The Effect of Oestradiol, Progesterone and Heparin on Bovine Spermatozoa Function After Thawing

Abstract

The present experiment was designed to determine the effects of various biologically active substances, such as oestradiol (OE), progesterone (P4) and heparin (Hep) alone or in combination on sperm plasma membrane scrambling, capacitation and acrosome reaction (AR) of post-thaw bovine spermatozoa. Spermatozoa were incubated for 180 min in capacitation medium supplemented with (i) 1 mug/ml OE; (ii) 1 mug/ml P4; (iii) 1 mug/ml OE and 1 mug/ml P4; (iv) 1 mug/ml OE and 5 mug/ml Hep; (v) 1 mug/ml P4 and 5 mug/ml Hep; (vi) 1 mug/ml OE, 1 mug/ml P4 and 5 mug/ml Hep. At predetermined time intervals aliquots were taken to assess sperm plasma membrane scrambling, or capacitation (AR induced by lysophosphatidylcholine) in spermatozoa. The second experiment was aimed to study the effects of OE, P4 and OE/P4 as potential inducers of AR in Hep-capacitated spermatozoa. Plasma membrane scrambling was assessed by a flow cytometer, using Merocyanine staining. Acrosomal status and viability of spermatozoa were evaluated under epifluorescence microscope with Ethidium homodimer-1/peanut agglutinin fluorescein isothiocyanate staining method (EthD-1/PNA-FITC). The results show that OE, P4 and a combination of OE/P4 at concentrations used did not affect sperm viability. Heparin significantly (p < 0.001) increased sperm plasma membrane scrambling of OE and P4-treated spermatozoa. P4 significantly affected the rate of sperm capacitation (p < 0.001) and AR (p < 0.05), but OE expressed membrane-stabilizing properties (p < 0.05). It can be concluded that in frozen-thawed bovine spermatozoa OE presents plasma membrane stabilizing properties that can be abolished by Hep, but not by P4. Progesterone possesses capacitating and AR-inducing properties in frozen-thawed bovine spermatozoa that can be alleviated by OE.

Effect of different media additives on capacitation of frozen–thawed ram spermatozoa as a potential replacement for estrous sheep serum

“…Moreover, when added to bovine capacitation medium, heparin supported downstream time-dependent increases in protein tyrosine phosphorylation [15]. Similarly, hypotaurine, a precursor of taurine, added in combination with heparin further supported sperm motility and viability [10,[21][22][23] and acted as an oxygen radical scavenger [24].…”

“…In this regard, addition of P4 to the capacitation medium exerted a positive effect on sperm membrane cholesterol efflux, hyperactivation, and the acrosome reaction [9][10][11][12]. However, the effect of estrogens on sperm capacitation has been controversial [7,10,13].…”

“…For instance, several hormones present in uterine and oviductal fluid at the time of in vivo fertilization and those controlling the estrous cycle, such as progesterone (P4) or estrogens, may be good candidates for triggering in vitro sperm capacitation [7,8]. In this regard, addition of P4 to the capacitation medium exerted a positive effect on sperm membrane cholesterol efflux, hyperactivation, and the acrosome reaction [9][10][11][12]. However, the effect of estrogens on sperm capacitation has been controversial [7,10,13].…”

The effect of oestradiol-17β on the motility, viability and the acrosomal status of bull sperm

“…Higher doses of E 2 are detrimental for sperm survival. A study was conducted by Lukoseviciute et al (2005) to determine the effect of 1 μg E 2 /mL on sperm plasma membrane scrambling, capacitation and acrosome reaction of post-thaw bovine sperm. According to the result of their study, the addition of 1 μg E 2 /mL did not affect sperm viability.…”

“…The most potent oestradiol is oestradiol-17β (E 2 ) which is synthesized through the aromatization of testosterone by the enzyme P 450 aromatase in the testis (Rommerts et al, 1982;Tsai-Morria et al, 1985). It plays an important role in reproduction by regulating the function of the efferent ductules and the epididymis where sperm mature and gain fertilizing ability (Hess et al, 1997;Carreau et al, 2003;Hess & Carnes, 2004;Lukoseviciute et al, 2005).…”