X-Message-Number: 24515 From: "Basie" <> Subject: Improving the success rate Date: Sun, 15 Aug 2004 23:13:23 -0400 Maybe we can learn something. Salts, nutrients, buffer compounds? <For successful cryopreservation, sperm must be emersed into a special solution before freezing. The solution, called an extender, always contains salts, nutrients, buffer compounds and most importantly, a cryoprotectant> Frozen semen is a hot topic among horse breeders by Jenny Tye (Guelph, October, 1996) Improving the success rate of stallion sperm preservation will mean an increase in breeding options, say U of G researchers. Cryopreservation is the process of deep-freezing cell, tissues or gametes for later use. However, in horses, variation in sperm survival following freezing, between individual stallions, hinders this technology. Veterinarian researchers Claire Plante, John Pollard, Walter Johnson and DVSc Student Tracey Chenier, theriogenology, are identifying factors which underlie semen variability in response to freezing. By studying and improving individual elements of the freezing process they hope to gain new insight into why variation exists -- and how to overcome it. "Currently, only a third of stallion sperm will freeze well; the rest freezes poorly or not at all," says Chenier. "We want to develop a freezing method which, upon thawing, will result in the highest number of viable sperm for the highest number of stallions." For successful cryopreservation, sperm must be emersed into a special solution before freezing. The solution, called an extender, always contains salts, nutrients, buffer compounds and most importantly, a cryoprotectant. A cryoprotectant is a chemical -- commonly glycerol -- which protects the cell from damage during freezing. Chenier is testing the toxicity and permeability of different cryoprotectants in equine sperm. These chemicals, after long exposure, are potentially poisonous to cells, yet under the right circumstances they can be used to the researcher=s advantage. Permeability is an issue because the more easily a cryoprotectant can cross a sperm's membrane, the more efficiently it protects the cell during freezing. Plante and Pollard are determining which extender (other than cryoprotectants) offer the best support for sperm during freezing. They're examining the individual performance of each ingredient, as well as how different extender ingredients perform together. To this end, the researchers are freezing semen in different extenders and then evaluating the sperm's motility and viability after thawing. Once the researchers know which cryoprotector and maintenance ingredients work best in horse semen cryopreservation, the information will be combined to generate more effective extenders. The next challenge will be to determine what concentration of each ingredient, when combined, provides the best freezing support for the majority of horse semen. Since there are hundreds of possible concentration variations, Plante and Pollard will be using a computer program called Simplex Optimization, developed by Statistical Programs, to help with the elimination process. The sperm from those extender variations which result in good sperm viability after freezing and thawing will be used to artificially inseminate mares to collect pregnancy rates. For each group of solutions used, pregnancy rates, sperm motility and other variables will be entered into the program. It will sort and analyze the data and inform the researchers how to alter individual concentrations to get increased pregnancy rates in future trials. There are many advantages to deep-freezing techniques in the horse breeding industry. Of primary interest, the costly and dangerous transport of animals for the purpose of breeding between countries can be replaced by the transport of the sperm itself. Long term semen storage is also a valuable means of banking genetic information for the future. "Even at this early stage of the project, we're learning new information about the nature of horse sperm everyday," says Plante. "If we can develop improved extenders, artificial insemination with preserved semen will become a more reliable and popular breeding option for horse owners and veterinarians." This study is sponsored by the Ontario Veterinary College and the Ontario Ministry of Agriculture, Food and Rural Affairs. Frozen semen is a hot topic among horse breeders by Jenny Tye (Guelph, October, 1996) Improving the success rate of stallion sperm preservation will mean an increase in breeding options, say U of G researchers. Cryopreservation is the process of deep-freezing cell, tissues or gametes for later use. However, in horses, variation in sperm survival following freezing, between individual stallions, hinders this technology. Veterinarian researchers Claire Plante, John Pollard, Walter Johnson and DVSc Student Tracey Chenier, theriogenology, are identifying factors which underlie semen variability in response to freezing. By studying and improving individual elements of the freezing process they hope to gain new insight into why variation exists -- and how to overcome it. "Currently, only a third of stallion sperm will freeze well; the rest freezes poorly or not at all," says Chenier. "We want to develop a freezing method which, upon thawing, will result in the highest number of viable sperm for the highest number of stallions." For successful cryopreservation, sperm must be emersed into a special solution before freezing. The solution, called an extender, always contains salts, nutrients, buffer compounds and most importantly, a cryoprotectant. A cryoprotectant is a chemical -- commonly glycerol -- which protects the cell from damage during freezing. Chenier is testing the toxicity and permeability of different cryoprotectants in equine sperm. These chemicals, after long exposure, are potentially poisonous to cells, yet under the right circumstances they can be used to the researcher=s advantage. Permeability is an issue because the more easily a cryoprotectant can cross a sperm's membrane, the more efficiently it protects the cell during freezing. Plante and Pollard are determining which extender (other than cryoprotectants) offer the best support for sperm during freezing. They're examining the individual performance of each ingredient, as well as how different extender ingredients perform together. To this end, the researchers are freezing semen in different extenders and then evaluating the sperm's motility and viability after thawing. Once the researchers know which cryoprotector and maintenance ingredients work best in horse semen cryopreservation, the information will be combined to generate more effective extenders. The next challenge will be to determine what concentration of each ingredient, when combined, provides the best freezing support for the majority of horse semen. Since there are hundreds of possible concentration variations, Plante and Pollard will be using a computer program called Simplex Optimization, developed by Statistical Programs, to help with the elimination process. The sperm from those extender variations which result in good sperm viability after freezing and thawing will be used to artificially inseminate mares to collect pregnancy rates. For each group of solutions used, pregnancy rates, sperm motility and other variables will be entered into the program. It will sort and analyze the data and inform the researchers how to alter individual concentrations to get increased pregnancy rates in future trials. There are many advantages to deep-freezing techniques in the horse breeding industry. Of primary interest, the costly and dangerous transport of animals for the purpose of breeding between countries can be replaced by the transport of the sperm itself. Long term semen storage is also a valuable means of banking genetic information for the future. "Even at this early stage of the project, we're learning new information about the nature of horse sperm everyday," says Plante. "If we can develop improved extenders, artificial insemination with preserved semen will become a more reliable and popular breeding option for horse owners and veterinarians." This study is sponsored by the Ontario Veterinary College and the Ontario Ministry of Agriculture, Food and Rural Affairs. Rate This Message: http://www.cryonet.org/cgi-bin/rate.cgi?msg=24515