The main objective of this investigation was to compare conception rate between Ovsynch and a modest presynchronization strategy consisting of simultaneous administration of PGF2α and GnRH 7 days before Ovsynch (Pre-OV) in indigenous Hariana breed of zebu cow. High producing dairy cow subjected to presynchronization of estrous cycle before timed-AI resulted with increased fertility (Moreira et al. 2001). Additionally, the researchers have reported increased fertility response subsequent to presynchronization of estrous cycle compared to Ovsynch alone (Navanukraw et al. 2004; Galvão et al. 2007). In an experiment on dairy cows, proportion of cows found pregnant were tended to be higher (P < 0.08) following presynchronization with G6G protocol (PGF2α followed 2 day later by GnRH, administered 6 days prior to start of Ovsynch protocol), in comparison with Ovsynch alone (50% vs 27%, Bello et al. 2006). Herlihy et al. (2012) observed that presynchronization either with double Ovsynch or double PG (14 days apart) followed by Ovsynch resulted in 46.3% and 38.2% conception rate in lactating dairy cows, respectively. Similarly, in present study, presynchronization with simultaneous administration of PGF2α and GnRH 7 days before Ovsynch resulted in better pregnancy outcomes than Ovsynch alone (Control). Additionally, previous studies based on similar presynchronization strategy, have reported comparable pregnancy outcomes as obtained with other laborious and logistically challenging presynchronization strategies like G6G and Presynch-10 (Yousaf et al. 2016; Martins et al. 2017). Moreover, FSCR obtained in presynchronized cows during present study was similar to previous reports. Contrary, Hubner et al. (2020) reported lower proportion of Holstein dairy cows pregnant (33.0%) subsequent to presynchronization with PGF2α and GnRH 7 days before Ovsynch. The discrepancies in the current and previous result could be attributed to involvement of post-partum cows in early stage (48 days post-partum) than current study. Thus, results of this study clearly show that presynchronization with simultaneous administration of GnRH and PGF2α 7 days before Ovsynch improved the FSCR than Ovsynch alone (control), and underlying reasons could be: i) greater ovulation rate in response to 1st GnRH of Ovsynch, ii) greater number of CL and higher plasma progesterone present on day 7 of Ovsynch, iii) improved synchronization rate, iv) greater LPF size on day of AI. Thus, above findings support our hypothesis that Pre-OV treatment resulted in improved pregnancy rate as compared to control.
In present study, Pre-OV treated cows had smaller size of CL at start of Ovsynch as compared to Ovsynch initiated at random stage of estrous cycle in control. Also, on the day of first GnRH of Ovsynch, a tendency towards higher variation in plasma P4 conc. (σ2 = 0.29 vs 0.08; P < 0.1; control vs Pre-OV) and size of LF (σ2 = 12.7 vs 3.2; P < 0.08 control vs Pre-OV) was observed in control, which demarks randomness among control cows. Above findings implies that Pre-OV treatment had led to initiation of a new estrous cycle in cows and support our hypothesis that Pre-OV treatment would synchronize cows to 6-7th day of estrous cycle at the start of Ovsynch, while majority of cows might be in mid-cycle stage at initiation of treatment in controls. Indeed, greater CL size present on day 0 of Ovsynch in control than Pre-OV also indicates that the cows in control should have been at mid-cycle stage. In response to GnRH, ovulation of LF is size dependent (Sartori et al. 2001). The acquisition of ovulatory capacity of LF develops as it reaches > 8mm size in Nellore breed of zebu cattle (Barros et al. 2008). In present study number of cows with LF of size < 8mm were higher (8 cows vs 2 cows; P < 0.05) in control as compared with Pre-OV on day of initiation of Ovsynch. Indeed, presence of larger sized LF present at the time of 1st GnRH in Pre-OV compared with control resulted in higher ovulation rate in response to first GnRH of Ovsynch in Pre-OV than control. This observation forced us to speculate that Pre-OV treatment strategy might have increased the number of cows at 6-7th day of estrous cycle at start of Ovsynch which resulted in greater FSCR. Further, this high ovulatory response to first GnRH in Pre-OV and presence of younger functional CL at first GnRH of Ovsynch in a greater number of cows from presynchronized than control is the possible explanation of the higher FSCR in Pre-OV. Previous study has also shown that P/AI was higher when a young functional CL was present at first GnRH of Ovsynch (Carvalho et al. 2018). In response to first GnRH, ovulation rate observed in present study was similar to (68%) reported by Yousaf et al. (2016) using similar presynchronization strategy in Holstein Friesian cows. On contrary, Hubner et al. (2020) reported higher (90%) ovulation rate subsequent to presynchronization with PGF2α and GnRH 7 days before Ovsynch in lactating Holstein dairy cows. Moreover, such disparities in ovulation rate were previously reported for various presynchronization strategy like G6G (67–85%, Yousaf et al. 2016; Bello et al. 2006) and Double-Ovsynch (37.2–90.3%, Giordano et al. 2012b; Ozturk et al. 2010). Various factors like environment, nutrition and genetics of treated cows could account for such discrepancies (Hubner et al. 2020). While, ovulatory response to first GnRH in control cows of present study was similar to previous reports involving lactating dairy cows (Giordano et al. 2012b; Lopes et al. 2013). Giordano et al. (2012a) reported that progesterone levels at the time of GnRH administration suppress the LH surge which hampers the ovulation. Further suggested that, despite the capacity of follicles to ovulate, the reduced ovulation in response to GnRH, is associated with alterations in mechanism that triggers the ovulation process.
Although, mean CL area at the time of PGF2α of Ovsynch was smaller in Pre-OV compared to control but due to greater number of CLs observed in Pre-OV resulted into higher plasma P4 in Pre-OV cows. As various studies have indicated that ovulation to first GnRH leads to formation of accessory CLs and resultant high P4, thus high ovulatory response to first GnRH can implied as reason for more CLs (Vasconcelos et al. 1999; Bello et al. 2006). Carvalho et al. (2018) speculated that cows experienced with elevated P4 at the time of PGF2α, could have greater probability to become pregnant as also mentioned in previous reports (Bello et al. 2006; Wiltbank and Pursely 2014; Herlihy et al. 2012). Higher plasma P4 in Pre-OV treatment could also be attributed to reduced chances of spontaneous luteolysis before PGF2α of Ovsynch in Pre-OV treated cows; as in the present study, PGF2α of presynchronization was intended to cause luteolysis of all mid and late-cycle CL, and a functional CL formed after ovulation to either GnRH of presynchronization or first GnRH of Ovsynch would persist until administration of PGF2α of Ovsynch.
In present study, high P4 conc. at time of PGF2α of Ovsynch was identified as possible predictor of pregnancy at 45 days post-AI. During the development period of LF, elevated circulating P4 conc. might have decreased LH pulsatility, likely leads to enhancing competency of LF which consequently, improved quality of the LPF (Mihm et al. 1994; Revah and Butler 1996), similar observations have been noticed in present study; however, LH conc. was not measured. Ovulatory response to first GnRH influences luteolysis at PGF2α of Ovsynch and synchronization rate with increased frequency of complete luteolysis in cows ovulated to first GnRH (Bello et al. 2006), and similar findings were observed in the present study as higher proportion of cows had complete luteolysis in Pre-OV. The luteolysis rate in present study (86.8%) was lower than reported by Yousaf et al. (2016) (97%) after using similar Pre-OV strategy in dairy cows. The higher luteolysis rate reported by Yousaf et al. (2016) might be owing to administration of an extra PGF2α injection on day 8 of Ovsynch. In present study, luteolytic response in control animals was less in comparison to previous reports 83.9% (Giordano et al. 2012b) and 83% (Carvalho et al. 2015). The exact reason for this discrepancy is not known, however, this difference could be attributed to the stage of estrous cycle at starting of Ovsynch and physiological differences between cows. Average ovulatory follicle diameter at 2nd GnRH of Ovsynch and at the time of AI was higher and found lesser variable in Pre-OV as compared to control. Less variability in ovulatory follicle size at 2nd GnRH of Ovsynch in Pre-OV was attributed to higher ovulatory response to first GnRH in this group, which increased the circulating P4 conc. thus, allowing the development of the LF less variable and closer to the ideal size at the time of the second GnRH (Bello et al. 2006). Circulating conc. of P4 at 2nd GnRH is identified as probable marker of fertility with decreasing P4 resulting in increased likelihood of pregnancy at 45 days post-AI. This can be attributed to the fact that lower P4 at the time of 2nd GnRH resulted in elevated expression of estrus and improves uterine environment for successful conception (Vasconcelos et al. 2013). The present study also confirmed the improved FSCR in cows that exhibited estrus than those did not.
In this study, plasma E2 conc. on day of AI was found to be higher in Pre-OV cows as compared to control. Additionally, higher plasma E2 conc. on day of AI was recorded in pregnant cows as compared to their non-pregnant counterparts in both treatments (Pre-OV and Control). Thus, high E2 conc. in the present study was related to improve FSCR and the results were in accordance with the observations reported in earlier studies on dairy cows (Lopes et al. 2010; Vasconcelos et al. 2013) and buffalo (Pandey et al. 2018). It is well established fact that optimal E2 conc. on day of estrus is a key for proper growth and coordination in important events like triggering LH surge, ovulation and resumption of meiosis (Greenwald and Roy 1994). Further, increased E2 conc. on day of final GnRH of Ovsynch was also related to increasing probability of pregnancy at day 35 post-AI in cows receiving timed-AI (Bello et al. 2006). It seems logical that the higher plasma E2 on the day of AI would have led to the LH surge, consequently timely ovulation that might have resulted in increased conception rate.
In this study, LPF size on day of AI also affected luteal dynamics and function subsequent to ovulation on day 12 post-AI and a positive correlation was observed The previous studies including dairy cows (Pursely and Martins 2011) and buffalo (Pandey et al. 2018) also showed constructive relationship between LPF diameter and luteal profiles. Many studies observed that P4 conc. affected the pregnancy status as increased plasma P4 concentration was conducive for establishment of pregnancy in cows (Grimard et al. 2006; Pursely and Martins 2011). It is noteworthy that, the pregnant cows had larger CL and higher plasma progesterone on day 12 post-AI compared to non-pregnant cows of respective treatment. Retrospective analysis of data obtained from both treatments (Pre-OV and control) indicated that ovulation in response to first GnRH of Ovsynch affects the outcome for each injection of Ovsynch; resultant luteal profiles and conception rate, which is a key determinant in success of FTAI programme based on Ovsynch strategy (Bello et al. 2006).
Based on this study, it can be concluded that simpler presynchronization strategy involving simultaneous administration of GnRH and PGF2α 7 days before Ovsynch (Pre-OV) increased the ovulatory response to first GnRH of Ovsynch, increased CL area day 12 post-AI and resulted with higher conception rate. Plasma P4 conc. at PGF2α and at 2nd GnRH of Ovsynch was identified as significant predictor of pregnancy outcome at 45 days PAI. Further, it was determined that LPF size on day of AI influences luteal profiles and pregnancy outcomes post-AI