Time and incidence of ovulation and conception rates after incorporating estradiol cypionate into a timed artificial insemination protocol.

Abstract

Two experiments were conducted to determine the effect of estradiol cypionate (ECP), when incorporated into a conventional GnRH-PGF(2alpha)-GnRH timed artificial insemination protocol (Ovsynch), on systemic estradiol (E(2)), time and incidence of ovulation, luteal development, and conception rate in Holstein cows. Our objective was to determine if administration of 0.25 mg of ECP at the time of the second GnRH injection would effectively synchronize ovulation and increase conception rate. In Experiment 1, lactating Holstein cows (n = 23; 58.7 +/- 1.2 d in milk) were synchronized with PGF(2alpha) (at d -10). Ten days later, Ovsynch was initiated with the administration of 100 mug of GnRH (d 0) followed by PGF(2alpha) on d 7. On d 9, cows were assigned randomly to be treated with either GnRH + 0.25 mg of ECP (OVS-ECP; n = 11) or GnRH and 1 mL of cottonseed oil (OVS-C; n = 12). Ovarian activity was monitored by ultrasonography on d 0, 7, and 9. To determine the time of ovulation, ultrasound examinations were conducted at 12 and 20 h posttreatment and then at least every 3 h until either 36 h posttreatment or ovulation was observed. Blood samples were collected on d 0, 7, 9, and 16 for progesterone analysis. Blood samples also were collected at the time of treatment (d 9, 0 h) and at 6, 12, 20, and 28 h for E(2) analysis. Incidence of ovulation did not differ between treatments. Mean ovulation time relative to the second GnRH administration was similar between treatments. Serum progesterone concentration did not differ between treatments at any time. Serum E(2) concentration was not different at the time of treatment (0 h); however, mean E(2) concentration was greater for the OVS-ECP group at 6 and 12 h after treatment compared with OVS-C. In Experiment 2, lactating dairy cows (n = 333) in 3 commercial herds were randomly assigned to OVS-ECP (n = 169) or OVS-C (n = 164). Cows were inseminated 22 to 24 h posttreatment. Conception rates did not differ between treatments. Estradiol cypionate treatment was successful in increasing serum E(2) when administered at the time of the second dose of GnRH in the Ovsynch protocol. Conception rates, however, were not affected by treatment.

Full text

Page 1

J. Dairy Sci. 89:620–626
 American Dairy Science Association, 2006.
Time and Incidence of Ovulation and Conception Rates
After Incorporating Estradiol Cypionate into a Timed
Artificial Insemination Protocol
C. B. Sellars,* J. C. Dalton,* R. Manzo,* J. Day,† and A. Ahmadzadeh*1
*Animal and Veterinary Science Department, University of Idaho, Moscow 83844
†Dairy Health Services, Jerome, ID 83338
ABSTRACT
Two experiments were conducted to determine the
effect of estradiol cypionate (ECP), when incorporated
intoaconventionalGnRH-PGF2α-GnRHtimedartificial
insemination protocol (Ovsynch), on systemic estradiol
(E2), time and incidence of ovulation, luteal develop-
ment, and conception rate in Holstein cows. Our objec-
tive was to determine if administration of 0.25 mg of
ECP at the time of the second GnRH injection would
effectively synchronize ovulation and increase concep-
tion rate. In Experiment 1, lactating Holstein cows (n =
23; 58.7 ± 1.2 d in milk) were synchronized with PGF2α
(at d −10). Ten days later, Ovsynch was initiated with
the administration of 100 ?g of GnRH (d 0) followed by
PGF2αon d 7. On d 9, cows were assigned randomly to
be treated with either GnRH + 0.25 mg of ECP (OVS-
ECP; n = 11) or GnRH and 1 mL of cottonseed oil (OVS-
C; n = 12). Ovarian activity was monitored by ultraso-
nography on d 0, 7, and 9. To determine the time of
ovulation, ultrasound examinations were conducted at
12 and 20 h posttreatment and then at least every 3 h
until either 36 h posttreatment or ovulation was ob-
served. Blood samples were collected on d 0, 7, 9, and
16 for progesterone analysis. Blood samples also were
collected at the time of treatment (d 9, 0 h) and at 6,
12, 20, and 28 h for E2analysis. Incidence of ovulation
did not differ betweentreatments. Mean ovulation time
relativetothesecondGnRHadministrationwassimilar
between treatments. Serum progesterone concentra-
tion did not differ between treatments at any time.
Serum E2concentration was not different at the time
of treatment (0 h); however,mean E2concentration was
greater for the OVS-ECP group at 6 and 12 h after
treatment compared with OVS-C. In Experiment 2, lac-
tating dairy cows (n = 333) in 3 commercial herds were
randomly assignedto OVS-ECP(n = 169)or OVS-C(n =
164). Cows were inseminated 22 to 24 h posttreatment.
Received May 17, 2005.
Accepted October 3, 2005.
1Corresponding author: amin@uidaho.edu
620
Conception rates did not differ between treatments.
Estradiol cypionate treatment was successful in in-
creasing serum E2when administered at the time of
the second dose of GnRH in the Ovsynch protocol. Con-
ception rates, however, were not affected by treatment.
Key words: timed artificial insemination, estradiol
cypionate, conception rate
INTRODUCTION
Systematic breeding programs provide an organized
and efficient approach to administering AI and improv-
ing reproductive efficiency in dairy herds. To alleviate
the difficulties associated with detection of estrus and
to increase the AI submission rate, timed AI programs
have been developed such as Ovsynch (Pursley et al.,
1995). Ovsynch (d 0 GnRH, d 7 PGF2α, d 9 GnRH, timed
AI 8 to 24 h) was designed to synchronize ovulation,
thereby allowing timed AI of all cows without detection
of estrus. Synchronization of ovulation in 84 to 100% of
cows can be expected (Pursley et al., 1995; Fricke et
al.,1998;Vasconcelos etal.,1999;Cartmill etal.,2001).
Conception rates following timed AI associated with
Ovsynch rangefrom 22to 42%in dairycows (Stevenson
et al., 1996; Pursley et al., 1997a,b, 1998; Fricke et al.,
1998; Jobst, 1998; Stevenson et al., 1999). In practice,
although AI submission rates are 100% when using
Ovsynch, conception rate may be reduced with this pro-
tocol compared with cows that receive AI following de-
tected estrus (Stevenson et al., 1999; Santos et al.,
2004). Nevertheless, pregnancy rate (AI submission or
estrus-detection rate × conception rate) achieved either
after timed AI or after detected estrus and AI may
be comparable.
Although timed AI protocols such as Ovsynch are a
convenient method to increase AI submission rate, a
numberofstudiesindicatemechanismsbywhichinduc-
tion of ovulation by administration of GnRH during
proestruscandisturbnormalreproductivefunctionand
negatively affect conception rates. Lucy and Stevenson
(1986) found that an injection of GnRH during peries-
trus, and before the preovulatory LH surge, decreased
serum estradiol (E2) and reduced fertility compared

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MODIFICATION OF THE OVSYNCH PROTOCOL
621
with cows having a spontaneous LH surge. These re-
sults are further supported by Kobayashi (1995) who
showed that administration of GnRH after PGF2α
caused a cessation of estrogen secretion by the preovu-
latory follicle as evidenced by a decline in blood concen-
trations of E2. Thatcher and Chenault (1976) reported
thatGnRHtreatmentofcattle48hafterPGF2αreduced
the frequency of estrus compared with PGF2α-induced
estrus alone. Those authors suggested that this obser-
vation might be due to alterations in plasma progestins
and E2. In vitro studies (Uemura et al., 1994; Takekida
et al., 2000) have shown that GnRH agonists directly
inhibit ovarian and granulosa cell steroidogenesis [E2
and progesterone (P4)]. Lower frequency of estrus in
cows treatedwith GnRH48 hafter PGF2α(Rodriguez et
al.,1975;ThatcherandChenault,1976),takentogether
with suppressed estrual behavior following the second
GnRH injection in cows subjected to Ovsynch (Pursley
et al., 1995; Twagiramungu et al., 1995; Stevenson et
al., 1996, 1999, and 2000; Jobst, 1998) as well as in
vitro studies give further support to the premise that
induction of ovulation during the follicular phase, as
occurs with Ovsynch, suppresses E2secretion. Further,
exogenous E2increases uterine contractions, efficiency
of sperm transport, number of sperm in the oviducts,
retention and adhesion of sperm to oviductal epithe-
lium, enhances sperm capacitation and the true acro-
some reaction, and increases fertilization (Hawk and
Cooper, 1978; Hawk, 1983, 1987; Bathla et al., 1999;
Langendijk et al., 2002). Consequently, conception
rates observed following GnRH-induced ovulation and
timedAImaynotbeoptimizeddue,inpart,toasynchro-
nous timing of the GnRH-induced LH surge and final
follicular maturation coupled with limited or brief E2
secretion around the time of estrus (Lucy and Steven-
son, 1986; Stevenson et al., 1999; Taponen et al., 1999).
Another factor potentially limiting conception rate in
cows enrolled in the Ovsynch protocol (without presyn-
chronization) might be due to a 16 to 19% lack of syn-
chronization of ovulation after the second GnRH (Vas-
concelos et al., 1999).
Our hypothesis was that administration of 0.25 mg
of estradiol cypionate (ECP) at the time of the second
dose of GnRH in the Ovsynch protocol (OVS-ECP)
wouldimprovefertilitycomparedwiththeconventional
Ovsynch protocol (OVS-C). The objectives of these ex-
periments were to determine the effect of 0.25 mg of
ECP when incorporated into a conventional Ovsynch
protocolonsystemicE2,timeandincidenceofovulation,
luteal development, andfirst-service AI conception rate
in dairy cows.
MATERIALS AND METHODS
Experiment 1
Experiment 1 was conducted at the University of
Idaho Dairy Center using 23 lactating Holstein cows
Journal of Dairy Science Vol. 89 No. 2, 2006
(DIM = 58.7 ± 1.3 d). Mean (± SD) BW was 664 ± 92
kg and mean (± SD) BCS was 2.6 ± 0.5 (scale = 1 to
5, with 1 being emaciated and 5 being grossly over-
conditioned). Average 3.5% FCM yield at the DHI test
closest to the day of treatment was 41.1 ± 10.2 kg (mean
±SD).Basedontransrectalpalpationbytheherdveter-
inarian, experimental cows had no abnormalities of the
reproductive tract at the initiation of the experiment.
Ovulation Synchronization and Treatment. On
d −10, estrous cycles ofcows were presynchronized with
an i.m. injection of 25 mg of PGF2α(Lutalyse; Pfizer
Animal Health, New York, NY). Ond 0, and after detec-
tion of a corpus luteum (CL) by transrectal ultrasonog-
raphy (Sonovet 600, 5-MHz probe, Universal Ultra-
sound, Bedford Hills, NY), the Ovsynch protocol was
initiated with a dose of GnRH (100 ?g) administered
i.m. (Cystorelin; Merial, Athens, GA). Seven days later
(d 7), all cows received (i.m.) 25 mg of PGF2αto regress
the CL. Forty-eight hours after PGF2αtreatment (d 9),
cows were assigned randomly to treatment (n = 11;
OVS-ECP) or control (n = 12; OVS-C). The OVS-ECP
cows received (i.m.) GnRH (100 ?g) + 0.25 mg of ECP
(Pharmacia Animal Health, Kalamazoo, MI), whereas
the OVS-C cows received GnRH (100 ?g) + cottonseed
oil (1 mL). The ECP was diluted in purified cottonseed
oil (Sigma-Aldrich Corp., St. Louis, MO).
Ovarian Examination and Blood Collection. On
d 0, 7, and 9, ovaries were examined via transrectal
ultrasonography and structures recorded. Time of ovu-
lation after treatment (injection of GnRH or GnRH +
ECP)wasdeterminedbyultrasonographythatwascon-
ducted at 12 and 20 h after treatment and then at
least every 3 h thereafter until either ovulation or 36
h, whichever occurred first. Ovulation was defined as
thedisappearanceofanyantralfollicle≥10mmindiam-
eteratthetimeofanultrasoundexaminationcompared
with the previous ultrasound examination (Kaneko et
al., 1991). Time of ovulation was defined as the number
of hours from the time of treatment to the midpoint
of the 2 examinations between which ovulation had
occurred (Walker et al., 1996). On d 16, ultrasono-
graphic examinations of the ovaries were performed to
confirm the occurrence of ovulation, as evidenced by
the presence of a CL.
On d 0, 7, 9, and 16, coccygeal blood samples were
collected for later measurement of P4concentrations to
determine the ovarian response to the hormonal treat-
ments. Coccygeal blood samples were obtained at 0, 6,
12, 20, and 28 h after treatment to determine whether
ECPinducedasignificantchangeinserumE2compared
with controls. Blood samples were immediately placed
in ice and stored at 4°C for a minimum of 20 h to allow
clotting. All samples were centrifuged at 4°C for 30 min

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SELLARS ET AL.
622
at 2,750 × g. Serum was harvested and stored at −20°C
until assayed for E2or P4.
Hormone Assays. Serum E2concentrations were de-
termined by radioimmunoassay as described by Perry
etal.(1991)andwaskindlyperformedbythelaboratory
of Matthew Lucy (University of Missouri, Columbia).
The assay was conducted in nonequilibrium conditions
and the standard curve and all samples were assayed
in duplicate. Primary antiserum bound 35% of125I-E2
in the absence of unlabeled E2. Intra- and interassay
coefficients of variation (CV) were 9.3 and 5.5%, respec-
tively.
Serum P4concentrations were determined using a
solid-phase radioimmunoassay (Diagnostic Products
Corp., Los Angeles, CA). The assay was conducted un-
der equilibrium conditions. The standard curve ranged
from 0.1 to 40 ng/mL, and all samples were assayed in
duplicate. Intraassay CV was 7.8%.
Statistical Analyses: Experiment 1
Analysisofrepeatedmeasuresusingthemixedproce-
dure of SAS (Littell et al., 1998) was used to analyze
serum E2data. The statistical model included treat-
ment, the repeated factor time, and treatment × time
interaction. Cow within treatment was designated as
arandomeffectandpretreatmentserumE2valueswere
used as covariates in the model.
Serum P4data were analyzed by least squares AN-
OVA using the GLM procedure of SAS (SAS Inst. Inc.,
Cary, NC). The statistical model included treatment.
Separate analyses were performed for each sampling
day (d 0, 7, 9, and 16) to verify that cows in both treat-
ments had similar P4concentrations at the initiation
of OvSynch and to determine ovarian response to the
first GnRH, second PGF2α, and second GnRH adminis-
trations.
Experiment 2
Experiment 2 was conducted to determine the effect
of OVS-ECP on first-service conception rate in artifi-
cially inseminated dairy cows. Three commercial dair-
ies in southern Idaho participated in a field trial. Three
hundred thirty-three multiparous cows (OVS-ECP: n =
169; OVS-C: n = 164), having fewer than 80 DIM, were
submittedforthefirstpostpartuminseminationforthis
experiment.Hormonaltreatmentsweresimilartothose
in Experiment 1, except that all cows received PGF2α
14 d before the first dose of GnRH (d 0). Before d 9,
cows were assigned randomly to either OVS-ECP or
OVS-C by the herd manager at each farm and without
the knowledge of personnel who administered treat-
ments. On each dairy, cows were inseminated approxi-
Journal of Dairy Science Vol. 89 No. 2, 2006
Table 1. Serum progesterone concentrations (ng/mL; least squares
mean ± SEM) on d 0, 7, 9, and 16 in lactating Holstein cows treated
with Ovsynch with and without estradiol cypionate (Experiment 1)
Day of experiment
Treatment1
079 16
OVS-ECP (n = 11)
OVS-C (n = 12)
3.0 ± 0.8
4.0 ± 0.7
3.2 ± 0.8
4.8 ± 0.8
0.5 ± 0.3
0.8 ± 0.3
3.2 ± 0.4
3.1 ± 0.3
1OVS-ECP = Ovsynch was initiated with the administration of 100
?g GnRH (d 0) followed by PGF2αon d 7. On d 9 cows received (i.m.)
GnRH (100 ?g) + 0.25 mg of estradiol cypionate (ECP); OVS-C =
Ovsynch was initiated with the administration of 100 ?g of GnRH
(d 0) followed by PGF2αon d 7. On d 9, cows received GnRH (100 ?g)
+ cottonseed oil (1 mL).
mately 22 to 24 h after treatment by a single insemina-
tor.Cowsin all3herdswereobservedfor signsofestrus
oncedailybasedontailchalkremoval.Cowsthatexhib-
ited estrus on or before d 9 received AI immediately
and were removed from the experiment. Conception
rates (number of confirmed pregnancies divided by
number of cows inseminated) were calculated based
upon pregnancy diagnosis by the herd veterinarian via
palpation per rectum of uterine contents at 35 to 42 d
after AI.
Statistical Analyses: Experiment 2
Conception rate data were analyzed using the Logis-
tic procedure of SAS (SAS Inst.). The model included
the effects of treatment, herd, and the treatment × herd
interaction. Based on the number of cows used in this
experiment, a 13% difference in conception rate could
be detected. This sensitivity was calculated (Agresti,
1990) for α = 0.07 and β = 0.20 and an average preg-
nancy rate of 30%.
RESULTS
Experiment 1
Average BW and BCS did not differ between treat-
ments.Meandaily3.5%FCMproductionwasnotdiffer-
entbetween treatments.Moreover, meanserum P4con-
centrations did not differ between treatments at any
time (Table 1).
Atime×treatmentinteraction(P<0.05)wasdetected
for serum E2concentrations. At 0 h, mean serum E2
concentrations were 5.9 ± 0.4 pg/mL for OVS-ECP cows
and 5.3 ± 0.3 pg/mL for OVS-C cows and did not differ
(Figure 1). Mean serum E2concentrations were differ-
ent between treatments at 6 (P < 0.05) and 12 h (P =
0.05; Figure 1). At 6 h posttreatment, mean serum E2
concentration in OVS-ECP cows (6.0 ± 0.4 pg/mL) was
greater (P < 0.05) than that of OVS-C cows (4.4 ± 0.3 pg/

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MODIFICATION OF THE OVSYNCH PROTOCOL
623
Figure 1. Mean serum concentrations of estradiol in cows treated
with Ovsynch with (OVS-ECP) and without (OVS-C) estradiol cypio-
nate injection coincident with the second GnRH injection of Ovsynch
(Experiment 1). Treatment (0 h) with either GnRH or GnRH + estra-
diol cypionate occurred at 0 h. *Means differed (P ≤ 0.05) between
treatments at 6 and 12 h.
mL;Figure1).MeanserumE2concentrationdeclinedin
both groups between 6 and 12 h after treatment, but
remained greater(P =0.05) forthe OVS-ECPcows than
for the OVS-C cows at 12 h (3.77 ± 0.36 vs. 2.58 ± 0.33
pg/mL).
Time of ovulation did not differ between treatments
(Table 2). Range in time of ovulation was 16 to 32 h
and 26 to 29 h for the OVS-ECP and OVS-C cows,
respectively. Incidence of ovulation did not differ and
was 100% for OVS-ECP and 92% for OVS-C group.
Mean diameter of the ovulatory follicle, measured at
20 h after treatment, was similar between groups (18.3
± 1.2 vs. 18.1 ± 1.4 mm; Table 2). In the present study,
82% of all observed ovulations were on the right ovary
(18 of 22; Table 2).
Table 2. Time and incidence of ovulation, and size of ovulatory follicle in lactating Holsteins treated with
Ovsynch with and without estradiol cypionate (Experiment 1)
Diameter of
ovulatory
follicle,2
mm
Right
ovary
ovulation,
%
Time of
ovulation,2h
Incidence of
ovulation, %
Treatment1
OVS-ECP (n = 11)
OVS-C (n = 12)
26 ± 1
27 ± 1
100
92
18.3 ± 1.2
18.1 ± 1.5
82
83
1OVS-ECP = Ovsynch was initiated with the administration of 100 ?g GnRH (d 0) followed by PGF2αon
d 7. On d 9 cows received (i.m.) GnRH (100 ?g) + 0.25 mg of estradiol cypionate (ECP); OVS-C = Ovsynch
was initiated with the administration of 100 ?g of GnRH (d 0) followed by PGF2αon d 7. On d 9, cows
received GnRH (100 ?g) + cottonseed oil (1 mL).
2Means ± SE.
Journal of Dairy Science Vol. 89 No. 2, 2006
On d 16, all cows were subjected to ultrasonographic
ovarian examination of the ovaries and all had a CL
on the ovary where ovulation had been previously de-
tected. Mean serum P4concentrations on d 16 did not
differ between groups and were 3.2 ± 0.4 and 3.1 ±
0.3 ng/mL in the OVS-ECP and OVS-C, respectively
(Table 1).
Experiment 2
Neither treatment, herd, nor treatment × herd inter-
action influenced conception rate. Overall conception
rate for the OVS-ECP group was 31.4% and for the
OVS-C group was 36.6% (Table 3).
DISCUSSION
Experiment 1
In the present study, serum E2concentrations were
greater in OVS-ECP cows at 6 and 12 h after ECP
treatmentcomparedwithOVS-Ccows(Figure1).Tapo-
nen et al. (1999) found that when GnRH was adminis-
tered 24 h after PGF2αinjection, serum E2declined to
basal concentrations within 1 d. Mee et al. (1993) found
that serum E2declined after estrus in both saline- and
GnRH-treated cows; however, serum E2was less in the
GnRH-treated group. Serum E2profiles in OVS-ECP
cows differed from those in the OVS-C cows indicating
that 0.25 mg of ECP was able to alter circulating E2in
cows during the first 12 h after treatment (Figure 1).
Lucy and Stevenson (1986) administered GnRH or
salineat72hafterPGF2αandfoundthatGnRH-treated
cows had a mean serum E2 concentration approxi-
mately 3 pg/mL less than that of saline-treated cows.
In the present study, serum E2in OVS-C group was
36% less at 6 h after GnRH administration compared
with that in the OVS-ECP group. Serum E2profile of
the OVS-C group might suggest that the time of GnRH
administrationwasasynchronouswiththenaturaltim-
ing of neuroendocrine events following PGF2α; thus, the

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SELLARS ET AL.
624
Table 3. Conception rate (number of pregnant cows/total number of
cows inseminated) in lactating Holstein cows treated with Ovsynch
with and without estradiol cypionate (Experiment 2)
Treatment1
Herd OVS-ECPOVS-C
% (no.)
1
2
3
Total
29.4 (51)
33.9 (56)
30.6 (62)
31.4 (169)
40.4 (47)
38.6 (57)
31.7 (60)
36.6 (164)
1OVS-ECP = Ovsynch was initiated with the administration of 100
?g GnRH (d 0) followed by PGF2αon d 7. On d 9 cows received (i.m.)
GnRH (100 ?g) + 0.25 mg of estradiol cypionate (ECP); OVS-C =
Ovsynch was initiated with the administration of 100 ?g of GnRH
(d 0) followed by PGF2αon d 7. On d 9, cows received GnRH (100 ?g)
+ cottonseed oil (1 mL).
preovulatory follicle may not have matured adequately
before the GnRH-induced LH surge as originally hy-
pothesized by Lucy and Stevenson (1986).
Ovulations in both groups occurred between 16 and
32 h after GnRH administration (Table 2), which is
comparable to findings of other researchers (Pursley et
al.,1995;Ahmadzadehetal.,2002).Timeofovulationin
our study was similar to that after spontaneous estrus
(27.9 ± 5.6 h after the onset of estrus; Walker et al.,
1996).
The greater E2concentration observed in the OVS-
ECP compared with OVS-C group was not related to
size of the ovulatory follicle, because no difference in
mean diameter of the ovulatory follicle was detected
between groups at 20-h posttreatment. Results for both
groups are comparable to those described by Ahmadza-
deh et al. (2002) in which mean diameter of the ovula-
tory follicle was 18.6 ± 3.2 mm. Similarly, Vasconcelos
et al. (1999) measured the ovulatory follicle at the time
of the second dose of GnRH in Ovsynch-treated cattle
and reported the size of the ovulatory follicle to be
18.24 mm.
Experiment 2
Despite studies that have reported that exogenous
E2increases uterine contractions, efficiency of sperm
transport, number of sperm in the oviducts, total num-
ber of sperm retained in the female reproductive tract,
and proportion of total ova fertilized in laboratory ani-
mals and livestock (Hawk and Cooper, 1978; Hawk,
1983; Bathla et al., 1999; Orihuela et al., 1999; Langen-
dijk et al., 2002), no difference in conception rate was
detectedbetweenOVS-ECPandOVS-Cgroups.For333
dairy cows in 3 different herds, the conception rates
were similar (Table 3). Based on the number of cows
used in this experiment, a 13 percentage-point differ-
Journal of Dairy Science Vol. 89 No. 2, 2006
ence in conception rate could be detected. This sensitiv-
ity was calculated (Agresti, 1990) for α = 0.07 and β =
0.20 with anaverage pregnancy rate of 30%.The proba-
bility of detecting a smaller difference in conception
ratebetweentreatmentsinthisexperimentwaslimited
by the number of cows enrolled.
In contrast, Cerri et al. (2004) reported that use of 1
mg of ECP to induce ovulation as part of a timed AI
protocol improved conception at first postpartum in-
semination in dairy cows. It is possible that increased
conception rates observed for cows in the Heatsynch
protocol are the result of prolonged exposure to greater
concentrations of estradiol during proestrus (Cerri et
al., 2004). It has been proposed that estradiol during
proestrus influences sperm transport and might inhibit
PGF2αsecretion in the subsequent estrous cycle (Mann
and Lamming, 2000).
Serum E2for the OVS-ECP group was greater during
the first 12 h posttreatment (Figure 1). Nevertheless,
no difference in conception rate was detected between
treatments in Experiment 2. One possible explanation
forthesimilarityinconceptionratebetweentreatments
was the timing of AI in relation to ECP administration
and the systemic E2profile. Cows in Experiment 2 were
inseminated 22 to 24 h after ECP, and not likely during
the time of maximally elevated serum E2(Figure 1).
Therefore, although the OVS-ECP cows were exposed
to greater serum concentrations of E2than OVS-C cows
during the first 12 h posttreatment, by the time of AI
serum E2concentrations in most cows in the OVS-ECP
group had likely declined to concentrations similar to
the OVS-C group. Furthermore, it is likely that sperm
capable of fertilization did not reach the ampullary-
isthmus junction (site of fertilization) until 29 to 32 h
posttreatment (on average), a time when the 2 groups
were likely not different with regard to serum E2. In a
similar experiment using beef cattle, Ahmadzadeh et
al. (2003) reported that conception rate tended to im-
prove (68 vs. 57%, for OVS-ECP compared with OVS-
C groups, respectively) when AI was performed 6 to 8
h after GnRH + ECP, which corresponds to the time of
maximally elevated serum estradiol in OVS-ECP cows
in the current study. This may indicate that greater E2
in the OVS-ECP when timed AI was performed en-
hanced uterine activity, sperm transport, or fertiliza-
tion rates as previously suggested (Hawk and Cooper,
1978; Bathla et al., 1999; Langendijk et al., 2002).
CONCLUSIONS
Treatment with ECP at the time of the second dose
of GnRHin the Ovsynch protocolsuccessfully increased
serum E2in dairy cows. However, incorporating ECP
into the Ovsynch protocol had no effect on conception