The aim of study was to evaluate the effects of insulin-like growth factor I (IGF-I) on canine sperm function during cooled and freeze-thaw storage. Extenders supplemented with different IGF-I concentrations (0, 100 and 200 ng/ml) were added to canine spermatozoa, and the sperm samples were stored at 4°C for 48 hr or freeze-thawed. Sperm motility, morphology, plasma-membrane integrity (PMI) and mitochondrial membrane potential (MMP) were evaluated. IGF-I had no effect on PMI or morphology during cooling and freeze-thawing. However, IGF-I alleviated the reduction in progressive motility and MMP caused by cooled storage and led to an improvement in MMP after freeze-thawing. In conclusion, IGF-I can be helpful to maintain progressive motility of canine spermatozoa during hypothermic storage via increased MMP.