Effects of melatonin treatment on follicular development and oocyte quality in Chios ewes - short communication.

Follicular development and oocyte quality were assessed by laparoscopic observation and in vitro fertilisation, respectively, in melatonin-treated (Group M) and control (Group C) anoestrous Chios ewes (n = 10 in each group). Fourteen days after melatonin insertion, all ewes had laparoscopic evaluation of the follicular population followed by oocyte pick-up (OPU); on day 22 intravaginal progestagen sponges were inserted for 14 days. Two days after sponge removal the follicular population was re-evaluated and a second follicular aspiration was performed. Collected oocytes from the second OPU underwent in vitro maturation, fertilisation and culture. The number of large follicles was higher in Group M than in the control ewes during the first OPU and tended to be so (P = 0.06) at the second. Morphologically, oocytes collected from controls were of better quality than those from Group M; however, more oocytes collected from melatonintreated animals fertilised and developed in vitro . These results indicate that melatonin is a potent regulator of follicular development and oocyte competence during the anoestrous period of the ewe.

Effect of plasmin, plasminogen activators and a plasmin inhibitor on bovine in vitro embryo production.

In the present study, four experiments were conducted to investigate the possible effects of plasminogen activators (urokinase-type plasminogen activator (u-PA) and tissue-type plasminogen activator (t-PA)), plasmin, and a plasmin inhibitor (epsilon-aminocaproic acid (epsilon-ACA)) on different stages of bovine in vitro embryo production (IVP). The concentrations of these modifiers in IVP media were conditioned according to the plasminogen activator activity of bovine preovulatory follicular fluid. Media were modified in a single phase of IVP with an 18 h or 24 h incubation for in vitro maturation (IVM) and a 24 h or 48 h incubation for the IVF or in vitro culture (IVC), respectively. After IVM the oocytes were either fixed and stained or underwent IVF and IVC. The main findings were: (1) plasmin added to the 18 h IVM medium increased maturation rate without affecting fertilisation or embryo development rates; (2) t-PA added to the IVF medium significantly increased cleavage; (3) u-PA added to the IVC medium significantly increased embryo development rates; (4) the efficiency of all phases of IVP was reduced after the addition of epsilon-ACA; and (5) plasminogen addition had no effect in any IVP phase tested. We conclude that the members of the plasminogen activator-plasmin system contribute in different ways to bovine IVM, IVF and IVC.

Activity of glycosidases (beta-N-acetyloglucosaminidase, alpha-mannosidase, and beta-galactosidase) in the uterine luminal fluid of cows after multiple ovulation.

This study investigated the activity of beta-N-acetyloglucosaminidase (beta-NAGASE), alpha-mannosidase, and beta-galactosidase in the uterine luminal fluid of cows after superovulation treatment, along with the possible associations between the activity of these 3 glycosidases and the superovulatory response. Embryos and a sample of fluid flushed from each uterine horn were collected on day 7 after artificial insemination (on estrus day 0) from 32 cows in which superovulation was induced with porcine follicle-stimulating hormone. Glycosidase activity was assayed colorimetrically. The cows were classified as to superovulatory response according to the number of corpora lutea per ovary (group 1, 1 to 4; group 2, > 4) and according to the total number of embryos per horn (T1, 0; T2, 1 to 2; T3, 3 to 4; T4, > 4) and the number of transferable embryos per horn (TR1, 0; TR2, 1 to 2; TR3, 3 to 4; TR4, > 4). The mean activity of beta-NAGASE was significantly lower (P < 0.05) in group 2 than in group 1, at 95.99 (standard error 20.43) versus 226.72 (46.77) IU/L. It was also significantly lower (P < 0.01) in group T4 compared with groups T1, T2, and T3, at 50.09 (8.21) versus 129.25 (34.60), 222.27 (62.62), and 290.26 (93.77) IU/L, respectively, as well as in group T1 compared with group T3. There was a positive relationship between beta-NAGASE activity and both the total number of embryos (P = 0.047) and the number of transferable embryos per horn (P = 0.013) when 1 to 4 corpora lutea developed per ipsilateral ovary. No difference in alpha-mannosidase or beta-galactosidase activity was detected among the groups.

Plasminogen activator activity in cortical granules of bovine oocytes during in vitro maturation.

In this study, we provide evidence that plasminogen activator of tissue-type (t-PA), at least, is present in extracts of bovine oocyte cortical granules, and that its activity varies significantly with the duration of oocyte in vitro maturation. Cortical granules were collected from bovine oocytes by means of micromanipulation, after 0, 12, or 24 h of IVM. Our results show that plasminogen activator activity of cortical granule extracts was significantly higher after 24 h of IVM than after 12 h of IVM or before IVM. This activity was apparently due, at least partly, to tissue-type plasminogen activator as shown immunologically. No evidence was found for the presence of urokinase-type plasminogen activator, plasminogen activator inhibitors or plasmin inhibitors in bovine oocyte cortical granule extracts. Our findings further support the hypothesis that t-PA activity of oocyte origin may have a role in oocyte maturation or fertilization, as well as in post-fertilization events, such as cortical reaction and formation of the zona block to polyspermy.