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Theriogenology 78 (2012) 842–847

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Control of the estrous cycle in guinea-pig (Cavia porcellus)

A. Grégoirea,*, A. Allardb, E. Huamánc, S. Leóna,c, R.M. Silvac, S. Buffb, M. Berardd,
T. Jolyb
a
Institut Français d’Etudes Andines, UMIFRE17 CNRS/MAEE, Lima 18, Peru
b
Université de Lyon, VetAgro Sup/IsaraLyon, Unité ICE-Cryobio, 69 243 Lyon, France
c
CIETE - Ministry of Agriculture/La Molina, National Agrarian University, Lima, Peru
d
Institut Pasteur, Animalerie Centrale, 75 724 Paris, France

Received 31 January 2012; received in revised form 22 March 2012; accepted 24 March 2012

Abstract
The aim of this work was to look for a simple method to obtain synchronized ovulation in guinea pigs under farming conditions
while respecting animal welfare. The luteolytic activity of three different prostaglandins F2alpha (PGF2␣) analogs (D-clopros-
tenol, D,L-cloprostenol and luprostiol) and a daily treatment with oral progestagen (altrenogest) was tested successively at
different stages of the estrous cycle on the same group of females during a period of 8 mo. The estrous cycle length was not
modified by the administration of PGF2␣ analogs, whatever the stage of the estrous cycle when the treatment was initiated. Our
results led us to reject the use of PGF2␣ analog to induce practical synchronization of the estrus in this species. In females (n ⫽
29), given 15 days with altrenogest (0.1 mL po once a day), ovulation occurred 4.43 ⫾ 0.13 days after the end of the treatment.
Altrenogest treatment was followed by mating. No negative impacts of the treatment on the pregnancy rates, delivery rates and
litter sizes were observed. This standard method of guinea-pig estrus synchronization is less stressful for the animals compared
to techniques using progesterone tubing.
© 2012 Elsevier Inc. All rights reserved.

Keywords: Estrus; Synchronization; Guinea Pig; Altrenogest; Prostaglandin-F2␣

1. Introduction
The guinea-pig (Cavia porcellus), together with the
camelids and the dog, was domesticated in the Andes
[1] where it still plays an important role in highland
societies as a source of protein for many low income
indigenous people [2]. Peru is the country with the
highest population and consumption of guinea pigs.
About 16, 500 tons of meat are produced there every
year, out of a constant population of about 22 million
animals raised essentially upon family production sys-
* Corresponding author. Tel: (511) 4476070; fax: (511) 4457650.
E-mail address: anne.gregoire@gmail.com (A. Grégoire).
tems [3]. In this part of the world, guinea pigs are also
involved in many folklore traditions, exchanged as gifts
and used in traditional medicine to diagnose diseases
[4]. The conservation of native genetic diversity is a
longterm issue.
The guinea-pig has also been used commonly since
the late 18th century as a laboratory animal [5]. It
played a major role in the establishment of the germ
theory in the late 19th century, in particular through the
studies of Robert Koch on tuberculosis [6]. Despite its
important role in laboratory animal science, the use of
guinea pigs has decreased significantly since the middle
of the 20th century, mostly because mice and rats have
replaced them. In 2007, guinea pigs represented only

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 A. Grégoire et al. / Theriogenology 78 (2012) 842–847
2% of the total number of laboratory animals [7]. None-
theless, the guinea-pig is still a reference model for
very specific research, because its immune system pos-
sesses an antigen-macrophage interaction similar to that
of the human [8,9]. The relatively long gestation pe-
riod (68 days), compared to other rodents with ma-
ture central system at birth, makes it a very relevant
model for teratology studies. This characteristic is
particularly interesting for the study of deleterious
factors which act differentially with respect to the
stage of pregnancy, as do many noxious agents in
man [10]. The most common strains used in scientific
research are outbred animals, such as the albino
short-haired Duncan-Hartley guinea pig. Inbred
strains are also available, Strain 2 and Strain 13
being the most commonly used. New strains for
specific purposes are regularly established [11].
The establishment of an embryo cryobank will per-
mit the preservation of the genetic pool of both scien-
tifically valuable strains and native guinea pigs. It is
also an interesting strategy to reduce maintenance costs
related to livestock.
To produce guinea-pig embryos and to set up the
technique of embryo transfer, it is first necessary to
control the female estrous cycle. Controlling the heat
period is required to obtain embryos at morula or early
blastocyst stage and to prepare synchronous recipients
for embryo transfer. In the present work we sought
estrus synchronization through two approaches. One
was to block ovulation using a progestagen treatment,
and another was to induce estrus with a PGF2␣ analog
treatment.
A treatment to synchronize ovulation under labora-
tory conditions was proposed using a subcutaneous
implant filled with crystalline progesterone over a 4 wk
period[12]. The withdrawal of the implant induces ovu-
lation within 5 d. This treatment is difficult to set up in
farming conditions and presents ethical problems re-
garding the use of general anesthesia to insert a large
implant (1.0 cm long and 0.4 cm i.d.).
Most of the work to discover the luteolytic factor in
guinea pigs was carried out in the late 1960s and early
1970s. Blatchley and Donovan [13] established that
exogenous PGF2␣ was luteolytic in hysterectomized
guinea pigs. Poyser [14] indicated that guinea-pig uteri
are probably the major source of this luteolytic factor.
Further results [15] identified PGF2␣ as the uterine
luteolytic hormone in guinea-pig since the treatment of
females during their cycles with indomethacin (blocker
of PGF2␣ synthesis) lengthened their estrous cycles. In
1976, Blatchley and Donovan indicated that the re-
843
sponse of the corpus luteum to treatment with PGF2␣
changes during the estrous cycle. Treatment with
PGF2␣ over Days 4 – 6 or 6 – 8 of the cycle temporarily
depressed progesterone release without shortening the
life of the corpus luteum. When the drug was admin-
istered over Days 8 –10, 10 –12 or 12–14, the depres-
sion in progesterone was not followed by any recovery
(Day 1 is the day on which maximum cornification was
seen in smears) [16]. In hysterectomized guinea pigs,
the corpus luteum responds to the luteolytic action of
PGF2␣ only after Day 9 of the estrous cycle (Day 0 is
the first day when the vaginal membrane is fully per-
forated) [17].
The aim of this work was to look for a simple
method to obtain synchronized ovulation in guinea-pigs
under farming conditions while respecting animal wel-
fare. We tested the luteolytic activity of three different
PGF2␣ analogs (D-cloprostenol, D,L-cloprostenol and
luprostiol) currently used for mammal synchronization
and we tested the oral administration of a progestagen
(altrenogest) as it is used in sows and mares [18,19].
These treatments were administered to guinea pigs at
different stages of their estrous cycle.
2. Materials and methods
2.1. Animals
Forty-four multiparous female guinea pigs with nor-
mal cycles, aged between 18 and 24 mo, from the
Maria-Marcela Farm (Puente Piedra-Peru), weighing
from 1.0 to 1.5 kg, were used in this study. Females
were housed under farming conditions and were fed on
commercial pellets (vitamin C enriched) and tap water
ad libitum. To determine the day of ovulation, the
colony was checked twice a day for vaginal opening.
As perforation of at least half the vaginal membrane
was observed, vaginal smears were made at 12-h inter-
vals to determine the day of ovulation (Day 0), judged
by the first leukocytic smear following a cornified
smear [20]. The natural length of the estrous cycle in
the guinea pigs used in this study was 16.17 ⫾ 0.21 d
(observed on 83 cycles).
2.2. Treatments
In this study, four treatments to control the heat
period of the guinea-pig were tested. First, three treat-
ments with injections of PGF2␣ analogs had been per-
formed to clarify their capacity of luteolysis (D-clopro-
stenol; D,L-cloprostenol and luprostiol), and last, a
daily treatment with oral progestagen (altrenogest) was
844 A. Grégoire et al. / Theriogenology 78 (2012) 842–847
carried out to try to block ovulation. Each of these
treatments was tested successively at different stages of
the estrous cycle on the same group of females during
a period of 8 mo. To know exactly the stages of the
estrous cycle of each female when the treatments were
administered and to determine the return to estrus, the
colony was checked twice a day for vaginal opening
and ovulation was confirmed through vaginal smears.
We gave a single injection of D-cloprostenol to females
between Day 1 and Day 18 of their estrous cycle,
D,L-cloprostenol between Day 4 and Day 16, and lu-
prostiol between Day 3 and Day 12. The first adminis-
tration of altrenogest treatment was carried out in fe-
males between Day 1 and Day 14 of their estrous cycle.
We first tested the effect of D-cloprostenol (Ciclar,
Zoovet, Argentina) by i.m. injection of 7.5 ␮g/kg; 50
tests were performed. After the next return to estrus,
D,L-cloprostenol (Estrumate, MSD Santé Animale,
France) was administered by i.m. injection of 25 ␮g/kg;
17 tests were performed. Luprostiol (Prosolvin, Virbac,
France) was administered by i.m. injection of 0.75
mg/kg; 23 tests were performed. We then adapted the
progestagen treatment as described for mares. It con-
sisted of 15 d of administration of 0.1 mL (0.22 mg/kg)
Fig. 1. Effect of the PGF2␣ analogs and progestagen on the estrous cycle length and the treatment-ovulation interval.
p.o. s.i.d. (semel in die, i.e., once a day) altrenogest
(Regumate equine, MSD Santé Animale, France); 29
tests performed. The treatment-to-ovulation interval
corresponds to the interval between the single injection
and the moment of ovulation for the prostaglandins
treatments, and to the interval between the last admin-
istration of hormone and the moment of ovulation for
the altrenogest treatment.
2.3. Statistical analyses
Data were reported as means ⫾ SD. The statistical
analyses have been performed by means of linear re-
gression (Analysis Tool, Microsoft excel). The graphs
a, b, c and d in Fig. 1 represent for each treatment
(D-cloprostenol, D,L-cloprostenol, luprostiol, and al-
trenogest) the estrous cycle length (full circles) and the
treatment-ovulation interval (empty circles). For all
graphs in Fig. 1, Day 0 is considered as reference day
of previous ovulation. After altrenogest treatment, each
female was mated with males of proven fertility to
observe the litter size.
The litter size data were analyzed using Student t
test (Analysis Tool, Microsoft excel).
 A. Grégoire et al. / Theriogenology 78 (2012) 842–847
3. Results
3.1. Regarding the cycle length
For each prostaglandin treatment (D-cloprostenol;
D,L-cloprostenol and luprostiol) the estrous cycle
length was not modified by the administration of
PGF2␣ analogs (P value ⬎ 0.05): 15.76 ⫾ 0.31,
17.32 ⫾ 0.42, and 16.17 ⫾ 0.36 d for D-cloprostenol,
D,L-cloprostenol and luprostiol, respectively. In con-
trast, altrenogest treatment prevented ovulation. More-
over, as long as altrenogest was administered, the es-
trous cycle lengthened.
3.2. Regarding the treatment-ovulation interval
The duration of the interval between the end of the
treatment and the ovulation was independent of the
cycle stage when altrenogest was initiated (P ⬎ 0.05).
After the end of the treatment all the 29 females
ovulated 4.43 ⫾ 0.13 d after the last administration p.o.
of altrenogest, 93% of the females (27/29) had ovulated
by Day 5 (Fig. 2).
After the altrenogest treatment, the 29 females
with induced ovulation were mated with males of
proven fertility. The pregnancy rate estimated by
abdominal palpation after one mo was 79% (23/29),
the delivery rate was 82% (19/23) and the litter size
was 3.05 ⫾ 0.19 pups. The litter size of the treated
females was not statistically different (P value ⬎
0.05) from the litter size of untreated females from
the same farm (3.09 ⫾ 0.27 pups, n ⫽ 25). Unfor-
tunately, four females died because of uncontrollable
environmental conditions.
Fig. 2. Distribution of ovulated females after the last Altrenogest
administration.
845
4. Discussion
Although some previous papers [13–15,17,21] in-
dicated that PGF2␣ is luteolytic in guinea pigs, the
present work failed to induce ovulation when ani-
mals were given exogenous PGF2␣ analogs. The
doses administered in our work were lower than the
doses administered in former works from the 1960s
and 1970s. At that time, studies were performed with
natural PGF2␣ that required higher doses. The posol-
ogy we used in the present study was higher than the
posology used with similar synthetic PGF2␣ analogs
in cattle [22]. Even when we doubled the dose of
D,L-cloprostenol, no luteolytic effect was detected
(unpublished data). Moreover, all the studies to con-
trol the guinea-pig estrous cycle worked on hyster-
ectomized females [13,17]. Up to now, no control of
estrous cycle by PGF2␣ analogs treatment has been
described in females with intact uteri. Our results led
us to reject the use of PGF2␣ analogs to induce
practical synchronization of the estrus. Osamu Su-
zuki (personal communication, 2009) has tried to
synchronize guinea pigs with several doses of exog-
enous PGF2␣ analogs several times and it seemed to
perturb the progesterone effect, but did not stop
luteal phases.
In contrast, our results show that the administration
of 0.1 mL (0.22 mg/kg) p.o. s.i.d. altrenogest over 15 d
can induce in guinea-pig synchronized ovulation within
4 to 5 d after the end of treatment independently of the
stage of the estrous cycle when altrenogest is initiated.
As far as the length of the treatment is concerned, we
decided to administer the hormone over 15 d to avoid
the presence of residual active corpora lutea at the end
of the treatment, because the natural length of the
estrous cycle in the guinea pigs used in this study was
16.17 ⫾ 0.21 d. Moreover, the length of the treatment
should be adapted to the population with significantly
different natural cycle length.
Compared to other methods, ours was easy to im-
plement in farming conditions because there was no
need to anesthetize the animals to insert and remove
progesterone implants.
Appropriate doses of altrenogest cannot be supplied
to each animal by feed mixed with altrenogest. Because
of the alimentary behavior of the guinea pigs, the top
dressing of altrenogest is not possible in this species,
contrary to what can be performed, for instance, in pigs
and horses. Guinea-pigs’ free feeding behavior is char-
acterized by a large number of small meals with brief
periods of feeding, interspersed with non-feeding peri-
ods [23]. It is then impossible to ensure that each
846 A. Grégoire et al. / Theriogenology 78 (2012) 842–847
animal of a group housed together receives the appro-
priate dose of hormone. Though the daily oral admin-
istration of the treatment is demanding for the operator,
guinea-pigs maintain a docile disposition when handled
frequently and gently and become used to the treatment
within a few days. Compared to the subcutaneous im-
plant filled with crystalline progesterone over a 4-wk
period [12], this way of administration is less trauma-
tizing and invasive for the animal, achieving the pur-
pose of estrus synchronization while respecting animal
welfare.
Altrenogest is a safe and effective drug. The dose
of 0.1 mL altrenogest represents a high dose of
progestagen (0.22 mg/kg). It is five times the dose
used in mares, for example [24]. The toxicologic
assessment of hormonal manipulation described by
Peters [25] showed that altrenogest is neither geno-
toxic nor mutagenic. The surplus of altrenogest
seems to be eliminated in the feces [26]. In our study,
there was no negative impact of the treatment on the
pregnancy rates, delivery rates or litter sizes of the
females treated with altrenogest.
The altrenogest method of estrus synchronization
has been used with other populations of guinea pigs.
The results showed that this method should be efficient
in Peruvian guinea pigs but also in inbred strains. We
applied altrenogest treatment on inbred strains from the
Institut Pasteur used in biomedical research and we
obtained 87.5% of estrus synchronization (7 of 8 fe-
males were synchronized, 3 nulliparous and 5 multip-
arous— unpublished data).
5. Conclusions
A standard method of estrus synchronization re-
specting animal welfare has been determined in guinea
pigs. It consists of a 15-d treatment of 0.1 mL altreno-
gest p.o. s.i.d. which induces ovulation within 4 to 5 d.
This method presents a scientific interest to synchro-
nize the estrus of embryo-donor females that can be
used for biomedical research. It also represents an ad-
vance from a zootechnical point of view as it allows the
use of artificial insemination and the consecutive im-
provement of both production and sanitary control on a
farm. However, this practice is subject to the standard-
ization of the sperm collection in this species that re-
mains to be defined. As this method has been developed
in farming conditions, it is possible to use it in the field
to produce embryos from superovulated females. It will
be used to implement a cryobank of both native and
scientifically valuable guinea pigs.
Acknowledgments
The authors wish to express their gratitude to Gon-
zalo Valentin for providing animals from the Maria-
Marcela Farm (Puente Piedra, Peru), José Sarria
Bardales for lending the Small Animals Research Lab-
oratory of the La Molina National Agrarian University
(Lima, Peru) and Enrique Alvarado Malca, coordinator
of the CIETE - Ministry of Agriculture/La Molina Na-
tional Agrarian University (Lima, Peru). We wish to
thank Osamu Suzuki, Senior researcher of the Labora-
tory of Experimental Animal Models for Human Dis-
eases from the National Institute of Biomedical Inno-
vation (Osaka, Japan), for reading the manuscript and
offering valuable suggestions. We are grateful to Eddie
Maranghi for technical assistance at the Institut Pasteur
(Paris, France).
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