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Studies on the use of Ovuplant Induction
of ovulation at a precise time may be an advantage to the equine practitioner
for the following reasons: (a) to ensure ovulation occurs within 36-48 h of
mating to stallions with normal fertility and thus prevent rebreeding, (b)
appointment servicing for mares transported to stallions, (c) to ensure a single
insemination for a problem broodmare, (d) to ensure ovulation close to the time
of artificial insemination of cooled transported or frozen semen or for mares
mated to stallions with low fertility associated with poor sperm longevity, and
(e) to separate inseminations when many mares are presented in oestrus at the
same time. Initially
human chorionic gonadotrophin (hCG) was the only commercially available
preparation for induction of ovulation at 36-48 h when administered to a mare in
early oestrus with a 30-40 mm follicle. Drawbacks to the use of hCG are that it
is of human derivation which may influence purity or supply and refractoriness
has been noted after multiple treatments associated with antibody formation. An
alternative agent to hCG for the induction of ovulation such as GnRH or
analogues may be beneficial for the following reasons: (a) its smaller molecular
weight make it less antigenic thus allowing multiple administration within the
same breeding season while still reliably inducing ovulation, (b) it may be used
in alternate cycles with hCG, (c) because it is a pure synthetic product there
is no probability of viral contamination, and (d) supply would not be related to
availability of pregnant women. In
this study below we examined whether an analogue of GnRH (deslorelin-“Ovuplant”)
released from an implant in a controlled fashion over at least 72 h could
consistently induce a luteinising hormone (LH) surge capable of predictably
hastening ovulation. 1) A. O. McKinnon, A. M.
Nobelius, S. T. D. Figueroa, J. Skidmore, J. R. Vasey, and T. E. Trigg.
Predictable Ovulation in Mares Treated with an Implant of the GnRH Analog
Deslorelin. Equine.Vet.J. 25:321-323,
1993. The
experimental animals were part of a herd of 95 mares in good body condition,
kept on pasture and given lucerne hay and grain supplement as necessary. They
were studied between October 1990 and March 1991 at the Goulburn Valley Equine
Hospital. Mares
were teased daily and ultrasonography was used to monitor follicular size and to
determine ovulation and number of ovulations per cycle. Mares were examined
every 3 days during dioestrus, daily during early oestrus and twice daily (06.30
and 18.30 h) when a follicle > 30 mm in diameter was detected. Ovulation was
diagnosed by absence of a previously identified pre-ovulatory follicle (>30
mm diameter) and visualisation of the characteristic echogenic structure formed
by the collapse of the follicular walls. All cycles were induced by luteolysis
effected by administration of PGF2
on Days 7-10 after ovulation from mares that had follicles < 20 mm in
diameter. Only
61% of mares showed oestrus at some time before assignment. But 87% of mares
became oestrus at some time before ovulation. This percentage was not markedly
different from that expected and can be used to illustrate difficulties involved
with assigning mares on teasing data only. In the absence of behavioural
oestrus, assignment was accurately performed on follicle size, relaxing cervix
and presence of endometrial folds. Mean
time to ovulation in higher doses of GnRH analogue (Ovuplant) were not different
from that in hCG and both drugs significantly decreased time to ovulation
compared to controls. These results suggested that the appropriate dose and
release profile of the GnRH analogue (deslorelin) was equally efficacious in
inducing ovulation in cyclic mares when compared with hCG and were useful in
deciding if other international studies would be commissioned. Other studies
confirmed these findings and later the drug was released for commercial use. Since
the release of Ovuplant it has been widely accepted by veterinarians as a
reliable ovulating induction agent. It has saved many mares from being re-bred
when for a variety of reasons it was not possible to accurately predict the
precise time of ovulation. After
the initial release and acceptance of the breeding industry of the new drug, it
became apparent that other possible uses should be explored. Mares
in winter anoestrous have reduced gonadotropin releasing hormone (GnRH)
synthesis and storage in the hypothalamus and decreased quantities of
luteinising hormone (LH) in the anterior pituitary. Induction of ovulation in
seasonally anovulatory mares in deep winter anoestrous cannot be recommended on
commercial breeding farms due to 1) poor response to GnRH or analogues or
multiple injections of equine pituitary extracts, or 2) failure of the mare to
maintain the corpus luteum (CL) that resulted from induced ovulations. In
one study it was concluded that constant administration of GnRH agonists may induce ovulation in
mares during seasonal anoestrus; however, percentage of mares ovulating and the
lack of reproducibility of effect indicated that this approach was inappropriate
for use as a reliable method to manipulate breeding activity in commercial
broodmares. Pulsatile
administration of GnRH produced better results compared to constant infusion or
absorption. More success is obtained however, when mares enter the transition period when the combined
effects of photoperiod, nutrition and warmth, result in increased GnRH synthesis
and storage. Progression through the transition period can be monitored
hormonally or by determining the follicular population with ultrasonography.
Expected response of early or mid transition mares to constant infusion of GnRH
or analogues is 50-75% ovulation rate. Most mares will maintain the CL and late
in the transition, mares respond more predictably. The GnRH analogue Deslorelin
(“Ovuplant”) has been shown effective in increasing LH and predictably
hastening ovulation in cycling mares. Published LH profiles in cycling mares
suggest a dose dependent response with maximal effect in 12-24 hours and
duration of 3-4 days after implantation with Ovuplant. The
report below describes our experiences with experimental administration of
Ovuplant to mares in the transition period prior to their first ovulation of the
breeding season. 2) A. O. McKinnon, J. R.
Vasey, T. B. Lescun, and T. E. Trigg. Repeated use of a GnRH analogue deslorelin
(Ovuplant) for hastening ovulation in the transitional mare.
Equine.Vet.J. 29 (2):153-155, 1997. The
experimental animals were part of a herd of 55-65 mares in fair to good body
condition, kept on pasture and given lucerne hay and grain supplement as
necessary. The studies were performed between July and November in 1994
(experiment 1) and 1995 (experiment 2) at the Goulburn Valley Equine Hospital,
Victoria, Australia. Mares
were teased daily and ultrasonography was used to monitor follicular size,
determine ovulation and monitor the CL. Mares were examined every 2 days during
transition and then daily after assignment to treatment. Ovulation was diagnosed
by absence of a previously identified pre-ovulatory follicle (>30 mm
diameter) and visualisation of the characteristic echogenic structure formed by
the collapse of the follicular walls. Whenever possible mares were monitored to
determine the time of the subsequent ovulation (inter-ovulatory interval). The
experiments were conducted over two breeding seasons. Mares were not bred. Experiment 1:
In
1994, 21 early transitional mares were randomly assigned to either a control
(n=10) or treated (n=11) group on the same day (September 29). Treated mares
were implanted every other day with one implant of Deslorelin until either
ovulation had occurred or six implants had been administered. Experiment 2:
In
1995, 20 late transitional mares were randomly assigned to either control or
treatment groups after they were identified with a ³ 30 mm follicle, were in heat and had endometrial folds demonstrated
using ultrasonography. Treated mares were implanted every other day as above.
Implants were placed subcutaneously in the neck in both experiments. In
1994, utilising mares in early transition, we were able to demonstrate an
apparent effect of the implants in hastening ovulation (ovulation in <10
days) in treated mares (6/11) versus controls (0/10). However, the mean
ovulation date was not different between groups and reflected the effect of 5
mares that did not ovulate after treatment. Those 5 mares may have underwent
some form of “down regulation” in GnRH receptors as their follicular size
decreased quite clearly in comparison to control mares. This experiment
highlighted the difficulty in selecting mares randomly for treatment during the
early part of the transition period. The responses are similar to previous
reports with GnRH therapies early in the transition. In
experiment 2 in 1995, we were able to demonstrate the ability of the implants to
hasten ovulation late in the transition. The number of mares with a ³ 30 mm follicle ovulating within 3 days of treatment (8/10) suggests that
the procedure would be useful in predicting ovulation in transitional mares.
Failure of mares to ovulate during transition into the breeding season is a
major source of frustration to clients, breeding farm managers and
veterinarians. The average number of implants needed was 2.1 and this may result
in savings in decreased veterinary costs, decreased boarding charges, decreased
teasing and handling and better usage of the stallion. Each
year, one mare in the treated group became anoestrus or transitional again and
thus demonstrated ovulation may have been forced upon them prematurely. From
this perspective it would appear prudent to wait until a transitional mare has
demonstrated maturity of the reproductive axis by maintaining large (>30mm)
follicles for a few days before initiating induction of ovulation. The increased
inter-ovulatory interval detected in treated mares in experiment 2 may have been
associated with premature entry into the cyclic season or may suggest a
leuteotropic action of the implants. No progesterone measurements were taken and
would be a logical area for further investigation. In
summary, it was demonstrated that accurate, timed ovulation was achieved during
the late transition using multiple implants of Ovuplant.. This regime
is simple, may be cost effective and is expected to be useful for practicing
veterinarians. At
this stage no studies had actually looked at what the expected time to ovulation
(in hours) was after administration. This has important ramifications (see
below). We decided that we should take a close look at some of the effects that
various drugs may have on the reproductive cycle, in particular on time to
ovulation. 3)
A.O.
McKinnon, W.J. Perriam, T.B. Lescun, J. Walker, J.R. Vasey and T.E. Trigg*. Effect of a GnRH analogue
(Ovuplant), hCG and dexamethasone on time to ovulation in cycling mares. World
Equine Veterinary Review Vol 2 page 16-18, 1997. Induction
of ovulation may be an advantage to the equine practitioner for the reasons
listed above: In addition, induction of ovulation at a precise time is necessary
for oocyte retrieval in gamete intrafollopian transfer (GIFT) in vitro
fertilisation (IVF) or intracytoplasmic sperm injection (ICSI) programmes. Most
studies have examined hastening ovulation, however there appears to be little
information on delaying ovulation. Delaying ovulation would be particularly
useful when organising transport of cooled semen. One study found that by
administering the synthetic glucocorticoid,
dexamethasone (30 mg/day) in midcycle (day 10
from ovulation) that only 1 of 8 mares exhibited
behavioural oestrus, compared to 7 of 8 control mares and ovulation occurred in
1 and possibly in 2 treated mares, compared to all 8 control mares. In addition
there was a significant reduction in mean maximum luteinizing hormone (LH)
concentration and follicle size. Studies on hastening ovulation using human
chorionic gonadotrophin (hCG) began
in the 1940’s. Human chorionic gonadotrophin reliably induces ovulation
at 36-48 h when administered to a mare in early oestrus with a 30-40 mm
follicle, however some question the use of repeated hCG in consecutive cycles as
refractoriness has been noted after multiple treatments, perhaps associated with
antibody formation. A GnRH analogue, “Ovuplant”, has been effective as an
alternative for induction of ovulation in cycling mares and transitional mares.
Apparently only two studies have directly compared hCG and Ovuplant and reported
an ovulation interval of 1.88 and 1.98 days and 43 and 46.9 h respectively.
However both those and other studies have allowed an interval of 12 hours
between examinations. The
purpose of this study was to examine the effect of dexamethasone on delaying
ovulation when administered to mares with a mature follicle and in oestrous and
to more accurately characterise the time interval from administration to
ovulation with Ovuplant and hCG. Experiment 1 When
mares were detected with a follicle of ³ 30 mm, a softening cervix and endometrial folds, they were assigned to
one of three treatments 1) Treatment intramuscularly every 12 hours until
ovulation with 20mg dexamethasone (n=10 cycles), 2) Single subcutaneous
administration of Ovuplant (n=10 cycles) and 3) No treatment (n=10 cycles).
Mares were examined for ovulation every twelve hours. Experiment 2 Using
the same assignment criteria as Experiment 1, mares were treated with either
2500 IU hCG intravenously (n=26 cycles) or a single subcutaneous implant of
Ovuplant (n=26 cycles). Mares were examined with ultrasonography for ovulation
at least every 2 hours from ~ 28 h after administration. Experiment 1 Dexamethasone
did not increase time to ovulation compared to controls. Ovuplant administration
resulted in a significant decrease in time to ovulation (40.5 ± 6.2 h) compared to either controls (61.3 ± 31.4 h) (P<0.05) or dexamethasone treatment (54.6 ± 16 h) (P<0.05). Experiment 2 Time
to ovulation was significantly different (P<0.001) between hCG (35.9 h ± 3.8 h) compared to Ovuplant (40.7 ± 3.2 h) (Figure 1). Data
was removed for ovulation < 24 h from three cycles in hCG treated mares and 2
cycles in Ovuplant treated mares. Techniques
to delay time to ovulation would be of benefit to practitioners and researchers
by allowing delayed breeding or treatments. Dexamethasone was not able to delay
ovulation as used in this study. Other techniques that may be considered are
antisera against either the pituitary, follicle stimulating hormone (FSH),
oestrogen, luteinizing hormone or even GnRH, or administration of inhibin,
follicular fluid or progesterone. For any substance to be useful and practical
it would need to be effective late in the cycle, after selection of the
preovulatory follicle had occurred. The
finding of significant difference in time to ovulation of Ovuplant versus hCG
has important clinical ramifications for those needing accurate ovulation times.
At the GVEH we have used Ovuplant for breeding mares to frozen semen in the last
three breeding seasons. A total of 26 and 54 mares were bred for a total of 39
and 98 cycles in the 1995 and 1996 breeding seasons. Whenever possible if hCG
had been used the previous cycle, Ovuplant was used the next, however due to the
recognised difference in time to ovulation between the two drugs, most commonly
Ovuplant was administered. In our hands Ovuplant was administered at 6-7 P.M. on
one day and then the mare next examined at + 24 h, again at +36 h (6-7 A.M.) and
then hourly as indicated until ovulation. With this regime the mares ovulated
during hours when there were many people available to handle and examine them.
When using hCG to mimic the same examination schedule (daylight hours) the drug
had to be administered at 12 midnight and then the mares examined at +12 h, +24
h, +31 h (7A.M.) and then hourly as indicated. Later
in the breeding season it became even more critical to know the expected time of
ovulation for in vivo matured oocyte collection for ICSI experiments. In this
study, due to time constraints, mares were all subjected to follicular
aspiration at 39 hours after Ovuplant. A total of 59 mare cycles were programmed
for follicular aspiration. On 10 occasions (17%) ovulation had occurred prior to
the time of aspiration, sometimes between identification of the preovulatory
follicle and the aspiration attempt. Thirty two oocytes were recovered from 49
attempts (65%). Oocyte recovery was different ( P< 0.001 ) for matured
irregular follicles (30/38-79%) versus not matured follicles (2/11-18%). Knowing
the time of expected ovulation will have profound influences on accuracy of
programming mares to ovulate at times were personnel of maximum experience and
efficiency are available. Detected differences in time to ovulation of these two
drugs will enable clinicians to determine which compound is most appropriate to
be administered given the availability of personnel and time management.
Figure
1 Number
of mares responding (ovulating) to either hCG or Ovuplant as a function of time
(values grouped into 2 hourly increments and the curve smoothed). Acknowledgement Portions
of the studies reported above were funded by RIRDC (Australia). Summary Ovuplant
has been demonstrated to be effective to accurately predict the time of
ovulation and has resulted in an increased awareness amongst the horse breeding
fraternity that one breed per cycle is all that is necessary. We (GVEH) aim at a
number of serves per cycle of 1.05. In other words we expect that after 100 mare
cycles the stallions will have to serve 105 times. This has critical
ramifications for stallions with poor libido, low sperm numbers and shuttle
stallions with limited services per season. Using Ovuplant as outlined above
will help achieve these objectives. Adams,
G.P., Kastelic, J.P., Bergfelt, D.R. and Ginther, O.J. (1987) Effect of uterine
inflammation and ultrasonically-detected uterine pathology on fertility in the
mare. J. Reprod. Fertil. Suppl. 35,
445-454. Ginther
OJ (1992) Reproductive biology of the mare: Basic and applied aspects.
Equiservices, Cross Plains, Wisconsin, pp 1-642 Loy,
R.G. (1980) Characteristics of postpartum reproduction in mares. Vet. Clin. North Am. [Large. Anim. Pract]. 2,
345-349. McKinnon
AO (1998a) Why is my mare loosing her pregnancy?
Equine Research Seminar 119-144(Abstract) McKinnon
AO, Rantanen NW (1998b) Twins. In: Rantanen NW, McKinnon AO (eds) pp 141-156 McKinnon,
A.O., Squires, E.L., Carnevale, E.M., Harrison, L.A., Frantz, D.D., McChesney,
A.E. and Shideler, R.K. (1987) Diagnostic ultrasonography of uterine pathology
in the mare. Proc. AAEP 605-622. McKinnon,
A.O., Squires, E.L., Harrison, L.A., Blach, E.L. and Shideler, R.K. (1988)
Ultrasonographic studies on the reproductive tract of mares after parturition:
Effect of involution and uterine fluid on pregnancy rates in mares with normal
and delayed first postpartum ovulatory cycles. Journal
Of The American Veterinary Medical Association 192, 350-353. Merkt,
H., Jacobs, K.O., Klug, E. and Aukes, E. (1979) An analysis of stallion
fertility rates (foals born alive) from thebreeding documents of the Landgestut
Celle over a 158-year period. J.
Reprod. Fertil. Suppl. 73-77. Morris,
L.H.A. and Allen, W.R. (2001) Reproductive efficiency of the intensively managed
Thoroughbred. Equine Veterinary
Journal Osborne,
V.E. (1966) An analysis of the pattern of ovulation as it occurs in the annual
reproductive cycle of the mare in Australia. Aust.
Vet. J. 42, 149-154. Ricketts,
S.W. and Young, A. (1990) Thoroughbred mare fertility [letter]. Vet. Rec. 126, 68-68. Sanderson,
M.W. and Allen, W.R. (1987) Reproductive efficiency of Thoroughbred mares in the
United Kingdom. Proc.
Bain-Fallon. Mem. Lect. ,. Sydney. 31-41.
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