Fertilization - fusion of 2 gametes ensure encounter of sperm & oocyte/egg External fertilization # of gametes stimulus for release Internal fertilization copulatory organs receptors PH-30 humans activation of metabolic dormant oocyte/egg - different stages of oogenesis fusion of 2 haploid nuclei - syngamy to produce diploid embryo

The Acrosomal Reaction. Acrosomal reaction: when exposed to the jelly coat the sperm’s acrosome discharges it contents by exocytosis. Hydrolytic enzymes enable the acrosomal process to penetrate the egg’s jelly coat..

B locks to Polyspermy 1. Fast Block - short-lived Na + channels in the egg’s plasma membrane opens. Na + flows into the egg and the membrane depolarizes 2. The Cortical Reaction - slow, long-lasting Ca 2+ from the eggs ER is released into the cytosol and stimulates cortical granules to fuse with the plasma membrane and release their contents into the perivitelline space. The vitelline layer separates from the plasma membrane. An osmotic gradient draws water into the perivitelline space, swelling it and pushing it away from the plasma membrane. The vitelline layer hardens into the fertilization envelope: a component of the slow block to polyspermy. The plasma membrane returns to normal and the fast block to polyspermy no longer functions.

Activation of the Egg, High concentrations of Ca 2+ in the egg stimulates an increase in the rates of cellular respiration and proteins synthesis. The sperm nucleus swells and merges with the egg nucleus  diploid nucleus of the zygote. DNA synthesis begins and the first cell division occurs.

In amphibians a rearrangement of the egg cytoplasm occurs at the time of fertilization. The plasma membrane and cortex rotate toward the point of sperm entry. The gray crescent is exposed and marks the dorsal surface of the embryo.

Fertilization in Mammals. Capacitation, a function of the female reproductive system removes inhibitory enzymes on the sperm head which allows the acrosomal reaction to occur. A capacitated sperm migrates through a layer of follicle cells before it reaches the zona pellucida. Binding of the sperm cell induces an acrosomal reaction.

Voltage Clamps and Fast Block to Polyspermy

Cleavage follows fertilization. Functions of cleavage: Multicellular for differentiation The zygote is partitioned into blastomeres. Each blastomere contains different regions of the undivided cytoplasm and thus different cytoplasmic determinants. Restores Somatic Nuclear to Cytoplasmic Ratio 1:500 -> ->-> 1:6 sea urchin fertilized egg at end of cleavage - somatic cell Controls on # of cleavage divisions? 2n = 6 divisions 4n = ? n = ?

Holoblastic unequal where there is more yolk at the vegetal pole. Cleavage occurs more rapidly in the animal pole than in the vegetal pole. first two cleavages are vertical. The third division is horizontal. The result is an eight-celled embryo with two tiers of four cells Yolk is most concentrated at the vegetal pole and least concentrated at the animal pole. In animals with less yolk there is complete division of the egg: holoblastic cleavage. Holoblastic equal

A blastocoel forms within the morula  blastula

In birds the yolk is so plentiful that it restricts cleavage to the animal pole: meroblastic cleavage.

Superficial cleavage in insects Holoblastic equal in mammals

What Controls Cleavage? Information in Cytoplasm – Source? Spiral Cleavage in Snails Rt = Dextral (dominant) Left= recessive Female dd X male DD F1 genotype vs. phenotype F1 cross dD X dD Prediction? Genotype Phenotype

 Gastrulation rearranges the embryo into a triploblastic gastrula. –The embryonic germ layers are the ectoderm, mesoderm, and endoderm. Gastrulation rearranges the blastula to form a three-layered embryo with a primitive gut Copyright © 2002 Pearson Education, Inc., publishing as Benjamin Cummings

Fate map – vital stain Invagination

Involution Epiboly

Avian Development. Cleavage is meroblastic, or incomplete. Cell division is restricted to a small cap of cytoplasm at the animal pole. Produces a blastodisc, which becomes arranged into the epiblast and hypoblast that bound the blastocoel, the avian version of a blastula.

During gastrulation some cells of the epiblast migrate (arrows) towards the interior of the embryo through the primitive streak Ingression (immigration)

Once again, the embryonic membranes – homologous with those of shelled eggs. Chorion: completely surrounds the embryo and other embryonic membranes. Amnion: encloses the embryo in a fluid-filled amniotic cavity. Yolk sac: found below the developing embryo. Develops from the hypoblast. Site of early formation of blood cells which later migrate to the embryo. Allantois: develops as an outpocketing of the embryo’s rudimentary gut. Incorporated into the umbilical cord, where it forms blood vessels.

Activation of embryonic genome Mexican axolotl o-mutant strain The “o” gene is a recessive gene “O” gene is the normal, dominant gene In embryos obtained from female axolotls homozygous for gene “o”, development is always arrested during gastrulation. WHY? “O” protein is necessary to activate the embryonic genome. PROOF?

The eggs can be rescued by injecting eggs of o/o females with cytoplasm from normal eggs. Where is the "corrective component" (O protein) produced? Rescue Experiments Cytoplasm from Egg vs. Primary Oocytes Cytoplasm vs. Primary Oocyte fluid from germinal vesicle

What is the “O” protein doing?

activation of the embryonic genome. How can we prove it?

When is mammalian embryonic genome activated? Two-cell stage determined by radioactive labeling experiments Imprinting of mammalian gamete’s nucleus Experiments combining: Female pronucleus combined with Female pronucleus Male pronucleus combined with Male pronucleus