of Bovine Spermatozoa1

This study was conducted to determine the osmotic proper-ties of bull spermatozoa, including the effects of osmotic stress and cryoprotectant agent (CPA) addition and removal, on sperm motility. Semen from beef bulls was collected by electroejacu-lation and extended 1:3 in TL-Hepes containing 100 mg/ml py-ruvate and 6 mg/ml BSA. In solutions of 150–1200 mOsmolal (mOsm), bull spermatozoa behaved as linear osmometers (r2 5 0.97) with an osmotically inactive cell volume of 61%. The is-osmotic cell volume was 23.5 mm3. Motility was determined af-ter exposure to anisosmotic solutions ranging from 35 to 2400 mOsm and after return to isosmotic conditions. Retention of at least 90 % of isosmotic motility could be maintained only be-tween 270–360 mOsm. Bull spermatozoa were calculated to retain 90 % of their isosmotic motility at 92–103 % of their is-osmotic cell volume. Motility following a one-step addition and removal of 1 M glycerol, dimethyl sulfoxide, and ethylene glycol was reduced by 31%, 90%, and 6%, respectively, compared with CPA addition only. These data indicate that, during bull spermatozoa cryopreservation, osmotically driven cell volume excursions must be limited by exposure to a very narrow range that may be facilitated by the use of ethylene glycol as a CPA. gamete biology, sperm, sperm motility and transpor