Mouse sperm begin to undergo acrosomal exocytosis in the upper isthmus of the oviduct

Abstract

Recent evidence demonstrated that most fertilizing mouse sperm undergo acrosomal exocytosis (AE) before binding to the zona pellucida of the eggs. However, the sites where fertilizing sperm could initiate AE and what stimuli trigger it remain unknown. Therefore, the aim of this study was to determine physiological sites of AE by using double transgenic mouse sperm, which carried EGFP in the acrosome and DsRed2 fluorescence in mitochondria. Using live imaging of sperm during in vitro fertilization of cumulus-oocyte complexes, it was observed that most sperm did not undergo AE. Thus, the occurrence of AE within the female reproductive tract was evaluated in the physiological context where this process occurs. Most sperm in the lower segments of the oviduct were acrosome-intact; however, a significant number of sperm that reached the upper isthmus had undergone AE. In the ampulla, only 5% of the sperm were acrosome-intact. These results support our previous observations that most of mouse sperm do not initiate AE close to or on the ZP, and further demonstrate that a significant proportion of sperm initiate AE in the upper segments of the oviductal isthmus.

Natriuretic peptide type C induces sperm attraction for fertilization in mouse

“…The number of spermatozoa with NPR2-positive staining was further increased after treatment with NPPC, suggesting that only the spermatozoa with functional NPR2 are attracted by NPPC. This is consistent with recent studies that only a few spermatozoa migrate from the isthmus to the ampulla during the progression of fertilization616263. Thus, the capacitated spermatozoa show attraction by exposing the functional receptor NPR2, and travel from the isthmus of the oviduct to the ampulla during ovulation.…”

Designing 3-Dimensional In Vitro Oviduct Culture Systems to Study Mammalian Fertilization and Embryo Production

“…Usually, such ex vivo intact organ experiments are hampered by a rapid loss of cell viability, which significantly limits the duration of any experiments (several minutes to a few hours). Nevertheless, ex vivo organ incubations have been widely used to study sperm migration through the oviduct by video microscopy57–59,103,107,108 and epifluorescence microscopy 61. These techniques are especially applicable to species, like the hamster and the mouse, with a transparent wall to the ampullary region or entire oviduct 107…”

Mammalian egg coat modifications and the block to polyspermy

“…This model was supported by sperm‐binding studies performed using a bacterially‐expressed and refolded N‐terminal fragment of human ZP2 (Tsubamoto et al, 1999), whereas experiments with recombinant ZP proteins refolded from inclusion bodies of baculovirus‐infected insect cells suggested that, in human, acrosome‐intact sperm binds to ZP1, ZP3, and ZP4 (Chakravarty, Suraj, & Gupta, 2005; Ganguly et al, 2010). More recently, it was shown that the acrosome reaction can already occur before direct ZP contact in the upper isthmus of the oviduct (Jin et al, 2011; la Spina et al, 2016); moreover, it was reported that mice lacking the putative C‐terminal O ‐glycosylation sites of ZP3 are fertile (Gahlay et al, 2010). Together with experiments using recombinant ZP2 expressed in insect cells as well as oocytes with hybrid egg coats containing different combinations of mouse and human ZP proteins (Avella, Baibakov, & Dean, 2014; Baibakov, Boggs, Yauger, Baibakov, & Dean, 2012), this led to the suggestion that sperm of both species bind to the N‐terminus of ZP2 independent of N ‐ and O ‐glycosylation (Tokuhiro & Dean, 2018).…”