The Vanguard Sperm Cohort of the Boar Ejaculate is Overrepresented in the Tubal Sperm Reservoir In Vivo

In boars, sperm cohorts are sequentially emitted in epididymal cauda fluid and resuspended in different Mixtures of accessory sex gland secretions while ejaculated in various fractions. During natural mating, these ejaculate fractions sequentially enter the cervix-uterine lumen, are quickly transported towards the tips of the uterine horns and colonize the oviductal sperm reservoirs (SR). Using a simple experiment, we tested the hypothesis that the first ejaculated sperm subpopulation (fortuitously present in the peak portion of the sperm-rich fraction [SRF], the so-called Portion 1, P1) is, by reaching first the SR, overrepresented there. Spermatozoa from P1- and of P2- (last portion of the SRF and the Post-SRF) were collected from 3 fertile boars. PI-spermatozoa were fluorophore DNA-stained, while P2-spermatozoa were kept unstained. Weaned estrous sows were conventionally inseminated (12 h after onset of estrus) with similar sperm numbers (approx 10(.) x 10(9) spermatozoa) per portion but in different orders as follows: (i) a mix of P1 and P2 aliquots (control, P1+P2, n=5), or testing (ii) a sequential order (P1-P2, Treatment A, n=5) or (iii) an inverse order (P2-P1, Treatment B, n=5) of cohort AI. Sows were euthanized approx 3 h post-AI, and the SRs were flushed to recover the spermatozoa, which were accounted for as stained or unstained. The total number of spermatozoa flushed did not differ between groups or boars (NS, ranging 0.9 to 2.0 x 10(9)). Sequential, in vivo-like, sperm deposition (P1-P2, Treatment A) yielded the highest proportion of stained PI-spermatozoa in the SRs (59.8 +/- 5.66%, means +/- SEM) compared with when the order was reversed (P2-P1, Treatment B; 15.6 +/- 2.1% P1-spermatozoa, P<0.05) or P1 and P2 sperm Suspensions were mixed (control, 36.9 +/- 2.70% P1-spermatozoa, P<0.05). The tested hypothesis proved valid; if inseminated in the same order as ejaculated in vivo, P1-spermatozoa become overrepresented in the SR. The physiological consequences of this skewed SR-colonization are discussed in this paper alongside the advantageous use of P1-spermatozoa for handling, including cryopreservation.