Cryopreservation of rainbow trout sperm by deep-freezing

Cryopreservation trials on rainbow trout (Salmo gairdneri) sperm were carried out using two basic extenders: Mounib's medium (M) and Ménézo's medium (Me) to which were added bovine serum albumin (BSA) and tellurite egg yolk (Institut Pasteur). After 10 p. 100 of DMSO was added to these different deep-freeze diluents (DD), they were mixed with the sperm and then deep-frozen into 100 microliter pellets on dry ice. The pellets were stored in liquid nitrogen for periods lasting from 3 days to 6 months. The intensity of sperm motility and fertilizing ability were measured before and after cryopreservation. After the sperm was diluted in Ménézo's medium, slight spermatozoon motility was noticed, which probably caused their early exhaustion and would explain the lower fertilizing ability observed after thawing. Mounib's medium gave better results, especially after 10 p. 100 of egg yolk was added. The optimal deep-freeze conditions were: 1/3 dilution, no equilibration after dilution but immediate deep-freezing at a rate of 10 to 40 degrees C/min. Thawing had to be carried out rapidly in 10 sec. However, the spermatozoa were altered during the freezing-thawing process, and during insemination more frozen spermatozoa had to be used to equal the fertilization rate obtained with non-frozen sperm. However, the fertile spermatozoa gave normal embryogenesis and no abnormal development was seen up to the vesicle resorption stage. At the end of spermiation, sperm fitness for deep-freezing decreased, perhaps due to sperm senescence. Pooling the sperm of several males partially compensated for the loss of fertilizing ability seen at the end of the reproductive period.