Sperm calcium levels and chlortetracycline fluorescence patterns are related to the in vivo fertility of cryopreserved bovine semen

Cryopreserved bovine semen is less fertile than fresh semen for reasons tha t have not been fully elucidated. Cryopreservation is known to disrupt the sperm plasma membrane and it induces premature capacitation of a sperm subp opulation, which may be a result of the increased internal calcium levels a fter thawing. To test the hypothesis that sperm intracellular calcium level is correlated with in vivo fertility, we used the fluorescent calcium indi cator, indo-1, and flow cytometry to assess intracellular calcium levels in frozen-thawed sperm from bulls of varying degrees of fertility. We also te sted a second hypothesis that the physiological status of sperm, as assesse d by the chlortetracycline (CTC) fluorescent assay, is correlated with fert ility. As detected by indo-1 fluorescence, the intracellular calcium level is negatively correlated with bull fertility immediately after thawing (P = .0362; n = 3 ejaculates from each of 10 animals). Moreover, there was a si gnificant difference between the 3 most and least fertile bulls over 4 hour s of incubation (P < .05; n = 3 ejaculates per bull). Finally, there was a positive correlation between sperm displaying the CTC acrosome reaction pat tern and fertility (P = .0014; n = 3 ejaculates from each of 10 bulls).