Freezing of boar semen can be simplified by handling a specific portion of the ejaculate with a shorter procedure and MiniFlatPack packaging

Abstract

Low sperm survival post-thaw and time-consuming procedures for conventional freezing (CF) hamper the commercial application of cryopreserved boar semen. We had previously proven that boar spermatozoa in the first 10mL of the sperm-rich fraction, SRF (the so-called P1, the sperm-peak portion of the ejaculate) sustain best handling in vitro, since they probably bathe in an aliquot of seminal plasma (SP) with specific composition. Here, we performed three experiments to determine: Exp I: the concentration of bicarbonate among portions of the ejaculate; Exp II: the effects of bicarbonate doses on sperm motility and; Exp III: the outcome of a faster, simpler freezing method (SF), handling P1-spermatozoa packed in MiniFlatPacks (MFP) vs. CF and vs. SRF-spermatozoa (2x2 factorial design). The bicarbonate content in SP was, among portions/fractions of the ejaculate, lowest in P1 (13.71mM/L, P<0.0001, Exp I). Boar spermatozoa require bicarbonate in the extender (to the levels present in P1) to maintain acceptable motility over a 120-h period at 16-17 degrees C (Exp II). Sperm freezing was dramatically shortened (from 8 to 3.5h) by the SF-procedure. P1- and SRF-spermatozoa survived equally both CF- and SF-freezing (% total motility 30min PT; P1-CF: 65.2+/-5.4% and P1-SF: 68.9+/-2.4%; SRF-CF: 64.4+/-2.7%; SRF-SF: 55.8+/-3.1%, ns). Interestingly, in contrast to SRF, there were no significant variations in 30-min PT-survival among either ejaculates or boars when the P1 was frozen, independent of the handling method (CF or SF). In conclusion, such a faster freezing protocol of semen packed in MFP could be advantageously applied to P1-spermatozoa (P1-SF), while the rest of the ejaculated spermatozoa could still be used for production of conventional artificial insemination (AI) doses, thus allowing for a maintained routine management of commercially relevant stud boars.