Synchronization rate, size of the ovulatory follicle, and pregnancy rate after synchronization of ovulation beginning on different days of the estrous cycle in lactating dairy cows

Recently a protocol was developed that precisely synchronizes the time of ovulation in lactating dairy cows (Ovsynch; GnRH-7d-PGF2 alpha-2d-GnRH). We evaluated whether initiation of Ovsynch on different days of the estrous cycle altered the effectiveness of this protocol. The percentage of cows (n = 156) ovulating to the first GnRH was 64% and varied (P < 0.01) by stage of estrous cycle. Treatment with PGF2 alpha was effective, with 93% of cows having low progesterone at second GnRH. The overall percentage of cows that ovulated after second GnRH (synchronization rate) was 87% and varied by response to first GnRH (92% if ovulation to first GnRH vs 79% if no ovulation; P < 0.05). There were 6% of cows that ovulated before the second injection of GnRH and 7% with no detectable ovulation by 48 h after second GnRH. Maximal diameter of the ovulatory follicle varied by stage of estrous cycle, with cows in which Ovsynch was initiated at midcycle having the smallest follicles. In addition, milk production and serum progesterone concentration on the day of PGF2 alpha affected (P < 0.05) size of the ovulatory follicle. Using these results we analyzed pregnancy rate at Days 28 and 98 after AI for cows (n = 404) in which Ovsynch was initiated on known days of the estrous cycle. Pregnancy rate was lower for cows expected to ovulate larger follicles than those expected to ovulate smaller follicles (P < 0.05; 32 vs 42%). Thus, although overall synchronization rate with Ovsynch was above 85%, there were clear differences in response according to day of protocol initiation. Cows in which Ovsynch was initiated near midcycle had smaller ovulatory follicles and greater pregnancy rates.     

Conception rate after fixed time insemination following ovsynch protocol with and without progesterone supplementation in cyclic and non-cyclic Mediterranean Italian buffaloes (Bubalus bubalis)

The aim of this study was to test the effect of progesterone supplementation to Ovsynch protocol in cyclic and non-cyclic Mediterranean Italian buffaloes on conception rate after fixed time artificial insemination. From 169 pluriparous buffaloes, 2 groups were identified and subjected to: (1) Ovsynch protocol (OV; n=83) and (2) Ovsynch protocol with the supplementation of progesterone from days 0 to 7 (OV+PROG.; n=86). All cows were inseminated 16-20 h after the second GnRH administration. Within each group, non-cyclic buffaloes were identified (OV=21 and OV+PROG.=20). Overall conception rate was significantly higher in cyclic compared to non-cyclic buffaloes: 43.7% versus 17.0%, respectively, P=0.001. A significant effect of progesterone supplementation on conception rate was observed in non-cyclic buffaloes (30% versus 4.7%, P=0.04) but not in cyclic buffaloes (51.5% versus 35.7%, P=0.077). Collectively, the presence of a large follicle (>or=10 mm) detected at the beginning of the Ovsynch protocol by ultrasound significantly affected conception rate (44% versus 8%, P=0.01). The findings of the present study suggest that (i) progesterone supplementation to the Ovsynch protocol in buffaloes increases conception rate in non-cyclic animals, (ii) the presence of a large follicle at the beginning of the Ovsynch protocol is a determining factor for a successful synchronization of ovulation and high conception rates and (iii) ultrasound monitoring can improve the overall efficiency by selectively identifying more suitable cycling animals carrying a responsive follicle at the time of first GnRH administration.     

Reproductive performance of dairy cows with ovarian cysts after synchronizing ovulation using GnRH or hCG during the warm or cool period of the year

This study was designed to compare the reproductive response to timed AI of lactating dairy cows with cystic ovarian follicles treated with GnRH or hCG to synchronize ovulation. The effectiveness of treatment during the warm or cool period of the year was also compared. Cows were given 12 microg GnRH-agonist i.m. on day 0 of the protocol, 15 mg PGF(2alpha) i.m. on day 7, and either GnRH-agonist (GPG treatment) or 3000 IU hCG i.m. (GPH treatment) on day 9, followed by timed AI. The cows were randomly chronologically assigned to GPG (n=130) or GPH (n=136) group. All cows were inseminated at fixed time 16-22 h after the end of treatment. During the warm period the pregnancy rate to first AI was 12% (7/60) and 21% (14/68) for the GPG and GPH groups, respectively, there being no significant differences between groups; the cumulative pregnancy rate was 22% (13/60) and 21% (14/68) for the GPG and GPH groups, respectively, again with no significant intergroup differences. During the cool period pregnancy rate to first AI was not different between groups: 29% (20/70) for GPG and 32% (22/68) for GPH, respectively; whereas the cumulative pregnancy rate was significantly higher (P<0.05) for the GPH groups than for the GPG group: 56% (39/70) and 78% (53/68), respectively. These findings indicate that during the warm period, the pregnancy rates of the cystic cows were similar whether they received GPG or GPH treatment, during the cool period, there is a beneficial effect to use hCG at day 9 of the ovsynch protocol compared GnRH on cumulative pregnancy rate.     

Persistent ovarian follicles in dairy cows: a therapeutic approach

Anestrus is common during the postpartum period in high-producing dairy cows. In a previous investigation, we were able to diagnose persistent follicles of 8 to 12 mm in anestrous cows. This report describes 2 consecutive studies. The objectives of the first were to 1) assess the association of persistent follicles with anestrus; and 2) evaluate 2 therapeutic treatments. In the second study, we compared the effectiveness of the best treatment established in Study 1 with the Ovsynch protocol. For Study 1, anestrous cows were considered to have a persistent follicle if it was possible to observe a single follicular structure > 8 mm in the absence of a corpus luteum or a cyst in 2 ultrasonographic examinations performed at an interval of 7 d. At diagnosis (Day 0), cows were assigned to 1 of 3 treatment groups. Cows in Group GnRH/PGF (n=17) were treated with 100 microg GnRH i.m., and 25 mg PGF2alpha i.m. on Day 14. Cows in Group PRID (n=18) were fitted with a progesterone releasing intravaginal device (PRID, containing 1.55 g of progesterone) for 9 d and were given 100 microg GnRH i.m. at the time of PRID insertion, and 25 mg PGF2alpha i.m. on Day 7. Cows in Group Control (n=18) received no treatment. The animals were inseminated at observed estrus and were monitored weekly by ultrasonography until AI or 5 weeks from diagnosis. Blood samples were also collected on a weekly basis for progesterone determination. The mean size of persistent follicles on Day 0 was 9.4 +/- 0.04 mm. Progesterone levels were < 0.2 ng/mL during the first 35 d in 16 of 18 Control cows. Cows in the PRID group showed a lower persistent follicle rate (16.7% < 70.6% < 88.9%; P < 0.0001; PRID vs GnRH/PGF vs Control, respectively); a higher estrus detection rate (83.3% > 29.4% > 11.1%; P < 0.0001) and a higher pregnancy rate (27.8% > 5.9% > 0%; P = 0.02). For the second study, 145 cows with persistent follicles were randomly assigned to 1 of 2 treatment groups: cows in Group Ovsynch (n=73) were treated with 100 microg GnRH i.m. on Day 0, 25 mg PGF2alpha i.m. on Day 7, and 100 microm GnRH i.m. 32 h later. Cows in this group were inseminated 16 to 20 h after the second GnRH dose (Ovsynch protocol). Cows in Group PRID (n=72) were treated as those in the PRID group of Study 1, and were inseminated 56 h after PRID removal. Cows in the PRID group showed a higher ovulation rate (84.8% > 8.2%: P < 0.0001); a higher pregnancy rate (34.2% > 4.1%; P < 0.0001) and lower follicular persistence rate (22.2% < 63%; P < 0.0001) than those in Ovsynch. Our results indicate that persistent follicles affect cyclic ovarian function in lactating dairy cows. Cows with persistent follicles can be successfully synchronized and time inseminated using progesterone, GnRH and PGF2alpha but show a limited response to treatment with GnRH plus PGF2alpha.     

Effect of presynchronization using prostaglandin F2alpha and a milk-ejection test on pregnancy rate after the timed artificial insemination protocol, Ovsynch

The objective of this research was to determine if PGF2alpha-induced milk letdown (ML) is an accurate indicator of luteolysis, allowing cows to be synchronized to begin the Ovsynch protocol (GnRH-7d-PGF2alpha-2d-GnRH-24h-AI) at the most beneficial time of the estrous cycle (days 5-9), and determine if this would improve pregnancy rate (PR). Lactating Holstein cows between 55 and 70 days in milk were used to evaluate the ML test and PR after the Ovsynch protocol, when initiated on the basis of the test result (PROSYNCH). PROSYNCH cows (n = 60) had one teat cannulated to test for ML and were treated with 500 microg cloprostenol, PGF2alpha analogue (PG). Cows with ML were started on Ovsynch 10 days later, and those without started 3 days later. Cows in the control group (OVSYNCH, n = 64) were injected with physiological saline and observed for ML. This group was started on Ovsynch 10 days after saline treatment. Milk samples were collected thrice weekly to determine progesterone concentrations. ML indicated luteolysis with a sensitivity of 98% and a specificity of 60%. The positive and negative predictive values were 83 and 92%, respectively. Pregnancy rates were 48% for PROSYNCH and 52% for OVSYNCH (P = 0.72). When data from both groups were combined, PR was greater in cows that started the Ovsynch protocol in stage 2 of the estrous cycle (days 5-9, 67%) than all other stages (stage 1: days 1-4, 35%; stage 3: days 10-16, 45%; stage 4: days 17-21, 42%; P < 0.01). The proportion of animals with ovulation after GnRH#1, luteolysis after PGF2alpha, and ovulation after GnRH#2 were all greater in the PROSYNCH group (77% versus 55%, P < 0.02; 83% versus 66%, P < 0.03; 97% versus 84%, P < 0.03, respectively). Therefore, the ML test indicated luteolysis with sufficient precision to time the initiation of the Ovsynch protocol between days 5 and 9 of the cycle, however, this did not alter PR compared to starting the protocol randomly throughout the cycle. Initiating the Ovsynch protocol between days 5 and 9 of the cycle increased PR, and improved the efficacy of each injection.     

Reproductive performance of dairy cows with ovarian cysts after different GnRH and cloprostenol treatments

Cystic ovarian disease is an important cause of reproductive failure and economic loss for the dairy industry. This report describes two consecutive studies. The objective of the first was to evaluate the response of cows with ovarian cysts to two therapeutic treatments. In the second study, we compared the effectiveness of the best treatment established in Study 1 with that of the Ovsynch protocol. For Study 1, cows were considered to have an ovarian cyst if it was possible to observe a single follicular structure with a follicular antrum diameter > 25 min in the absence of a corpus luteum in three ultrasonographic examinations performed at 7 days intervals. At diagnosis (Day 0), cows were assigned to one of two treatment groups. Cows in Group GnRH/CLP (n = 31) were treated with 100 microg GnRH i.m. and 500 microg cloprostenol (CLP) i.m. on Day 14. Cows in Group GnRH-CLP/CLP(n = 32) were treated with 100 microg GnRH i.m. plus 500 microg CLP i.m. on Day 0, and 500 microg CLP i.m. on Day 14. The animals were inseminated at observed estrus and monitored weekly by ultrasonography for 4 weeks or until Al. Cows in the GnRH-CLP/CLP group showed a lower cystic persistence rate (15.6% < 45.2%; P = 0.01); a higher estrus detection rate (84.4% > 41.9%; P < 0.0001); a higher ovulation rate (75% versus 32.3%; P < 0.0001) and a higher early response rate (31% > 3%; P = 0.02) than those in the GnRH/CLP group. For the second study, 128 cows with ovarian cysts were randomly assigned to one of two treatment groups: cows in Group Ovsynch (n = 64) were treated with 100 microg GnRH i.m. on Day 0, 500 microg CLP on Day 7, and 100 microm GnRH i.m. 36 h later. Cows in this group were inseminated 24 h after the second GnRH dose (Ovsynch protocol). Cows in Group GnRH-CLP/CLP/GnRH (n = 64)were treated as those in the GnRH-CLP/CLP group of Study 1 but received GnRH 32 h after the second CLP treatment and were inseminated 24 h after this. A further group of cows without ovarian cysts inseminated at natural estrus served as the Group Control (n = 64). Cows in the GnRH-CLP/CLP/ GnRH group showed a lower cystic persistence rate (10.9% < 46.9%; P < 0.0001); higher ovulation rate (79.7% > 17.2%; P < 0.0001); higher return to estrus rate (34.3% > 12.5%; P < 0.01) and higher pregnancy rate (28.1% > 3.1%; P < 0.01) than those in Ovsynch; and a similar pregnancy rate (28.1% versus 35.9%) to Control cows. These findings indicate that lactating cows with ovarian cysts can be successfully synchronized and time inseminated using a protocol that combines GnRH and CLP, starting treatment by simultaneously administering both products. This protocol also allows the insemination of cows showing estrus within the first week of treatment. Ovarian cysts were less responsive when treatment was started with GnRH alone.     

Luteal activity at the onset of a timed insemination protocol affects reproductive outcome in early postpartum dairy cows

This study was designed to compare two timed insemination protocols, in which progesterone, GnRH and PGF2alpha were combined, with the Ovsynch protocol in presynchronized, early postpartum dairy cows. Reproductive performance was also evaluated according to whether cows showed high or low plasma progesterone concentration, at the onset of treatment. One hundred and six early postpartum dairy cows were presynchronized with two cloprostenol treatments given 14 days apart, and then assigned to one of the three treatment groups. Treatments for the synchronization of estrus in all three groups started 7 days after the second cloprostenol injection, which was considered Day 0 of the actual treatment regime. Cows in the control group (Ovsynch, n=30) were treated with GnRH on Day 0, PGF2alpha on Day 7, and were given a second dose of GnRH 32 h later. Cows in group PRID (n=45) were fitted with a progesterone releasing intravaginal device (PRID) for 9 days, and were given GnRH at the time of PRID insertion and PGF2alpha on Day 7. In group PRID/GnRH (n=31), cows received the same treatment as in the PRID group, but were given an additional GnRH injection 36 h after PRID removal. Cows were inseminated 16-20 h after the administration of the second GnRH dose in the Ovsynch group, and 56 h after PRID removal in the PRID and PRID/GnRH groups. Ovulation rate was determined on Day 11 postinsemination by detecting the presence of a corpus luteum in the ovaries. Lactation number, milk production, body condition at the onset of treatment and treatment regime were included as potential factors influencing ovulation and pregnancy after synchronization. Logistic regression analysis for cows with high and low progesterone concentration on treatment Day 0 revealed that none of the factors included in the models, except the interaction between progesterone and treatment regime, influenced the risk of ovulation and pregnancy significantly. In cows with high progesterone concentration at treatment onset, Ovsynch treatment resulted in a significantly improved pregnancy rate over values obtained following PRID or PRID/GnRH treatment. In cows with low progesterone concentration, PRID or PRID/GnRH treatment led to markedly increased ovulation and pregnancy rates with respect to Ovsynch treatment. These findings suggest the importance of establishing ovarian status in early postpartum dairy cows before starting a timed AI protocol, in terms of luteal activity assessed by blood progesterone.     

Effect of CIDR-based protocols for timed-AI on the conception rate and ovarian functions of Japanese Black beef cows in the early postpartum period

Our objectives were to compare: (1) conception rates (in early postpartum Japanese Black beef cows) to timed-artificial insemination (timed-AI) among Ovsynch and Ovsynch plus CIDR protocols, and a protocol that used estradiol benzoate (EB) in lieu of the first GnRH of the Ovsynch plus CIDR; and (2) the effects of these protocols on blood concentrations of ovarian steroids. Cows in the control group (Ovsynch; n=35) underwent a standard Ovsynch protocol (GnRH analogue on Day 0, PGF(2 alpha) analogue on Day 7 and GnRH analogue on Day 9), with timed-AI on Day 10, approximately 20 h after the second GnRH treatment. Cows in the Ovsynch+CIDR group (n=31) received a standard Ovsynch protocol plus a CIDR for 7 days (starting on Day 0). Cows in the third treatment group (EB+CIDR+GnRH; n=41) received 2mg of EB on Day 0 in lieu of the first GnRH treatment, followed by the same treatment as in the Ovsynch+CIDR protocol. The conception rate tended to be greater in the Ovsynch+CIDR group (67.7%, P<0.15) and was greater in the EB+CIDR+GnRH (73.2%, P<0.05) and CIDR-combined (both CIDR-treated groups were combined) groups (70.8%, P<0.05) than in the Ovsynch group (48.6%). Plasma progesterone concentrations were higher on Day 7 (P<0.01) and lower on Days 14, 17 and 21 (P<0.001) in the CIDR-combined group than in the Ovsynch group. Plasma estradiol-17beta concentrations were higher on Day 7 in the Ovsynch group of non-pregnant cows than in the CIDR-combined group of non-pregnant cows and in an all-combined group (all treatment groups combined) of pregnant cows (P<0.01). Furthermore, estradiol-17beta concentrations were lower on Day 9 in the Ovsynch and CIDR-combined groups of non-pregnant cows than in the all-combined group of pregnant cows (P<0.05). In conclusion, both protocols using CIDR improved conception rates following timed-AI in early postpartum suckled Japanese Black beef cows relative to the Ovsynch protocol. Treatment with a CIDR may prevent early maturation of follicles observed in non-pregnant cows treated with the Ovsynch protocol, by maintaining elevated blood progesterone concentrations until PGF(2 alpha) treatment.     

Use of medroxyprogesterone acetate (MAP) in lactating Holstein cows within an Ovsynch protocol: follicular growth and hormonal patterns

To evaluate the effects of incorporating medroxyprogesterone acetate (MAP) in an Ovsynch protocol, cyclic lactating dairy cows were assigned randomly to two groups (control and MAP, n=8 each). Ovsynch treatment (Day 0: GnRH, Day 7: PG, Day 9: GnRH) was initiated at random stages of the estrous cycle (control) and an intravaginal polyurethane sponge impregnated with 300mg of MAP was inserted intravaginally in the MAP group at Day 0 and removed at Day 7 of the Ovsynch protocol (MAP treatment). Ovaries were scanned daily from Day 0 until the second GnRH treatment on Day 9 and from then every 6h for 36 h. Milk samples were collected three times weekly starting 17 days before the initiation of treatment to determine the stage of the cycle at the beginning of the Ovsynch protocol. Blood samples were collected to monitor estradiol (E2), progesterone (P4), LH, and 15-ketodihydro-PGF(2alpha) (PGFM) by RIA. Response to the first GnRH treatment varied with the stage of the cycle at the time of initiation of treatment, as cows in metestrous and late diestrous did not ovulate. In cows ovulating, growth rate of the new follicle was not affected by the addition of MAP. No treatment differences were found in E2 concentrations which reached a maximum at Day 9, consistent with the maximum follicular size. At Day 7, cows with luteal concentrations of P4 had increased concentrations of PGFM, but cows with basal P4 did not show an active release of prostaglandins. There were no treatment differences in the ovulatory response to the second GnRH-induced ovulation, with 11 of the 16 cows ovulating between 16 and 32 h. The addition of MAP to the Ovsynch protocol could not mimic the normal high progesterone levels needed to prevent premature ovulations in those cows with premature CL regression.     

Timing of final GnRH of the Ovsynch protocol affects ovulatory follicle size,subsequent luteal function,and fertility in dairy cows

Synchronization of ovulation (Ovsynch) is an effective method for controlling time of first and subsequent AI in lactating dairy cows. However, validation of the original Ovsynch program did not include testing the optimal time to deliver the final treatment of GnRH. In Experiment 1, the effect of administering the final dose of GnRH on the same day as prostaglandin F2alpha (PGF2alpha) administration was tested. Lactating dairy cows (n = 218) were randomly assigned to receive either Ovsynch (OV; cows were given 100 microg GnRH, then 7 days later cows were administered 25mg PGF2alpha followed by a subsequent treatment of 100 microg GnRH 2 days after the PGF2alpha or the modified version of Ovsynch (MOV; cows were given 100 microg GnRH, then 7 days later cows were administered 25mg PGF2alpha followed immediately with 100 microg GnRH). In both treatment groups, AI took place 16 h after the final administration of GnRH. In Experiment 2, cows (n = 457) were randomly divided into four treatment groups that were administered GnRH 0, 12, 24 and 36 h following PGF(2alpha). The 36 h treatment group served as control. Pregnancy diagnoses were performed by palpation per rectum 36 days post-AI in Experiment 1 and by ultrasonography on Day 28 in Experiment 2. In Experiment 1, pregnancy rate/AI (PR/AI) was greater (P<0.025) in OV versus MOV. In a subset (n = 85), percentage of cows with both synchronized ovulations and regressed CL following administration of PGF2alpha were similar (P>0.1) between OV and MOV, respectively. All cows that became pregnant in the MOV subset group showed regression of the CL in response to the PGF2alpha. Diameter of the ovulatory follicle at the time of final GnRH administration was greater (P<0.05) in OV versus MOV. In Experiment 2, the synchronization rate was once again similar among treatments (P>0.28). There was a linear effect of treatment on follicle size (P<0.05) and PR/AI (P<0.0001) as time increased between administration of PGF2alpha and GnRH, with the greatest PR/AI at 36 h. There was a trend for a greater percentage of cows with short luteal phases in the 0 h group (P<0.10). In summary, delivering the final treatment of GnRH of the Ovsynch program at the same time as PGF2alpha, or in the 24h following PGF2alpha, resulted in lower fertility compared to controls.     

Use of Ovsynch in dairy herds--differences between primiparous and multiparous cows

Ovsynch protocols are used to increase service rate and decrease days open and cullings for infertility. Recent reports have indicated better results after Ovsynch in primiparous than in older cows. However, this was not observed in all investigations on the subject. The objective of the study was to evaluate differences between primiparous and multiparous cows after synchronization of ovulation with an Ovsynch protocol in six trials. A total of 1584 cows (583 primiparous and 1001 multiparous cows, respectively) on three dairy farms were synchronized with an Ovsynch protocol consisting of a GnRH-analogue at Days 0 and 9, and a prostaglandin F(2alpha) analogue on Day 7. AI was carried out in all cows 16-20 h after the last treatment. Cows were categorized into primiparous and multiparous cows for analysis. Conception rate (CR) to timed AI, to further AI, overall conception rate and proportion of cows pregnant by 200 days in milk were compared between the age groups. Finally, two logistic regression models were calculated with conception to first service and conception by 200 DIM as the outcome variables. Independent variables were trial (categorical) and age group (primiparous versus multiparous). Conception rates to TAI were higher in primiparous than in older cows (37.9% versus 31.6%, P=0.015). Likewise pregnancy rates by 200 DIM were higher in primiparous cows (81.8% versus 75.4%, P=0.003). However, the extent of the difference varied between trials. Results indicate that Ovsynch protocols are more effective in primiparous than in older cows.     

Strategies to optimize reproductive efficiency by regulation of ovarian function in yak (Poephagus grunniens L.)

Ovulatory response to the first GnRH of Ovsynch is the critical determinant for successful synchronization of ovulation in animals. An attempt was made in this study to design a pre-Ovsynch hormonal strategy in yaks to increase the ovulatory response to the first GnRH injection of Ovsynch so that overall synchronization rate to Ovsynch could be improved. Non-lactating cyclic yak cows (n=33) were assigned to receive either no treatment before Ovsynch (control) or 0.375 mg of PGF2alpha (PreP) followed 2 d later by 10 microg of GnRH (PreG), administered 4 (G4G), 5 (G5G), or 6 (G6G) d before initiating the Ovsynch protocol. Rectal palpation was performed to assess ovulation and blood samples were collected to measure progesterone concentrations during pre-treatment, treatment and post-treatment periods. All the animals received timed AI 12 and 24h after the final GnRH of Ovsynch. Diagnoses for pregnancy were performed by rectal palpation and profiles of plasma progesterone 35 d after AI. Percentage of yak cows that ovulated in response to the first GnRH injection of Ovsynch, synchronized to Ovsynch treatment, had a functional CL at PGF2alpha of Ovsynch, had circulating concentrations of P4 at PGF2alpha of Ovsynch and likely to be pregnant after 35 d after AI, were greater in G6G and G5G compared with control, whereas G4G did not differ from controls. In addition, animals that ovulated in response to first GnRH of Ovsynch had greater response to PGF2alpha of Ovsynch and greater synchronization rate to the overall protocol than those that did not ovulate. In summary, PGF2alpha-and-GnRH-based pre-Ovsynch strategies consisting of 5 or 6-d interval between PreG and first GnRH of Ovsynch resulted in a greater ovulatory and luteolytic response to first GnRH andPGF2alpha of Ovsynch, respectively, compared with control animals. These, in turn, optimized synchronization rate to Ovsynch in yaks.     

Effect of milk production on the incidence of double ovulation in dairy cows

To determine the effect of parity and milk production on the incidence of double ovulation, the synchronization of ovulation, using GnRH and prostaglandin F2 alpha followed by timed AI (Ovsynch), was initiated at a random stage of the estrous cycle in lactating Holstein cows (n = 237). Ovulatory response at 48 h after the second GnRH injection and conception rate at 28 d post AI were determined by transrectal ultrasonography. Ovulation was synchronized in 84% of cows receiving the Ovsynch protocol. Of the synchronized cows, 14.1% exhibited a double ovulation and 47.6% conceived. Conception rate tended to be greater (P = 0.08) for cows exhibiting double (64.0%) rather than single ovulation (45.2%). To determine the effect of milk production on the incidence of double ovulation, cows were classified into low (< or = 40 kg/d) or high (> 40 kg/d) milk production groups based on the average milk production of 40.5 +/- 0.8 kg/d collected 2 d before AI. Although the incidence of double ovulation tended to increase linearly (P = 0.09) with increasing parity, the incidence of double ovulation was nearly 3-fold greater (P < 0.05) for cows in the high (20.2%) than the low (6.9%) milk production group. Furthermore, the increase in the incidence of double ovulation with parity apparently occurred because, within a parity group, the proportion of cows with high milk production was greater for the older cows. Twinning rate of cows that calved (n = 58) was 5.2%. In a secondary objective, cows were retrospectively classified as cystic or normal based on ultrasonographic ovarian morphology at the time of the second GnRH injection. Incidence of ovarian cysts was 11%, and the synchronization and conception rate of cows classified as cystic was 73.1 and 36.8%, respectively, which did not differ from that of normal cows. We conclude that milk production is the primary factor affecting the incidence of double ovulation in lactating dairy cows and may explain the effect of parity on twinning rate. In addition, Ovsynch appears to be an effective method for establishing pregnancy in lactating dairy cows with ovarian cysts.     

Effect of season on luteal activity during the post partum period of dairy cows in temperate areas

Seasonal effects on luteal activity during post partum were evaluated in two consecutive studies in 253 dairy cows in Northern Italy. In study 1, plasma progesterone concentrations were determined on days 14, 21, 28, 35, 42, 49 and 56 post partum and in study 2 cows were synchronized and inseminated at a fixed time using two regimes based on the 'Ovsynch' protocol. Study 1: Animals were classified as luteal (progesterone >1.5 ng/ml in at least two consecutive samples) or non-luteal (progesterone <1.5 ng/ml in all samples). The proportion of cows without luteal activity from calving to day 56 post partum was 47/253 (18.5%). Of the 47 cows without luteal activity, 42 (89%) were detected during the warm months of the year and five were detected during the cold months of the year, and the effect of season was highly significant (P < 0.001). Study 2: Three study groups were established; control (CONT, untreated cows, n = 92), GPG (cows receiving gonadotropin-releasing hormone (GnRH) on day 0, PGF2α on day 7 followed by a second dose of GnRH 24 h later, n = 80); and HPH (the same as the GPG group, but with human chorionic gonadotropin (hCG) substituted for GnRH, n = 81). In the GPG and HPH groups, cows were inseminated 16 to 22 h after the second GnRH or hCG injection. Untreated cows were inseminated at first estrus after a voluntary weaning period. Because the effects of the GPG and HPH regimes on pregnancy rate were not significantly different, data were pooled into a single treatment group (TREAT). Pregnancy rates during the warm months of the year were 16% and 15% at first service and 65% and 66% at day 135 post partum for CONT and TREAT groups, respectively. Pregnancy rates during the cold months of the year were 36% and 38% at first service and 72% and 76% at day 135 post partum for CONT and TREAT groups, respectively. There was an effect of season (P < 0.05) but not of treatment on pregnancy rate. Treatment reduced the number of days from calving to conception during both the cold (101 ± 3.2 v. 121 ± 3.1 days; P < 0.001) and warm seasons (122 ± 3.2 v. 145 ± 3.1 days; P < 0.001). In conclusion, the present study shows that (i) heat stress during the warm season can compromise luteal activity and (ii) that regimes based on the Ovsynch protocol did not improve pregnancy rate at first service or by 135 post partum, but they had a positive effect on the calving-to-conception interval.     

Resynchronization of ovulation and timed insemination in lactating dairy cows, II: assigning protocols according to stages of the estrous cycle, or presence of ovarian cysts or anestrus

Pregnancy rates were compared in lactating dairy cows (n = 1083) assigned to protocols for resynchronization of ovulation based on stages of the estrous cycle, or presence of ovarian cysts or anestrus. Cows were detected not pregnant by ultrasonography 30 d after a previous AI (study day 0) and classified as diestrus, metestrus, proestrus, with ovarian cysts or anestrus. Cows in diestrus (January-May) were assigned to either Ovsynch (GnRH day 0, PGF2alpha day 7, GnRH day 9, and timed-AI [TAI] 16 h later; n = 96), or Quicksynch (PGF2alpha day 0, estradiol cypionate [ECP] day 1, AI at detected estrus [AIDE] on day 2, or TAI on day 3; n = 96). Cows in diestrus (June-December) were assigned to either Ovsynch (n = 156) or Modified Quicksynch (PGF2alpha day 0, ECP day 1, AIDE days 2 and 3, and to Ovsynch on day 4 if not detected in estrus; n = 142). Cows in metestrus were assigned either to Ovsynch (n = 68), Heatsynch (GnRH day 0, PGF2alpha day 7, ECP day 8, AIDE day 9, or TAI day 10; n = 62), or GnRH + Ovsynch (GnRH on day 0, followed by Ovsynch on day 8; n = 64). Cows in proestrus, with ovarian cysts, or anestrus were assigned to either Ovsynch (proestrus n = 89, ovarian cysts n = 97, anestrus n = 8) or GnRH + Ovsynch (proestrus n = 87, ovarian cysts n = 109, anestrus n = 9). Pregnancy rate was evaluated 30, 55 and 90 d after resynchronized AI. For cows in diestrus (January-May), pregnancy rates were higher for Ovsynch (35.9, 29.2 and 26.0%) than for Quicksynch (21.7, 16.7 and 15.6%). For cows in diestrus (June-December), pregnancy rates were similar for Ovsynch (34.4, 24.0 and 23.6%) and Modified Quicksynch (27.1, 26.2 and 21.6%). For cows in metestrus, pregnancy rates were higher for GnRH + Ovsynch (33.3, 24.5 and 20.3%) than for Heatsynch (20.3, 12.9 and 9.8%). For cows with ovarian cysts, pregnancy rates were higher for GnRH + Ovsynch (30.3, 26.6 and 22.9%) than for Ovsynch (20.2, 18.5 and 14.7%). Assignment to resynchronization protocols based on the stages of the estrous cycle, or presence of ovarian cysts improved pregnancy rates.     

Resynchronization of ovulation and timed insemination in lactating dairy cows I: use of the Ovsynch and Heatsynch protocols after non-pregnancy diagnosis by ultrasonography

The objective was to compare pregnancy rates and pregnancy losses in lactating dairy cows that were diagnosed not pregnant and re-inseminated following either the Ovsynch or Heatsynch protocols. Also evaluated were the effects of stages of the estrous cycle, ovarian cysts and anestrus on pregnancy rates for both treatments. Non-pregnant cows (n = 332) as determined by ultrasonography on day 27 post-AI (study day 0) were divided into two groups. Cows in the Ovsynch group (n = 166) received GnRH on day 0, PGF2alpha on day 7, GnRH on day 9, and timed AI (TAI) 16 h later (day 10). Cows in the Heatsynch group (n = 166) received GnRH on day 0, PGF2alpha on day 7, estradiol cypionate (ECP) on day 8, and TAI 48 h later (day 10). Cows detected in estrus on days 8 and 9 were inseminated and included in the study. On day 0, cows were classified according to different stages of the estrous cycle, or presence of ovarian cysts or anestrus. Pregnancy rates were evaluated 27, 45 and 90 days after resynchronized AI. Overall, there was no difference in pregnancy rates on days 27, 45 and 90 between cows in the Ovsynch (25.2, 17.5, and 13.9%) and Heatsynch (25.8, 19.9, and 16.1%) groups. There was no difference in pregnancy losses from days 27 to 45 and days 45 to 90 for cows in the Ovsynch (25.0 and 17.9%) and Heatsynch (14.7 and 10.3%) groups. However, pregnancy rates were increased when cows in metestrus were subjected to the Heatsynch protocol and cows with ovarian cysts were subjected to the Ovsynch protocol.     

Ultrasonographic evaluation of endometrial thickness near timed AI as a predictor of fertility in high-producing dairy cows

The objectives were to evaluate changes in endometrial thickness (ET) near the time of a synchronized ovulation and to assess the relationship of ET and fertility in lactating Holstein cows, with or without estrogen supplementation near timed ovulation. In Experiment 1, eight cows were examined with transrectal ultrasonography, once daily for 5 d, starting concurrent with PGF_2α (PGF) treatment during an Ovsynch protocol (GnRH - 7d - PGF - 72h - GnRH). The ET increased rapidly after PGF (from ∼7 to ∼9.5 mm), remained > 9 mm for the next 2 d, then decreased to ∼8 and 7.4 mm, 1 and 2 d, respectively, after the second GnRH. In Experiment 2,642 cows (total of 758 breedings) were subjected to an Ovsynch protocol (GnRH - 7d - PGF - 56h - GnRH - 16h - timed AI); cows received either no further treatment (Ovsynch) or 1 mg of estradiol-17β im 8 h before the second GnRH (Ovsynch + E2). For both uterine horns, ET was measured (∼2 cm from the internal uterine body bifurcation) before E2 treatment (48 h after PGF). In cows with ET ≤ 8 mm vs > 8 mm, rates of ovulation were 86.0% (n = 136) vs 98.1% (n = 472; P < 0.01), respectively, and percentage pregnant per AI (P/AI) were 26.7% (n = 146) vs 42.7% (n = 524; P < 0.01). Treatment with E2 increased P/AI in cows with lower ET (Ovsynch + E2 = 37.0% vs Ovsynch = 23.3%; P = 0.07), but did not significantly improve P/AI in cows with ET > 8 mm (Ovsynch + E2 = 43.4% vs Ovsynch = 42.1%). In conclusion, a single ultrasonographic evaluation of ET in Holstein cows 48 h after PGF treatment in an Ovsynch program was a good predictor of ovulation failure and pregnancy success. Perhaps poor fertility in cows with reduced ET was low peripheral E2 concentrations near AI, poor P4 priming, or luteolysis failure during timed AI procedures.     

A progesterone-based timed AI protocol more effectively prevents premature estrus and incomplete luteal regression than an Ovsynch protocol in lactating Holstein cows

The objective of this study was to evaluate pregnancy rates in lactating Holstein cows treated with an Ovsynch protocol (GnRH-PGF(2alpha)-GnRH) or a progesterone-based timed AI (TAI) protocol, and to determine the factors that may influence pregnancy rate following protocol treatment. In experiment 1, lactating Holstein cows were randomly assigned to three treatments: (1) an injection of GnRH (Day 0), an injection of PGF(2alpha) on Day 7, a second injection of GnRH on Day 9, and TAI 16h after the second GnRH injection (GPG group, n = 34); (2) insertion of a CIDR intravaginal progesterone (1.9g) device combined with a capsule containing 10mg estradiol benzoate (Day 0), an injection of PGF(2alpha) and removal of the device on Day 7, an injection of GnRH on Day 9, and TAI 16h after the GnRH injection (CPG group, n = 34); (3) an injection of PGF(2alpha) after confirming the presence of CL by ultrasonographical observation and artificial insemination at estrus (AIE) (P group, n = 75). The pregnancy rate after TAI following the CPG protocol (41.2%) was higher (P<0.05) than that after TAI following the GPG protocol (20.6%) and that after AIE (20.0%). In experiment 2, lactating Holstein cows were randomly assigned to two treatments: a GPG group (n = 31) and a CPG group (n = 31). The GPG and CPG protocols were identical to those used in experiment 1. The proportion of cows with premature estrus prior to injection of PGF(2alpha) and with incomplete luteal regression tended (P = 0.056) to be or were greater (P<0.05) in the GPG group (4/31, 8/31) than in the CPG group (0/31, 2/31), respectively. Average diameters of dominant follicles (1.5+/-0.1mm versus 1.4+/-0.1mm) on Day 7 and preovulatory follicles (1.8+/-0.1mm versus 1.6+/-0.1mm) on Day 9, and the proportion of cows with synchronized ovulation by 40h after the second GnRH injection were not different (81.5% versus 87.1%, P>0.05) between groups, respectively. We conclude that the pregnancy rate after TAI following the CPG protocol was higher than that after TAI following the GPG protocol, probably due to a decreased incidence of premature estrus and incomplete luteal regression.     

Modifying the double-Ovsynch protocol to include human chorionic gonadotropin to synchronize ovulation in dairy cattle

The objectives were to determine whether rates of conception, ovulation, presynchronization, or follicle and CL characteristics were altered after modifying the Double-Ovsynch (DO) protocol to include hCG compared with the DO protocol. Primiparous and multiparous lactating dairy cows (N = 183), and nulliparous dairy heifers (N = 51) were used. Cows were blocked by parity and heifers were stratified by age and breed before being randomly assigned to one of two treatments. All females received either 100 μg GnRH or 2000 IU hCG im, at initiation of the Pre-Ovsynch (PO) portion of the DO protocol (PO: GnRH/hCG, 7 days PGF_2α and 3 days GnRH). After 7 days, females started the Breeding-Ovsynch portion of the DO protocol (Breeding-Ovsynch: GnRH, 7 days, PGF_2α, 48 or 56 h and GnRH 16 hours timed artificial insemination with sex-sorted semen). Transrectal ultrasonography and blood samples were used to assess ovarian structures, ovulation, pregnancy diagnosis, and concentration of progesterone in plasma. Conception rates were similar in females treated with GnRH or hCG in cows (32.2 and 25.0%) and in heifers (30.8 and 36.0%). Ovulation rates in cows at the onset of PO were increased with hCG compared to GnRH (77.2 vs. 62.2%, P < 0.05). Concentrations of progesterone 7 days post-hCG or GnRH were greater in cows treated with hCG compared with GnRH (least significant mean ± SEM; 4.3 ± 0.3 and 3.0 ± 0.3 ng/mL, P < 0.01), but did not differ in heifers (4.5 ± 0.9 and 2.9 ± 0.9 ng/mL). More cows ovulated within 7 days post-hCG and a greater proportion of these cows tended to have failed luteal regression by Day 3 post-PGF_2α compared with cows that had ovulated to GnRH (29.6 vs. 16.1%, P ≤ 0.10). The overall percentage of females which were synchronized to PO did not differ between GnRH- or hCG-treated cows (61.5% and 52.2%) and heifers (42.3% and 40.0%). In conclusion, no overall improvement in fertility was achieved by replacing the first injection of GnRH in the DO protocol with hCG.     

Strategic treatment of anovular dairy cows with GnRH

The primary objective was to evaluate fertility of anovular dairy cows given GnRH 4 d after first postpartum timed AI (TAI). Secondary objectives were to determine ovulatory response to treatment, effect of treatment on serum progesterone (P(4)) concentrations, and the proportion with a short luteal phase. Lactating Holstein cows (n=1047) were submitted for first postpartum TAI using a Presynch+Ovsynch protocol. Anovular cows were identified from an initial 1047 lactating Holstein cows using transrectal ultrasonography, based on the absence of a CL at the first GnRH injection of a Presynch+Ovsynch protocol, and anovular cows were randomly assigned to receive either no further treatment (Control, n=85), or 100 microg of GnRH 4 d after TAI (GnRH treated; n=71). For GnRH treated cows, 51% responded by ovulating a follicle in response to GnRH treatment 4 d after TAI; however, pregnancies per AI (P/AI) did not differ between GnRH treated cows that ovulated (36%) compared to GnRH treated cows that did not ovulate (21%). There was a quadratic effect of P(4) at the PGF(2 alpha) injection of Ovsynch on P/AI, and cows with P(4)>or=1 ng/mL at the PGF(2 alpha) injection of Ovsynch had greater P/AI (41%) than cows with P(4)<1 ng/mL (12%); however, no treatment difference was detected. Overall, P/AI did not differ between control (30.1%) and GnRH treated (29.6%) treatments for synchronized cows. Although treatment of anovular cows with GnRH 4 d after TAI failed to improve fertility, variation among cows in serum P(4) at the PGF(2 alpha) injection of Ovsynch dramatically affected fertility of anovular dairy cows.