Fertilization potential of cold-stored Fowler’s toad (Anaxyrus fowleri) spermatozoa: temporal changes in sperm motility based on temperature and osmolality.

Asynchrony of gamete release is problematic in amphibian captive breeding programs but can be overcome by short-term storage of spermatozoa. Hormonally induced sperm from the model species Anaxyrus fowleri were used to determine storage conditions for optimal fertilisation capacity. Sperm motility was measured over time, as a function of storage temperature (4°C or 22°C) and solution osmolality (7-40mOsm/kg). Sperm at 40mOsm/kg (spermic urine) stored at 4°C exhibited higher motility compared to 22°C. Also, sperm stored at 40mOsm/kg retained higher motility compared to sperm stored below 15mOsm/kg at both temperatures. Under optimal storage conditions (40mOsm and 4°C) a 30% decrease in sperm motility occurred within 24h, however, subsequent loss of sperm motility was lower (<10%/day) for days2-8 thereafter. Sperm samples stored for 1-8days under optimal conditions were tested for fertilising capacity by conducting in vitro fertilisation trials. Sperm stored for 8days yielded 48% neurula development, similar to sperm stored for 1day, which produced 60% neurula development. Overall, sperm stored for up to 8days at 4°C as spermic urine retained fertilising capacity and thus can be used to circumvent asynchronous gamete release in assisted breeding efforts for amphibians.