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Freezing Bovine Semen. IV. Effect of Freezing on the Metabolic Activity of Bovine Spermatozoa during and after Storage at –79° C.^{1, 2,}

Dairy Breeding Research Center, Department of Dairy Science, The Pennsylvania State University, University Park

ABSTRACT

No evidence of respiratory activity by bull spermatozoa diluted in skimmilk-glycerol during storage for 6 mo. at –79° C. could be demonstrated as measured by production of C¹⁴O₂ from radioactive substrates present in the frozen medium. However, freezing resulted in harmful effects on post-thawing metabolism of spermatozoa. After 6 mo. at –79° C., twice-washed spermatozoa showed 34% less C¹⁴O₂ production from glucose-U-C¹⁴ during post-thawing incubation for 6 hr. at 5° C. than during a similar prefreezing incubation period. Unwashed sperm, frozen and stored for 1 hr., produced 61% less lactic acid during post-thawing incubation for 3 hr. at 37° C. than sperm from the same semen samples which were not frozen. The 36% loss of motile sperm due to freezing was not sufficient to explain the greatly reduced lactic acid production. Sperm stored frozen for 1 hr. in skimmilk-glycerol showed significantly greater post-thawing lactic acid production during incubation under air than those frozen in 1:4 egg yolk-citrate-glycerol, even though fewer sperm survived freezing in skimmilk. Adding 1.25% fructose to skimmilk-glycerol and egg yolk-citrate-glycerol diluents resulted in greater lactic acid production by both frozen, thawed and unfrozen spermatozoa. However, frozen samples still showed greatly depressed metabolic activity as compared to unfrozen samples.

¹ Authorized for publication on June 19, 1858, as Paper No. 2268 in the journal series of the Pennsylvania Agricultural Experiment Station.

² Supported in part by grants from the Pennsylvania State Association of Artificial Breeding Cooperatives.

³ The data contained in this paper are part of a thesis submitted by the senior author to the Graduate School of The Pennsylvania State University in partial fulfillment of the requirements for the degree of Doctor of Philosophy.