Effect of progesterone on capacitation of buffalo (Bubalus bubalis) spermatozoa in vitro.

The aim of the present study was to evaluate whether progesterone (P4) promotes sperm capacitation in buffalo. Frozen-thawed sperm were treated by swim-up to select only the motile population. Spermatozoa (N=1740, equally distributed into 6 groups) were incubated in the presence of: A) 0.1 µg of P4; B) 0.01 mM heparin and C) in medium lacking capacitating agents (control) for 1 and 2 hours. Sperm were exposed for 15 min to 60 g mL-1 of lysophosphatidylcholine, known to induce acrosome reaction only on capacitated spermatozoa. Spermatozoa were fixed in 37 % formaldheyde and double stained with Trypan blue and Giemsa, for acrosome evaluation by microscopic examination. The proportion of acrosome-reacted (AR) spermatozoa in each group was used to assess the efficiency of capacitation. Differences between groups were analyzed by Chi Square. The 1-hour treatment with both P4 and heparin significantly (P<0.01) increased the percentages of AR spermatozoa compared to the control (35.7 %, 23.2 % and 13.2 % respectively). Sperm incubation with P4 for 1 h resulted in significantly (P<0.01) higher incidence of AR compared to that with heparin, that is the capacitating agent currently used for in vitro fertilization (IVF). However, following 2 hours of incubation, both P4 and heparin gave similar percentages of AR sperm (35.6 vs 34.5 % respectively) that were significantly (P<0.01) higher than the control (13.3 %). It was demonstrated that P4 induces buffalo sperm capacitation in vitro and may be considered as an alternative capacitating agent for buffalo IVF.