In vitro assays for vesper mice (Calomys laucha) sperm using heterologous substrates from nonrodent species

Abstract

Small vesper mice (Calomys laucha) may be considered as an animal model for in vitro fertilization studies, but limited data about in vitro evaluations of their sperm quality and fertility are available. The in vitro penetration (IVP) assay is used to estimate potential sperm fertility for many mammal species, but it still requires reduction in cost and labor. This study tested improvements in the IVP assay for C. laucha sperm using swine oocytes and perivitelline layers (PVL) of chicken eggs as substrates, and evaluated associations among C. laucha sperm quality, IVP, and in vivo fertility after natural mating. In the IVP assay, gametes coincubation was carried out flat-bottomed wells with M2, in water bath at 37°C for 2 hr. C. laucha sperm presented motility, normal morphology, membrane integrity, and acrosome integrity equal to 90.6 ± 5.6, 90.2 ± 6.6, 88.7 ± 9.6, and 90.5 ± 11.5%, respectively. The IVP rate was 39.8% in swine oocytes and 87.5% in the inner PVL. Considering in vivo fertility as the gold standard, the IVP assay in swine oocytes presented a sensitivity of 16.0% and specificity of 83.3%. The sensitivity of the IVP assay in the inner PVL was 84.0%, but the specificity was not determined because there were no true negative results. Sperm membrane integrity was correlated with parturition after natural mating (r = 0.38, P<0.01) and litter size (r = 0.54; P<0.0002).The IVP assay using swine oocytes as substrates can be performed in nearly 2 hr without gametes' coincubation in CO(2).

Sperm impairments in adult vesper mice (Calomys laucha) caused byin uteroexposure to bisphenol A

“…However, these abnormalities may not impair C. laucha reproduction, since normal sperm morphology of this species has been reported by Corcini et al . () to be equal to 90.2 ± 6.6%, which is lower than the BPA200 group in this study. Sperm motility has also been shown to decrease in rats perinatally exposed to BPA (Salian et al ., ), and lower motility related to urinary BPA in a dose–response manner in men has been reported (Li et al ., ).…”

“…All the parameters discussed above may influence the IVP test. The IVP test presents high specificity, detecting sub‐fertile males unable to sire a litter after natural mating by detecting those failing to penetrate swine oocytes in vitro (Corcini et al ., ). Since C. laucha sperm head morphology is similar to that of other mammals (Lasserre et al ., ), it has been shown that porcine oocytes are useful heterologous substrates for in vitro fertilisation studies of this species (Lasserre et al ., ; Corcini et al ., ).…”

“…Our results for the water group were similar to those found by Corcini et al . (), who studied in vitro assays for vesper mice sperm and reported normal membrane integrity as 88.7 ± 9.6% for C. laucha .…”

“…Oocytes were collected and processed; thirty oocytes were used for each sample and the test followed the protocol described by Corcini et al . (). Briefly, a M2 medium containing HEPES, 0.4% bovine serum albumin (A3311) and approximately 2 × 10 6 spermatozoa per ml was used.…”

“…Spermatozoa were collected after approaching the reproductive system by laparotomy. The cauda of both epididymes and part of the vas deferens were removed, ruptured with a 12 g × 40 mm needle and placed in a 35 mm Petri dish containing 200 μl of M2 medium with HEPES (M7167) (Corcini et al ., ) for sperm dilution. Sperm quality evaluations were done as described below after incubation of samples for 10 min at 37 °C in M2 medium.…”