Progesterone accelerates spermatozoa penetration time in homologous in vitro fertilization in the pig

Theriogenology

PROGESTERONE ACCELERATES SPERMATOZOA PENETRATION TIME IN HOMOLOGOUS IN VITRO FERTILIZATION IN THE PIG J.M. Vazquez, E.A. Martinez, C. Matas and J. Rota Department of Animal Pathology, Murcia University, Murcia, 30.071. Spain We have previously shown that progesterone may contribute to inducing the acrosome reaction and may prolong the viability of boar spermatozoa, suggesting a possible role during fertilization. However, this steroid does not increase overall sperm penetration rates of matured pig oocytes after 16-18 h incubation. In this study we investigated if the penetration time of unwashed and non-preincubated boar spermatozoa in in vitro matured pig oocytes is accelerated by the presence of progesterone in the culture medium. Spermatozoa (5~10~ cells/ml) were incubated in 2 ml of a control medium (M: TCM199 modified as described by Mattioli et al., Gamete Res., 20:177. 1988, supplemented with 2mM caffeine), MD: M+DMSO (0.16%; solvent for progesterone), or MP: MD + progesterone (15uM) with groups of 15 oocytes matured in vitro (Yoshida et al., Mol. Reprod. Dev., 31:68. 1992) at 39’C under 5 % CO2 in air. At 3, 6, 9 and 18 h after the begining of incubation, the oocytes were fmed for a minimum of 24 h with ethanol:acetic (3: 1 v/v), stained with 1% lacmoid and assessed for sperm penetration. The data from six replicates were analyzed by ANOVA and comparisons between means were made by Tukey-test. Results are summarized in the table below. Compared to the controls (M and MD), the percentages of in vitro oocytes penetrated were signiticantly greater in the presence of progesterone at 6 and 9 h incubation. The analysis of the data suggests that progesterone accelerates the time of sperm penetration, but does not increase polyspermia, in in vitro matured pig oocytes. Table 1. Effect of progesterone on sperm penetration rates at different lengths of exposure. Sperm Penetration Length of Medium Mature Spedoocyte exposure (n + SEM) (%k SEM) oocytes (n) 1.1+0.1 M 2.3OkO.7 85 MD 60 1.95+0.5 MP 1.1kO.l 92 3.17kO.4 1.2kO.2 M 191 23. 15f2.1a 6h

9h

MD

161

25.13f1.2a

1.2kO.3

MP

178

41.23k4.2b

1.4kO.4

M

180

55.27*2.ga

2.1kO.5

MD

141

53.15f3.1a

2.3kO.3

3.OkO.4 69.15k6.3b M 89.35k2.7 4.0+0.6 160 4.9f0.7 18h MD 140 91.81+_1.7 5.3kO.7 MP 182 92.85+3.5 Means with different superscripts within columns and groups are different @ < 0.05). MP

189

Funded by CICYT (AGF 95/1009) and EUREKA (EU 1713).