|MadSci Network: Cell Biology|
Dear Natalie, I did a little research into your question.The following link has a detailed description regarding cryoprotectans used in equine sperm preservation http:www.gvequine.com.au/more_on_frozen_semen.htm. Here's the short version: There are two general classes of cryoprotectants: 1) penetrating cryoprotectants. These pass thorugh the sperm membrane and act both intracellularly and extracellularly, and 2) non-penetrating cryoprotectants, which act only extracellularly. Glycerol and DMSO are commonly ultilized penetrating cryoprotectants. Whereas non-penetrating cryoprotectants include proteins found in milk or egg yolk; sugars such as fructose , lactose, mannose, raffinose or trehalose; synthetic polymers such as polyvinylpyrrolidone or methyl cellulose and amides. Most penetrating cryoprotectants serve as both as a solvent and a solute. All non-penetrating cryoprotectants can not serve as a solvent. Glycerol serves as a solute within the water and also penetrates the sperm. Glycerol contributes to the osmotic pressure both outside and inside the spermatozoan. The non-penetrating cryoprotectants such as egg yolk, although are soluble, do not penetrate through the plasma membrane of a living cell Thus they contribute to the osmotic pressure of the extender solution. Non- penetrating cryoprotectants move water out of the sperm which results in dehydration and shrinkage. When sperm is frozen for cyropreservation, the addition of glycerol helps prevent ice damage. Glycerol has a lower freezing temperature than water. Apparently the process of thawing and insemination is potentially very damaging to the sperm due to rapid changes in volume.Frozen stocks typically have an osmotic pressure >1000 mOsmol/kg whereas the environment of the uterus is >300 mOsmol/kg. This results in a rapid increase in volume for sperm as water moves in to equlibrate the high concentration of intracellular glycerol (left behind). This may result in rupture of the plasma membrane. There is also evidence that addition of egg yolk proteins to the freezing medium aids in the recovery of sperm motility.Here's the reference for that study: Alvarez JG, Storey BT. (1993) Evidence that membrane stress contributes more than lipid peroxidation to sublethal cryodamage in cryopreserved human sperm: glycerol and other polyols as sole cryoprotectant.J Androl May-Jun;14(3):199-209 Yildiz C, Kaya A, Aksoy M, Tekeli T. (2000) Influence of sugar supplementation of the extender on motility, viability and acrosomal integrity of dog spermatozoa during freezing.Theriogenology Sep 1;54(4):579- 85 To learn more about the osmotic effects of permeable versus non permeable reagents: Liu Z, Foote RH.(1998) Osmotic effects on volume and motility of bull sperm exposed to membrane permeable and nonpermeable agents. Cryobiology 1998 Nov;37(3):207-18 Devireddy RV, Swanlund DJ, Roberts KP, Pryor JL, Bischof JC.(2000) The effect of extracellular ice and cryoprotective agents on the water permeability parameters of human sperm plasma membrane during freezing. Hum Reprod. May;15(5):1125-35. Best wishes, Elsa .
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