Investigation of Various Cryopreservation Methods for Improvement of Canine Sperm Survival

Cryoprotectants are key factors affecting sperm survival during sperm cryopreservation. Apart from glycerol, the commonly used cryoprotectant, other permeating cryoprotectants (i.e. dimethyl sulfoxide, ethylene glycol and propylene glycol) have been tested in sperm freezing of various animal species. However, the efficiency of these permeating cryoprotectants (i.e. glycerol, dimethyl sulfoxide, ethylene glycol and propylene glycol) on canine sperm freezing has not been reported. In the present study, the cryoprotective effects of four cryoprotectants on Kunming dog sperm cryopreservation have been investigated based on two freezing extenders Tris-citric acid-fructose (TCF) and Lactose-Glucose DM (LG-DM). The performance of different concentration of cryoprotectants in Kunming canine sperm freezing was also been investigated. Sperm motility, motility recovery rate, plasma membrane and acrosomal integrity were examined to evaluate cryopreserved sperm function. The results showed that: Glycerol and ethylene glycol exhibited best cryopreserving effects while propylene glycol and DMSO showed little cryoprotection for Kunming canine spermatozoa, though propylene glycol displayed better effectiveness than that of dimethyl sulfoxide. Although 5% glycerol and 5% ethylene glycol yielded more than 40% of frozen-thawed sperm motility, which meets the need for in vitro fertilization (IVF) or artificial insemination (AI), 5% glycerol was the best for Kunming dog sperm freezing and had a significant difference compared with other treatments (P<0.05). On the contrary, propylene glycol or dimethyl sulfoxide was found to be unsuitable for the sperm freezing in Kunming dogs. Additionally, the function of permeating cryoprotectant appeared to be independent of extender types.