RESUMO
A large number of genetically modified mouse strains have been produced in recent years. Sperm cryopreservation is the most effective means of preserving these valuable strains, most of which have a C57BL/6 genetic background. However, the fertilization efficiency of sperm from several cryopreserved strains, including C57BL/6, is quite low. While new and improved methods of cryopreservation have been developed, the majority of sperm stocks have already been cryopreserved using traditional methods, such as storage in 18% raffinose and 3% skim milk (R18S3). Therefore, new thawing methods for these frozen stocks are needed. We have developed a new thawing method that involves selective collection of motile sperm and a preincubation medium that enhances capacitation. Motile sperm are selected simply by collecting a sample from the center of a dish, and capacitation is induced by the addition of methyl-beta-cyclodextrin, D-penicillamine, sodium citrate, and hypotaurine to modified Tyrode's solution. The fertilization rate of sperm prepared using this method was increased significantly compared to that of sperm thawed using the traditional method (63.9 vs 16.5%, P<0.01). These results demonstrate that this new in vitro fertilization method is an effective means of reviving C57BL/6 sperm cryopreserved in R18S3.
Assuntos
Criopreservação/métodos , Crioprotetores/farmacologia , Preservação do Sêmen/métodos , Capacitação Espermática/efeitos dos fármacos , Espermatozoides/efeitos dos fármacos , Animais , Feminino , Fertilidade/efeitos dos fármacos , Fertilidade/fisiologia , Fertilização in vitro , Nascido Vivo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Capacitação Espermática/fisiologia , Motilidade dos Espermatozoides/efeitos dos fármacos , Motilidade dos Espermatozoides/fisiologia , Espermatozoides/fisiologiaRESUMO
The DBA/2J mouse strain is a standard laboratory strain that is widely used for biomedical research. This strain, however, suffers from poor reproductive performance. In addition, the conditions for reliable embryo transfer (ET) of this strain have not been elucidated. The intention of this study was to determine the optimal number of embryos for transfer that allow the effective production of DBA/2J offspring. In the experiment, 7 to 15 embryos per oviduct were transferred into pseudopregnant ICR females. A relatively high success rate for pup production was observed when a large number of DBA/2J embryos (30 embryos per female) were transferred. This result shows that the ET efficiency of the DBA/2J strain can be improved by increasing the number of transferred embryos.