Effects of pre- and post-thaw thermal insults on viability characteristicsof cryopreserved bovine semen

The magnitude of damage to the viability of cryopreserved bovine spermatozo a by pre- and post-thaw thermal insults was compared. Semen collected by ar tificial vagina from 5 Holstein bulls was diluted in egg yolk-citrate-7% gl ycerol extender (EYCG) and cryopreserved in 0.5 mL French straws at a sperm concentration of 40 to 60 x 10(6) cells/mL. In Experiment 1, straws were s ubjected to 22, 5 or -18 degrees C static air temperature for a duration of 1, 2, 3, 4 or 5 min before or after thawing in a 37 degrees C water bath f or 1 min. Control straws were thawed in a 37 degrees C water bath for 1 min without further thermal insult. In Experiment 2, straws were thawed for 1 min in a 37 (control), 20 or 5 degrees C water bath, or were loaded into an insemination gun and plunged into a 37 degrees C water bath for 3 min. In both experiments, straws were returned to a 37 degrees C water bath for inc ubation prior to viability analysis. Viability evaluations, conducted in tr iplicate, included the percentage of motile spermatozoa at 1 min and at 3 h post thermal insult and the percentage of intact acrosomal membranes at 3 h post thermal insult. In both experiments, acrosomal integrity was more se nsitive than motility to thermal insult. In Experiment 1, a significant int eraction was observed between timing of thermal insult (pre- or post-thaw), static air temperature and duration of straw exposure. At 22 and 5 degrees C, thermal insults applied before thawing significantly (P<0.05) reduced a crosomal integrity at greater than or equal to 2 and greater than or equal to 4 min of exposure, respectively. However, post-thaw exposure to 22 and 5 degrees C for up to 5 min had no effect on any of the sperm viability para meters evaluated. In contrast, at -18 degrees C static air temperature, pos t-thaw exposure for greater than or equal to 3 min decreased acrosomal inte grity (P<0.05), while 5 min of pre-thaw exposure was required for alteratio n of acrosomal integrity. In Experiment 2, each alternative thawing method resulted in significantly (P<0.05) lower incubated acrosomal integrity rela tive to the controls. These findings suggest that bovine spermatozoa cryopr eserved in EYCG extender are more sensitive to pre-thaw than post-thaw ther mal insults and that acrosomal integrity following 3-h incubation at 37 deg rees C is superior to motility evaluations for detection of damage to sperm viability due to thermal insult. (C) 2000 by Elsevier Science Inc.