Capacitation status and fertility of fresh and frozen–thawed ram spermatozoa

Abstract

The effect of cryopreservation on the capacitation status and fertility of ram spermatozoa was observed. After the chlortetracycline staining technique was validated for ram spermatozoa, it was applied to fresh or long-term frozen-stored spermatozoa. Fresh spermatozoa displayed mainly the F pattern (non-capacitated; 61·3%), becoming B pattern (capacitated; 54%) and AR pattern (acrosome reacted; 41%) with incubation (6 h at 37°C). In contrast, frozen spermatozoa displayed the B pattern (65· 9%), becoming the AR pattern (64·2%) with incubation. This demonstrates that cryopreservation may cause membrane changes in ram spermatozoa functionally equivalent to capacitation. The differences in capacitation status did not affectin vitro fertilization rates between fresh and frozen spermatozoa, but pregnancy rates at Day 18 after intrauterine artificial insemination were higher for fresh than for frozen spermatozoa. This difference was not evident at Day 50, possibly as a result of the high embryonic loss between Days 18 and 50 when fresh unincubated and frozen incubated spermatozoa were inseminated. Further research is necessary to determine what part of the cryopreservation process is responsible for the membrane changes in ram spermatozoa.