Abstract: The object of this study was to investigate, the effect of PGF_2α administrated on 5 and 7 d following GnRH treatment on estrus and pregnancy during the breeding season in awassi ewes. A total of 60 ewes 3-5 years old were used in this experiment. The ewes were divided into 3 groups (20 ewes/group) in breeding season. First group (control) was treated with the PG-10 day protocol consisting of 2 intramuscular (i.m.) doses of PGF_2α analogue (250 μg of D-Cloprostenol/dose). The second group received intramuscular injection of GnRH analogue (busereline) and 5 day later an i.m. injection of PGF_2α. In the 3 groups were treated with GnRH followed, 7 days later, by an i.m. injection of 250 μg of prostaglandin F_2α. After the detection of estrus, the ewes were inseminated with 0.2 mL fresh sperm. Pregnancy was determined using ultrasonography on day 30 after the AI. Estrus response was 90% in Group 1, 90% in Group 2 and 35% in Group 3 (p<0.001). The pregnancy rate was 77.8% in Group 1, 83.3% in Group 2 and 42.8% in Group 3 (p<0.05). Lambing rate was 77.8% in Group 1, 83.3% in Group 2 and 42.8% in Group 3 (p<0.05). These results suggest that effect of induction estrus and pregnancy and lambing rate after GnRH and 5 days later an i.m. injection of prostaglandin F_2α were higher than GnRH and PGF_2α treatment followed 7 days injection of PGF_2α. But comparable to PGF_2α treatment with 10 days intervals.

INTRODUCTION

Estrus synchronization can be an effective means of increasing the proportion of ewes that become pregnant over a short time, resulting in more uniform lamb crops. Precise control of estrus cycles requires the manipulation of both follicular growth and luteal life Span (Husein and Kridli, 2003). The most frequently used estrus synchronization protocols, based on the use of equine chorionic gonadotropin after a pretreatment with progesterone or progestagen intravaginal devices (Evans and Maxwell, 1990; Menchaca et al., 2004).

Prostaglandin F_2α a great many of strategies have been developed in order to control ovarium function in sheep. Estrus synchronization can be controlled by manipulating either follicular development or Corpus Luteum (CL) (Husein and Kridli, 2003). Prostaglandin F_2α (PGF_2α) can be applied to the ewe to provide estrus synchronization (Gordon, 1997; Husein and Kridli, 2003). PGF_2α in sheep causes CL regression (Rubianes et al., 2003). PGF_2α applications are ineffective since CL does not respond to it between 0-2 days of sexual cycle. Likewise, since CL is luteolized spontaneously between 14-17th days, PGF_2α maintains ineffective. For this reason, it is applied in the form of double luteolitic dose with 9 days intervals in order to synchronize estrus and ovulation (Gordon, 1997). In fixed Timed Artificial Insemination (TAI) and natural breeding, the pregnancy rate in PGF_2α applications, for synchronizing estrus, are found low (Boland et al., 1978). Apart from this, in double dose PGF_2α applications applied with 10-14 days intervals, fixed timed artificial insemination applications are not preferred on big herds as a useful method (Evans and Maxwell, 1990; Menchaca et al., 2004). Estrus and ovulation formed following PGF_2α application can change depending on the existence of follicle developed in ovarium during the application of exogenous hormone (Husein and Kridli, 2003). Alternative applications have been developed in order to manipulate follicular development and provide estrus synchronization. One of these is PGF_2α treatment made after 5 days following GnRH administration on the sheep (Beck et al., 1996). Administering exogenous GnRH in any phase of sexual cycle causes the increase of LH that causes ovulation or atresia and a new follicular wave is occured in 2-3 days following this application (Webb et al., 1992; Wolfenson et al., 1994).

During the ovulatory cycle in sheep, follicles having 2-3 mm diameters appear and reach to the maximum 4-12 mm (Bartlewski et al., 1999; Vinoles et al., 2001). It has been reported that during the appearance of a new follicular wave, many small follicles do not gather and small follicles are not pressured during a development phase of a follicular wave. Moreover, it is suggested that a strong follicular dominance does not exist in the follicular wave appearing once in every 3-5 days (Duggavathi et al., 2003).

PGF_2α treated after 5 days following, the GnRH application has the ability to luteolize CL formation (Beck et al., 1996). Our hypothesis is that PGF_2α done on day 7 following the tretment of GnRH will be effective in increasing reproductive efficiency by prolonging the luteal function.

The objective of the experiment was to examine whether GnRH + PGF_2α (5 or 7 day later) method gives a sufficient on the fertility, compared with standard the method in which double doses of PGF_2α are applied in breeding season.

MATERIALS AND METHODS

Experimental animals and design: The study was performed Harran University sheep breeding unit, Sanliurfa, Turkey (37°07'N, 38°49'E). The material of the study consists of 60 Awassi ewes from different ages (2-5 aged) in breeding season. The same feeding conditions were applied to all animals. The sheep were kept in half-open folds during the study.

Animals were divided into 3 equal groups. The 1st group was given twice, at 10 day interval, i.m. of injections 250 μg of cloprostenol an analogue of PGF_2α (PGF_2α, dalmazine^®, intervet, Turkey). The second group received intramusculary (i.m) 10 μg of buserelin, an analogue of gonadotropine releasing hormone (receptal^®, intervet, Turkey) and 5 days later i.m injection of 250 μg of D-Cloprostenol. The 3rd group was received intramusculary i.m, 10 μg of buserelin and 7 days later i.m injection of 250 μg of D-Cloprostenol.

Determination of the estrus and insemination: The estrus cycle of each ewe was checked twice daily, using teaser rams, with a time interval of about 12 h with a 3 day period after second PGF_2α application. After the detection of estrus, the ewes were inseminated once during estrus and performance of artificial insemination was recommended between 12 and 24 h after detection of standing estrus. Pregnant ewes were determined using the realtime B-mode ultrasound (Scanner LC 100 Vet, Pie Medical, Netherlands) with the 6-8 MHz linear-array transrectal probe on day 30 following artificial insemination.

Parameters and statistical analyses: The reproductive parameters below were calculated in each study group:

The results were analyzed by χ² test. Statistical analyses were performed using SAS (1989).

RESULTS AND DISCUSSION

In estrus synchronization performed with double dose PGF_2α, only CL causes regression and has effect on follicular development. The first GnRH injection initiates a new follicular development and forms a more homogenous follicular wave. GnRH and PGF_2α applied following GnRH synchronize luteal function (Beck et al., 1996). In this study, GnRH + PGF_2α applications were performed in Awassi ewes.

Estrus response, as seen in Table 1; were found as 90% in Group 1 (control) and in Group 2. The Estrus rates in Group 1 and 2 were found considerably higher than Group 3 (p<0.001). These findings are similar to the results reported by Beck et al. (1996), which PGF_2α applied after 5 days following GnRH provides high level estrus synchronization. In addition, the present study confirms that GnRH with PGF_2α administration 7 days later is able to synchronize estrus in ewes and showed that this treatment is less effective for estrus synchronization than other estrus synchronization protocol studied. Thus, it was assumed that CL was sensitive to PGF_2α on day 5 day after GnRH administration. Beck et al. (1996) reported high levels of oestrous synchronization and fertility using a treatment of GnRH followed by PGF_2α 5 days later during the breeding season. However, it was seen that this sensitivity was considerably decreased in potential CL on day 7 day on ovarium.

Table 1:	The effect of different synchronization method in Awassi ewes on estrous response, pregnancy rate and lambing rate
a- dMeans in the same row with different superscripts differ significantly

When exogenous GnRH is administered to the ewes in any phase of estrus cycle, it causes LH secretion. Thus, it could initiate a new follicular wave causing ovulation or atresia (Webb et al., 1992; Wolfenson et al., 1994). The luteal tissue formed after GnRH injection can perform PGF_2α luteolizing performed 5 days later (Beck et al., 1996; Husein and Kridli, 2003). GnRH + PGF_2α protocol (7 days), the fact that development phase of either luteal function or dominant follicle in luteolizing time could decrease estrus synchronization rate. The extending luteal function might change the estrogen/progesterone rate in follicles (Bartlewski et al., 2000).

Pregnancy rate was found as 77.8% in Group 1 and 83.3% in Group 2. The pregnancy rate in Group 1 and 2 were found considerably higher than Group 3 (35.72%) (p<0.05). The pregnancy rate obtained from the inseminations that were performed after synchronized estrus was found similar in double dose PGF_2α and GnRH + PGF_2α (5 days) (Table 1). Beck et al. (1996) observed a high pregnancy rate in cyclic ewes treated PGF_2α after 5 days following the GnRH. When we compare GnRH + PGF_2α (7 days) group with the other 2 groups with regard to the pregnancy rate, the results are considerably low. This case is arised from the animals in GnRH + PGF_2α (7 days) group show low estrus rate than the other groups. So, it can be said that PGF_2α, applied on day 7 following GnRH application, causes increase in fertility. When each group is compared with regard to the pregnancy rate, no considerable difference was found among the groups (Table 1). It was suggested that the extension of the lifespan of the CL in the ewes caused increase in fertility (Evans et al., 2001). The fact that either pregnancy rate or breeding rate will be effective in increasing reproductive efficiency of PGF_2α performed on day 7 following GnRH application supports our hypothesis.

Lambing rate was found as 77.8% in Group 1, 83.3% in Group 2 and 35.72% in Group 3. The lambing rate in Group 2 were found significantly higher than Group 3 (p<0.05) depending on low estrus rate in Group 3.

CONCLUSION

These results suggest that effect of induction estrus and pregnancy and lambing rate after GnRH and 5 days later an i.m. injection of prostaglandin F_2α were higher than GnRH and PGF_2α treatment followed 7 days injection of PGF_2α.