Ovarian Responses to hCG, GnRH, and Progesterone Supplementation
in Lactating Dairy Cattle
Alicia Lloyd
Introduction
This experiment is a subset of a larger experiment that is attempting to improve
success rates of artificial insemination. To increase conception rates, we attempted to
increase endogenous post-insemination concentrations of progesterone (P4) in lactating
dairy cattle. There are a number of reasons exit for which we hypothesized that
increasing concentrations of P4 would impact conception rates.
Conception failure is coincident with less than normal concentrations of P4 as early
as 6 days after AI (Thatcher et al., 2001). Thus, if concentrations of P4 might be increased by
day 6, a decrease might occur in rates of embryonic death. In addition, blood P4 (timing
and magnitude) generally reaches greater concentrations earlier in pregnant than
nonpregnant cows suggesting that earlier exogenous increase in P4 may spare some
embryo loss... Embryo development is related to concentrations of P4 and the ability of
the conceptus to secrete the antiluteolytic hormone, interferon-t (Mann et al., 1999). More
exposure to P4 by the embryo may increase its chances of secreting interferon-t and thus
survive. Exogenous P4 can stimulate embryo development (Thatcher et al., 1994), so by
manipulating post-insemination concentrations of P4, conception rates may ultimately
improve.
We stimulated P4 early after AI by: inducing ovulation of a secondary corpus luteum
(CL) or multiple CL by injecting gonadotropin-releasing hormone (GnRH) or human
chorionic gonadotropin (hCG) because they either cause luteinizing hormone (LH)
release or mimic LH bioactivity, respectively. We hypothesize that more luteal tissue, or
more luteal volume might result in more biosynthesis of P4.In addition, we provided
supplemental progesterone (via an intravaginally placed P4-releasing CIDR insert). The
term CIDR is an acronym for controlled internal drug release.
Treatment of lactating dairy cows on day 5 after AI with 3,300 i.u. of hCG increased
conception rates at 28, 45, and 90 days after first postpartum AI (in cooler weather only).
Injection of hCG also improved conception rates in cows losing body condition between
AI and day 28 after AI. Furhter, hCG induced accessory corpus luteum (CL) in 86% of
cows and increased concentrations of blood P4 by 4 mg/mL.
We choose to use CIDR inserts as a source of exogenous progesterone because
treatment of lactating dairy cows with a CIDR between days 5 and 12 or days 12 and 19
increased conception rates, (but blood P4 is only increased in the earlier treatment;
Robinson et al., 1989). In addition, treatment with CIDR inserts during 6 to 12 d applied
before mid cycle (beginning 4 to 9 d after AI) improved conception rates compared with
controls (74.6%; n = 466 vs. 66.1%; n = 461; Macmillan and Peterson, 1993).
Administration of GnRH or Fertagyl was our third choice for increasing
concentrations of P4. Injection of GnRH or its agonists induced accessory CL when
administered while a dominant follicle was viable on days 5 to 12 of the estrous cycle
(Pursley et al., 1995; Vasconcelos et al., 1999). A GnRH agonist (8 g of Buserelin) was equally
effective as 3,000 i.u of hCG for inducing accessory CL when injected on d 5 or 6 after
estrus (Schmitt et al., 1996).
Our hypothesis was that increasing endogenous concentrations of P4 in lactating
dairy cattle after insemination will spare embryonic loss and improve overall conception
rates.
Methods
In my project we determined ovarian follicular and luteal response to treatments. We
also detected differences in P4 concentrations resulting from changes in luteal structures.
The experiment was carried out as follows in lactating dairy cows. On 5 to 8 days
after insemination we used transrectal ultrasonography to scan the cow’s ovaries to look
for and measure number and diameters of follicles and CL. Then cows were treated with
3,300 .i.u. of hCG (Chorulon, Intervet, Millsboro, DE; n =19), 100 g of GnRH
(Fertagyl, Intervet, Millsboro, DE; n = 21), a CIDR (Pfizer, New York, NY; n =21)
containing 1.38 g of P4, or designated as an untreated controls (n = 20). Treatments were
applied equally across lactation number to reduce variability. Blood samples also were
collected at treatment and 7 days later to test for concentrations of P4. Prior to the second
blood sample 7 days after treatment ovaries were again scanned and ovarian structures
were measured... Pregnancy was assessed at 30 and 60 days after insemination.
We measured diameter and volume of the ovarian structures present on the ovaries
via ultrasound. We took into account what structures were present from one week to the
next and also noted when follicles ovulated and additional CL were formed.
Results
Figure 1 illustrates that hCG reduced (P < 0.01) the number, but not the average
diameter of follicles (Figure 2) by 7days after treatment. With approximately 20 cows in
each treatment, the sample size was not sufficient to detect potential differences in
follicle diameter.
Figure 3 illustrates that hCG was effective in ovulating smaller (P < 0.01) follicles
than GnRH. In contrast, maximum diameter of the largest follicle detected before
treatment that ovulated after GnRH and hCG did not differ (data not shown).
The proportion of cows that had CL induced by hCG or GnRH was greater (P < 0.01)
than that in controls. Proportion of cows having induced CL and treated with P4 did not
differ from controls (Figure 4).
More (P< 0.01) total numbers of CL were present 7days after treatment for cows
treated with GnRH and hCG than for controls (Figure 5).
Figure 6 illustrates total volume of luteal tissue present 7 days after treatment and
reaffirms previous results showing that hCG did ovulate more follicles, but subsequentforming CL were smaller so have less volume and area.
Concentrations of P4 (Figure 7) did not differ among treatments, despite having a
greater amount of luteal tissue in GnRH- and hCG-treated cows. Numerically there is
more luteal volume in hCG-treated cows, but statistically this did not result in greater
concentrations of P4. So the CL is probably self-regulating.
Figure 8 documents conception rates. Sample size, however, was not sufficient to
detect a difference in treatments. The trend for conception rates to be greater than those
of controls. Because we detected a numerical increase in conception rate at the first
pregnancy diagnosis, there is probably a treatment effect on embryo survival between
conception and day 30-35 of pregnancy.
Conclusions
Injections of GnRH and hCG were effective in inducing ovulation (57% GnRH and
74% hCG). Treatment with hCG, however, ovulated smaller-size follicles than GnRH. A
tendency existed for cows treated with hCG to have fewer numbers of follicles 7 days
after treatment and less total follicular area. Cows treated with hCG had more luteal
tissue 7 days after treatment compared with control cows. Interestingly concentrations of
serum P4 did not differ among treatments, even in the presence of more luteal tissue.
Although not a main objective of the study, conception rates had a tendency to improve,
this agrees with our other data spanning hundreds of dairy cows.
References
Macmillan, K.L.,Peterson A.1993.A New Intravaginal Progesterone
Releasing Device for Cattle (CIDR B) for Oestrus Synchronization,
Increasing Pregnancy Rates and the Treatment of Postpartum Anestrus.
Animal Reproduction Sci. 33:1-25.
Mann, G.E., Laning G.E., Robinson R.s>, and Walhes D.C.1999.The
Regulation of Interferon Tau Production and Uterine Hormone Receptors
During Early Pregnancy. Reprod. Fertile. Suppl.54:317-28.
Pursley, J.R., M.O. Mee, and M.C. Wiltbank.1995.Synchronization of
Ovulation in Dariy Cows using PGF2alpha and GnRH. Theriogenology
44:915-923.
Robinson N.A.,Leslie K.E.,Walton J.S.1989.Effect of Treatment with
Progesterone in Pregnancy Rate and Plasma Concentrations of Progesteron
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Santos, J.E.P., W.W. Thatcher, L. Pool, and M.W. Overton.2001.Effect
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Gonadotropin Releasing Hormone Agonist or Human Chorionic Gonadotropin
for Timed Insemination in Cattle.J.Anim.Sci.74:1084-1094.
Thatcher, W.W.,C.R. Staples, G. Danet-Desnoyers,B-Oldick, and E.J.-P.
Schmidt.1994.Embryo Health and Mortality in Sheep and Cattle. J. Anim.
Sci 72:16-30.
Thatcher W.W., A. Guzeglu, R. Mattos, M. Binelli, T.R. Hansen, and
J.K. Pru.2001.Uterine Conception Interactions and Reproductive Failure
in Cattle. Theriogenology 56:1435-1450.
Vasconcelos, J.L.M., R.W. Sulcox, G.J.M. Rose, J.R. Pursley, and M.C.
Wiltbank.1999.Synchronizatin Rate, Size of Ovulatory Follicles, and
COnception Rate after Syncronization of Ovulation Beginning on
Different Days of Estrous Cycle in Lactating Dairy Cows.Theriogenology
52:1067-1078.
Figure 2.
Figure 3
Average Follicle Diameter 7 days
after Treatment
3
mm
2
1
Minimum Follicle Diameter that Ovulated
14
16
12
14
10
12
10
8
mm
No. of Follicles > 5 mm 7 days
after Treatment
6
8
6
4
4
2
2
0
0
0
hCG
hC
C
O
Figure 5
Figure 6
Total Luteal Volume 7 days after
Treatment
Total number of CL 7 days after treatment
% of cows having induced CL
80
3.0
70
2.5
50
2.0
40
1.5
cm3
60
30
1.0
20
0.5
10
0
0.0
hCG
CON
CIDR
GnRH
hCG
N
GnRH
C
O
CIDR
Figure 7
Figure 8
Conception Rates (%)
Serum Progesterone Concentrations
ng/mL
CON
20
18
16
14
12
10
8
6
4
2
0
10
50
8
40
6
30
4
20
2
10
0
CON
CIDR
GnRH
hCG
0
CON
CIDR
GnRH
hCG
C
ID
R
G
nR
H
Figure 4
hCG
G
GnRH
N
CIDR GnRH
C
ID
R
G
nR
H
CON
hC
G
Figure 1