Studies on the sperm reservoir of the pig oviduct

During sperm transport in the female pig a proportion of spermatozoa are arrested, often for 24 h or more, in a particular segment of the utero-tubal junction (UTJ) and the adjacent tubal isthmus, where a sperm reservoir (SR) is built up. The SR maintains, via its lining epithelium and the fluid it produces, sperm viability pre-ovulation. It also controls the release of potentially fertilising spermatozoa so that only a small sub-population reaches the site of fertilisation, thus diminishing the risk of polyspermy. In vitro research has focused on sperm binding as the main mechanism of sperm storage, sperm release and modulation of capacitation, but little attention has hitherto been paid to the isthmic oviductal fluid (IOF), its composition and the control of capacitation in vivo. This thesis aimed to study the intra-luminal milieu of the SR in sexually cycling gilts and sows, its contents of glycosaminoglycans (GAGs) — particularly of the non-sulphated hyaluronan (HA) — and the presence and expression of the specific HA receptor CD44 and of HA synthases. Both non-inseminated (controls) and inseminated animals were studied during specific moments of oestrus, in relation to spontaneous ovulation. Ultimately, the study aimed to determine the capacitation status of SR-stored spermatozoa and the effect of IOF and HA on capacitation of the SR-stored spermatozoa. The results showed that SR-stored spermatozoa are entrapped in a mucus-like IOF pre-ovulation. This IOF contains fluctuating levels of sulphated GAGs and HA. Hyaluronan is synthesised in the lining epithelium by HA synthase 3 (has3). Both the ligand and the specific HA-receptor CD44 were particularly present in the deep furrows of the SR, where most spermatozoa are trapped. Massive sperm capacitation does not occur in vivo in the porcine SR under spontaneous standing oestrus, particularly during pre- and peri-ovulation, but SR spermatozoa capacitate if exposed to the effector bicarbonate. Exposure of SR spermatozoa to IOF (or its component HA) in vitro was seen to reverse the bicarbonate influence during pre- and peri-ovulation but to potentiate capacitation post-ovulation, suggesting an active role for the intra-tubal fluid and/or HA in modulating sperm capacitation in pigs. The findings support the concept that the oviductal SR keeps the potentially fertile spermatozoa viable and uncapacitated during their pre-ovulatory arrest. The findings may help improve sperm preparation protocols for porcine in vitro fertilisation (IVF) and preservation of boar semen