 Responses)

1. Cortical & zona reactions - cortical granule release → oocyte membrane impenetrable + zona pellucida altered → prevents polyspermy
2. Resumption of 2nd meiotic division - oocyte was arrested in metaphase II; sperm entry triggers completion:
   • Forms mature oocyte + 2nd polar body (very little cytoplasm)
   • Maternal chromosomes (22 + X) form the female pronucleus
   • Trigger: phospholipase C-zeta from sperm → Ca²⁺ oscillations → cell cycle reactivation
3. Metabolic activation of the egg - initiates early embryogenesis (activating factor carried by sperm)

5. Pronuclei Formation & Syngamy

• Sperm nucleus enlarges → male pronucleus (tail degenerates)
• Oocyte nucleus → female pronucleus
• Both pronuclei are morphologically indistinguishable
• Each pronucleus: 1n (haploid), 2c (two chromatids) - they replicate DNA first
• The oocyte at this stage = ootid
• Pronuclei come together, nuclear envelopes break down → chromosomes arrange on cleavage spindle → ZYGOTE (2n, diploid)
• Fertilization ends at metaphase of first mitotic division

6. Results/Outcomes of Fertilization (High-Yield Summary)

1. Stimulates oocyte to complete 2nd meiotic division
2. Restores diploid number (46 chromosomes) in the zygote
3. Produces genetic variation (independent assortment + crossing over)
4. Determines chromosomal sex of embryo (X-bearing sperm → 46,XX female; Y-bearing sperm → 46,XY male)
5. Causes metabolic activation and initiates cleavage of the zygote

7. Block to Polyspermy - Two Mechanisms

8. Key Molecules - Must Know

9. Clinical Correlates

• IVF: Capacitation possible in vitro; sperm doesn't need to contact female reproductive tract
• ICSI: Overcomes oligozoospermia/azoospermia; single sperm injected directly into oocyte cytoplasm; risk of Y-chromosome deletions and higher birth defect incidence
• Primary sex ratio: 1:1 at fertilization (equal X and Y sperm); secondary sex ratio at birth ~1.05 (105 males:100 females)
• Ectopic fertilization: Can occur in other parts of the tube, NOT in the uterine body
• Polyspermy: Results in triploid embryo (69 chromosomes) → lethal; prevented by zona & cortical reactions

Spermatogenesis - Last Minute Revision Highlights

1. Overview

• Begins at puberty (~13 years) and continues throughout most of life
• 80-90% of testis volume = seminiferous tubules + germ cells at various stages
• Total time: ~74 days (Langman's, Ganong's); Costanzo states ~64 days
• ~300 million sperm produced daily (Langman's); Costanzo: 128 million/day (from 2 million spermatogonia x 64 spermatids each)

2. Temperature Requirement

• Normal spermatogenesis requires 35-36°C (1-2°C below body temperature)
• Maintained by countercurrent heat exchange between testicular arteries and veins in the pampiniform plexus (scrotum outside body cavity)
• Cryptorchidism → elevated temperature → impaired spermatogenesis → infertility

3. Cellular Sequence (The Pathway)

Primordial Germ Cells (PGCs)
        ↓  [migrate to testes during embryogenesis]
Spermatogonial Stem Cells
        ↓  [puberty - begin dividing]
Type A spermatogonia (dark Ad + pale Ap)  ← self-renewing stem cells
        ↓  [mitosis]
Type B spermatogonia
        ↓  [mitosis - last mitotic division]
Primary spermatocytes  ← LARGEST germ cells in seminiferous tubule; 46 chromosomes (2n, 4c)
        ↓  [Meiosis I - prolonged prophase ~22 days]
Secondary spermatocytes  ← ~half the size; 23 chromosomes (1n, 2c); VERY short-lived
        ↓  [Meiosis II - rapid]
Spermatids  ← ~half the size; 23 chromosomes (1n, 1c); HAPLOID
        ↓  [SPERMIOGENESIS - no further division]
Spermatozoa (mature sperm)

4. Three Phases of Spermatogenesis (Costanzo)

5. Spermiogenesis - Key Steps (No Division!)

1. Formation of the acrosome - from Golgi apparatus; covers anterior 2/3 of nucleus; lysosome-like organelle containing hydrolytic enzymes (hyaluronidase, acrosin/proteolytic enzymes, neuraminidase)
2. Condensation of the nucleus - chromatin compacts tightly
3. Formation of neck, middle piece, and tail (flagellum)
   • Axoneme: central skeleton of 11 microtubules (9+2 arrangement, like cilia)
   • Middle piece: mitochondria wrapped around axoneme (supply ATP for motility)
   • Tail movement: rhythmic longitudinal sliding of microtubules (ATP-driven)
4. Shedding of most cytoplasm as residual bodies → phagocytized by Sertoli cells

6. Structure of Mature Spermatozoon

• Sperm cell membrane and mitochondria do NOT enter the oocyte at fertilization (left outside)

7. Sertoli Cells - Must Know Functions

8. Leydig Cells (Interstitial Cells)

• Located in connective tissue between seminiferous tubules, near capillaries
• 20% of adult testis = connective tissue with Leydig cells
• Function: synthesis and secretion of testosterone
• Stimulated by LH from anterior pituitary
• Cytoplasm full of lipid droplets (typical of steroid-secreting cells)
• Testosterone has local (paracrine) effects on Sertoli cells to support spermatogenesis + endocrine effects on distant organs

9. Hormonal Regulation

10. Blood-Testis Barrier - High Yield

• Formed by tight junctions between Sertoli cells (NOT between germ cells)
• Creates two compartments:
  • Basal: spermatogonia + early primary spermatocytes (outside barrier, accessible to blood)
  • Adluminal: later spermatocytes → spermatids → spermatozoa (inside barrier, immunologically protected)
• Crossing the barrier: spermatogonia/early spermatocytes cross from basal → adluminal compartment via Sertoli cell junctions
• Analogy: like the blood-brain barrier

11. Cytoplasmic Bridges - Important Concept

• From the moment Type A spermatogonia divide, the daughter cells remain connected by cytoplasmic bridges (cytokinesis is incomplete)
• All progeny of a single spermatogonium form a clone that stays connected until late spermatid stage
• Purpose: synchrony of differentiation across the clone
• Residual bodies = shed cytoplasm when bridges finally break → phagocytized by Sertoli cells

12. Sperm Maturation in Epididymis

• Sperm from seminiferous tubules: only slightly motile initially
• Passively transported to epididymis (via contractile elements in tubule wall)
• Acquire full motility in the epididymis
• Remain viable in epididymis for several months
• Capacitation then occurs in the female genital tract (~7 hours)

13. Clinical Correlates - High Yield

14. Key Numbers Summary

Spermatogenesis - Simple Language

Definition

The 3 Phases of Formation

Phase 1 - Multiplication Phase (Mitosis)

• The starting cells are called spermatogonia (stem cells sitting on the wall of the tubule)
• They divide by mitosis (normal cell division) repeatedly to increase their number
• Some stay behind as stem cells (to keep production going forever)
• The rest move forward and become primary spermatocytes
• At this point, each cell still has the full 46 chromosomes

Think of it as: raw material being duplicated before the factory starts processing it.

Phase 2 - Meiotic Phase (Meiosis)

• Primary spermatocyte (46 chromosomes) divides
• Produces 2 secondary spermatocytes (23 chromosomes each)
• This step takes the longest - prophase alone lasts ~22 days
• Crossing over happens here → genetic variation

• Each secondary spermatocyte divides again very quickly
• Produces 2 spermatids from each secondary spermatocyte
• So from 1 primary spermatocyte → 4 spermatids total
• Each spermatid has 23 chromosomes (haploid)

Think of it as: cutting a document from 46 pages to 23 pages, so when sperm meets egg (also 23), they make a complete 46-page human.

Phase 3 - Spermiogenesis

• Spermatids are round, simple cells - not yet sperm
• They now get shaped and structured into proper spermatozoa (no more division happens here)
• Four key changes occur:

Think of it as: a rough-cut diamond being polished into its final shape.

Simple Flow Chart

Spermatogonia (46 chr)
      ↓ Phase 1: MITOSIS
Primary Spermatocyte (46 chr)
      ↓ Phase 2a: MEIOSIS I
Secondary Spermatocyte (23 chr)
      ↓ Phase 2b: MEIOSIS II
Spermatid (23 chr) — round, immature
      ↓ Phase 3: SPERMIOGENESIS
Spermatozoon (23 chr) — mature sperm ✓

One-Line Summary of Each Phase

Oogenesis - Simple Language, Last Minute Revision

Definition

The BIG Difference vs Spermatogenesis

The Key Numbers (Memorize These)

The rest degenerate (become atretic). Nature is very wasteful with eggs.

Phases of Oogenesis

Phase 1 - Multiplication Phase (Before Birth)

• PGCs migrate from yolk sac wall to the developing ovary
• Once there, they become oogonia and divide rapidly by mitosis
• Peak: 7 million oogonia by the 5th month of fetal life
• Then oogonia start dying off (atresia begins)
• Meanwhile, some oogonia stop dividing and enlarge → become primary oocytes
• Each primary oocyte gets surrounded by a single flat layer of follicular cells → this unit = primordial follicle
• By the 7th fetal month, all surviving oogonia have become primary oocytes
• No new oogonia are EVER formed after birth

Phase 2 - Growth Phase (Fetal life → Puberty)

• Primary oocytes begin Meiosis I but get ARRESTED in Prophase I (specifically at diplotene stage)
• This arrest is caused by Oocyte Maturation Inhibitor (OMI) - a small peptide secreted by follicular (granulosa) cells
• Primary oocytes stay frozen here for years - up to 40-50 years if the woman ovulates late in life!
• This is why older mothers have higher risk of chromosomal errors (e.g. Down syndrome) - oocytes are aging in arrest
• The zona pellucida forms around the oocyte during this phase

Think of it as: a factory product that is made and then placed in a freezer, waiting for the right signal to continue.

Phase 3 - Maturation Phase (Each menstrual cycle, from puberty onward)

• Primary oocyte (46 chr) finishes its first meiotic division just before ovulation
• Produces:
  • Secondary oocyte - large cell, gets almost ALL the cytoplasm (23 chromosomes, 2 chromatids each)
  • 1st polar body - tiny cell with hardly any cytoplasm; may divide again but eventually degenerates
• Division is unequal - one cell is large, one is tiny (opposite of spermatogenesis where 4 equal cells are made)

• Secondary oocyte immediately begins Meiosis II but gets ARRESTED at Metaphase II
• This is the stage at which ovulation occurs
• What is released at ovulation: a secondary oocyte (NOT a mature ovum) - still arrested!

• If a sperm penetrates the secondary oocyte → Meiosis II is completed
• Produces:
  • Mature ovum (ootid/egg) - large, gets all cytoplasm; 23 chromosomes (1 chromatid each = haploid)
  • 2nd polar body - tiny, degenerates
• If NO sperm → the secondary oocyte degenerates without completing meiosis

Complete Flow Chart (Simple)

Oogonium (46 chr)
  ↓  MITOSIS (fetal life, before birth)
Primary Oocyte (46 chr)
  ← ← ARRESTED in Prophase I (diplotene) for years ← ←
  ↓  MEIOSIS I completed (just before ovulation, after LH surge)
Secondary Oocyte (23 chr) + 1st Polar Body
  ← ← ARRESTED in Metaphase II (ovulated in this state) ← ←
  ↓  MEIOSIS II completed ONLY IF sperm enters
Mature Ovum (23 chr) + 2nd Polar Body
  ↓
ZYGOTE (46 chr) after fusion with sperm (23 chr)

The Two Arrest Points - Must Know

Exam tip: What is released at ovulation? = Secondary oocyte (arrested at Metaphase II), NOT a mature egg. The mature egg only forms after fertilization.

Follicle Development (Paired with Oogenesis)

Polar Bodies - Quick Summary

• Polar bodies exist only to discard extra chromosomes without wasting cytoplasm
• Ovum keeps all the cytoplasm (nutrients for early embryo) - that's why divisions are unequal

Clinical Highlights

One-Line Summaries