Small Ruminant Research 82 (2009) 152155

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Small Ruminant Research

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Short communication

Effect of a single subcutaneous injection of melatonin on estrous

response and conception rate in goats
Sudhir Kumar, G.N. Purohit
Department of Animal Reproduction, Gynaecology and Obstetrics, College of Veterinary and Animal Sciences, Rajasthan Agricultural University, Bikaner
334001, Rajasthan, India

a r t i c l e

i n f o

Article history:
Received 13 August 2008
Received in revised form 9 February 2009
Accepted 9 February 2009
Available online 14 March 2009
Keywords:
Estrus
Goat
Melatonin
Pregnancy

a b s t r a c t
The objectives of this study were to evaluate the effect of a single subcutaneous injection
of melatonin, on estrous induction and conception rate during the non-breeding season
at different times of the year. In Experiment 1 the melatonin powder was dissolved in an
oily vitamin A, D, E solution and injected subcutaneously randomly to goats in two dose
treatment groups of 20 mg (MLT-20; n = 20) and 40 mg (MLT-40; n = 20) in January (winter).
Twenty does were injected with 1 ml vitamin A and served as the control. In the MLT-20
treated goats 70% of the does were in estrus within 20.0 2.0 days, whereas in the MLT40 group 85% of the does were in estrus within 15.5 2.5 days and 100% and 80% of the
does, respectively, conceived on mating with the bucks. Only 10% of the untreated control
does exhibited estrus, but none conceived. The breeding season was thus initiated earlier
by 11.5 months in the treated goats. In Experiment 2, goats were treated with similar
MLT-20 and MLT-40 treatments in May (spring), with 20 goats in each treatment group
and 20 control goats. The proportion of goats that responded to the melatonin treatments
was 80% and 90% in the MLT-20 and MLT-40 treatments, respectively, with no signicant
differences recorded regarding the estrous response. However, in the MLT-20 treatment
group the estrous induction interval was signicantly longer (P < 0.05), compared to the
MLT-40 treatment. The pregnancy rates were not signicantly different for the MLT-20 and
MLT-40 treatments, with melatonin resulting in signicantly higher pregnancy rates than in
the control (88.4% versus 33.3%) and the breeding season initiated 2 months earlier. It could
be concluded that a single subcutaneous injection of melatonin can initiate the breeding
season (irrespective of the season of the year) earlier by 12 months in goats and this could
be advantageous when using accelerated breeding systems.
2009 Elsevier B.V. All rights reserved.

1. Introduction
Seasonality in small ruminants represents a major
limitation to farmers and the milk and meat industries
(Chemineau et al., 1995). Goats in the temperate zones
tend to be seasonal breeders, and as a result does breed
between August (end of summer) and March (onset of
spring) (Amoah et al., 1988). These seasonal sexual activ-

Corresponding author. Tel.: +91 151 2202396.

E-mail address: gnpvog@yahoo.co.in (G.N. Purohit).
0921-4488/$ see front matter 2009 Elsevier B.V. All rights reserved.
doi:10.1016/j.smallrumres.2009.02.005

ity changes are more pronounced in the female than the

male, with a cessation of cyclic ovulatory activity occurring
during the anestrous season in the more seasonal breeders.
So, for example in the Alpine goat, ovarian activity ceases
in March (spring) and resumes in September (autumn)
(Chemineau et al., 1991). For goats raised in Rajasthan
(India) maximum estrus and breeding occurs during the
rainy season (JuneJuly) (Gujar et al., 2007). Studies conducted in Georgia involving seven goat breeds revealed that
most goats breed seasonally, commencing approximately
in late June (summer) and reaching a peak in September
to November (autumn) (Amoah et al., 1996). For the Sirohi

S. Kumar, G.N. Purohit / Small Ruminant Research 82 (2009) 152155

goat breed, three seasons of breeding have been reported

namely spring (FebruaryMarch), summer (MayJune) and
autumn (SeptemberOctober) (Misra et al., 1983). Photoperiod appears to be the main environmental cue responsible
for the seasonality in sexual activity of sheep and goats
(Malpaux et al., 1999).
Goats can be induced to breed outside the natural breeding season (Amoah and Gelaye, 1989). The maintenance
of a constant winter milk supply and the production of
meat from goats for seasonal speciality markets are two
main reasons for the out of season breeding of goat (Haibel,
1990). The various strategies to induce out of the season breeding outside the natural breeding season in goats
include, manipulation of daylight length (Chemineau et
al., 1986; Malpaux et al., 1999) and the administration of
gonadotrophins following progestogen priming with either
intravaginal sponges or subcutaneous implants (Haibel,
1990). Controlled breeding systems requiring the regression of an active CL, will not be effective outside the
natural breeding season (Wildeus, 1999). Problems in the
manipulation of daylight length on most farms have created increased interest in the exogenous administration of
melatonin (as an oral administration, or a subcutaneous
implant) (Chemineau et al., 1995). It has been demonstrated
that ewes receiving a melatonin implant resume ovulatory
activity 1 month earlier than the controls (Ronayne et al.,
1989). As commercially available melatonin implants are
difcult to procure in any location in India as an alternative, it was envisaged to evaluate the effect of a single
subcutaneous injection of melatonin in oil (Chemineau et
al., 1995). Thus this study used different doses of melatonin
at different times of the year to test the does reproductive
response.
2. Materials and methods
The trials were conducted during two non-breeding season months
(January, winter and May, spring, 2006). Non-pregnant does of the Bikaner
breed (aged between 2 and 3 years, without any reproductive problems)
were selected and used in the trials during January. For the trials during
May, Sirohi goats of the same age (23 years) raised on a commercial goat
farm (Madhu Farms, Rajasthan, India) and farmers goats of the Bikaner
breed were included.
2.1. Melatonin treatment
Crystalline melatonin powder (Code M5250, Sigma Chemicals, USA)
was dissolved in a commercially available Vitamin A, D, and E injection
(Vetade Sarabhai Zydus, India). Vitamin A (as palmitate) I.P. 250,000 IU,
Vitamin D3 , I.P. 25,000 IU, Vitamin E (as d-alpha-tocophenyl-acetate I.P.)
100 IU, benzyl alchohol B.P. 2% (w/v) and sesame oil U.S.P Q.S per ml in
quantities sufcient to make a nal concentration of 20 mg/ml (MLT-20)
or 40 mg/ml (MLT-40). Once dissolved, the suspension was used on the
same day.
2.2. Experiment 1
Bikaner does were randomly allocated to the subcutaneous treatment
with 1 ml of either MLT-20 or MLT-40 (20 does per treatment) in January
(winter), 2006. An equal number of goats (n = 20) were treated with only
1 ml vitamin A, D, E and served as the controls. One week after the single melatonin treatment, daily teasing with the aid of vasectomised bucks
and transrectal ultrasonography (using 5.0 MHz Linear array probe, BPL,
India) was performed to record the follicular activity on the ovaries and
the occurrence of estrus. When in estrus, does were mated with bucks of
proven fertility. Pregnancy diagnosis was performed by transrectal ultrasonography at weekly intervals, starting from day 15 following mating

153

to day 30thereafter, at fortnightly intervals until day 60. The number of

kids born per goat and the abortions, if any, were recorded. The pregnancy
rates were calculated as the actual kiddings (including abortions).
2.3. Experiment 2
Bikaner does were treated similar to Experiment 1, with the same
treatment and following the same protocol until kidding. The Sirohi goats
originating from the farm were also treated with the same melatonin treatments during May (spring) 2006. The estrous response, time to estrous
induction and the pregnancy rates were recorded in both the trials as
previously described. The sensitivity, specicity and positive and negative
predictive values following ultrasonography for pregnancy diagnosis were
determined as described by Karen et al. (2001).
The estrous response and pregnancy rates between the treatments and
control for the two experiments were compared using the z test. The time
to estrous occurrence (estrous induction interval) between the treatments
and control was compared by the Tukey t test whereas, comparisons for
the same treatments between the farm and Bikaner goats were compared
by analysis of variance (Snedecor and Cochran, 1989).

3. Results
3.1. Experiment 1
During January (winter), treated does experienced
estrus within 20 days of treatment with melatoninwith
a signicantly (P < 0.05) shorter response period recorded
when 40 mg was injected, compared to 20 mg. The pregnancy rates however, were higher in the MLT-20 group,
compared to the MLT-40 group and pregnancy rates were
signicantly (P < 0.05) higher in both treatments compared
to the controls (Table 1). Ovarian follicular growth was evident in goats that responded to the treatment, with follicles
of 69 mm being recorded on the day of estrus. For the 40day observation period only 10% of the untreated or control
goats exhibited overt signs of estrus, but none of the does
conceived. Of the total 28 goats that conceived in Experiment 1, one abortion in MLT-40 groups was recorded and
38 kids were born (ve goats delivered twins and two goats
delivered triplets).
3.2. Experiment 2
During May (spring) similar to Experiment 1, a high
proportion of the treated does responded to the melatonin treatment (Table 2). However, contrary to Experiment
1, a signicantly (P < 0.05) longer reaction period was
required for estrous induction in the MLT-20 treated group
whereas, a signicantly shorter period was required for
estrous induction in the MLT-40 treated animals. The time
to estrous induction revealed no signicant differences
between the goats from different sources for the same
treatments, however, the effect of treatment was highly
signicant (P < 0.05). The estrous induction interval in the
MLT-20 and control goats was not signicantly different.
However, the number of Bikaner and farm goats demonstrating estrus was too small, compared to the treated goats.
The induced estrous period for goats in Experiments 1 and
2 varied between 24.5 1.5 and 40.6 2.06 h but, were
not signicantly different, although a longer period was
recorded during Experiment 1 and in the farm goats.
The sensitivity, specicity, positive and the negative
predictive value of ultrasonography in the diagnosis of

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S. Kumar, G.N. Purohit / Small Ruminant Research 82 (2009) 152155

Table 1
Estrous response, mean (S.E.) estrous response interval and pregnancy rate in goats treated with a single SC melatonin injection during January (winter).
Treatment

No. of goats treated

MLT 20
MLT 40
Control

20
20
20

abc

Estrous response (%)

14 (70%)a
17 (85%)a
02 (10%)b

Estrous induction interval (days)

Pregnancy rate (%)

20.0 2.0a
15.5 2.5 b
30.0 1.0c

14 (100%)a
14 (82.35%)a
0 (0%)b

Values with different superscripts in the same column are signicantly different.

Table 2
Estrous response, mean (S.E.) estrous response interval and pregnancy rate in goats treated with a single SC melatonin injection during May (spring).
Treatment

No. of goats treated

Estrous response (%)

Estrous induction interval (days)

Pregnancy rate (%)

Farm
Goats

MLT 20
MLT 40
Control

20
20
20

16 (80%)
18 (90%)a
04 (20%)b

27.18 0.12
10.89 0.44b
27.0 2.0a

14 (87.5)a
15 (83.3)a
02 (50)b

Bikaner
Goats

MLT 20
MLT 40
Control

20
20
20

18 (90%)a
17 (85%)a
02 (10%)b

29.5 1.08a
10.64 0.41b
28.0 1.13a

16 (88.8)a
16 (94.1)a
0 (0.0)b

ab

Values with different superscripts in the same column are signicantly different

pregnancy were 91.3%, 63.63%, 91.3% and 63.63% at 1522

days following mating. Pregnancy rates reached 97.8%,
100%, 100% and 91.6% at 45 days of gestation. The pregnancy
rates were signicantly higher (P < 0.05) in all treatments,
compared to the controls.
In Experiment 2, 2 of the 29 pregnant farm does aborted
at 3.54 months, and a total of 43 kids were born (8 goats
delivered twins and 4 delivered triplets), whereas 3 of the
32 Bikaner pregnant goats aborted, and 34 kids were born
(3 goats delivered twins and 1 delivered triplets).
4. Discussion
During the present study estrus could be successfully
induced by melatonin treatment, both during January and
May (winter and spring) in does raised by breeders and
those kept on a farm. The type of treatment used in this
study was different from a large number of previous studies on sheep and goats that used the oral feeding or ear
implants of melatonin (Wheaton et al., 1990; Wuliji et
al., 2003), intravaginal implants (Rajkumar et al., 1989)
or commercially available SC implants (Chemineau et al.,
1993, 1995; Brackel-Bodenhausen et al., 1994; Du Preez et
al., 2001; Zuniga et al., 2002; Pellicer-Rubio et al., 2007).
In several of the previous studies melatonin implants, or
short days were preceded by long days (7090 days) and
this was considered essential for the induction of estrus
during the non-breeding season (Chemineau et al., 1988,
1995; Brackel-Bodenhausen et al., 1994). However, a few
studies did not consider this to be essential (Arendt et al.,
1983; Chemineau and Malpaux, 1998; Chemineau et al.,
2004; Papachristoforou et al., 2007). The estrous induction treatment with the aid of melatonin during May in the
present study, was preceded by natural long days. Goats
treated during January on the other hand were denitely
not exposed to long days and the response in these does
using a single melatonin treatment is contrary to previous reports. March (onset of spring) is the supplementary
breeding season for goats in the area where the trials
were conducted. August to September (onset of autumn)
being the main breeding seasonhence treatment during

January advanced the breeding season by a shorter time,

compared to treatment during May. Advancement of the
breeding season by 11.5 months following photoperiod
manipulation, or melatonin treatments have previously
been demonstrated in sheep and goats (Wheaton et al.,
1990; Chemineau et al., 1993, 1995; Sempere et al., 1995;
Traldi et al., 2000; Zuniga et al., 2002). The estrous response
obtained during the present study varied between 70%
and 90%. Previous studies have recorded a 100% estrous
response (Rajkumar et al., 1989; Zuniga et al., 2002). The
double dose of melatonin (MLT-40) induced estrus earlier
both in Experiments 1 and 2, however, the subsequent pregnancy rates were higher in the MLT-20 treatment group.
The proportion of control does in Experiment 1, that came
into estrus was too low and only 50% of the farm goats
conceived. Melatonin treatments are known to increase
the fertility (Rajkumar et al., 1989; Sempere et al., 1995)
and fecundity (Wuliji et al., 2003; Papachristoforou et al.,
2007). The higher twinning rate achieved on the farm could
probably be ascribed to better nutrition. In a previous
study the kidding rate recorded was 70.8% in does treated
with melatonin compared to 37.5% in untreated controls
(Papachristoforou et al., 2007).
Likewise, the pregnancy rates were 90%, when ewes
treated with melatonin implants were mated to rams,
while, melatonin also increased the number of lambs born
per ewe or the fecundity (Gates et al., 1998). The mechanism
of action of melatonin appears not to be fully understood,
however it is known to suppress prolactin secretion (Wuliji
et al., 2003). Seasonal differences in the proles of circulating melatonin release during estrus are probably related
to the estradiol and thyroxine secretion (Blaszczyk et al.,
2004). An early response to estrus during the present study
could probably be ascribed to a low inherent level of melatonin. It has previously been shown that does treated with
lower melatonin levels showed an onset of cyclic ovarian
activity about 12 days after the introduction of the bucks,
while the other group only started 22 days later (Carcangiu
et al., 2005). It would thus appear that melatonin acts
directly on the GnRH neurons, but it appears to involve a
complex neural circuit of interneurons that include at least

S. Kumar, G.N. Purohit / Small Ruminant Research 82 (2009) 152155

dopaminergic, serotoninergic and the excitation of aminoacidergic neurons (Malpaux et al., 1999).
Estrus occurred earlier in the 40 mg treatment group
in the present study. Waller et al. (1988) recorded estrous
induction within 1215 days following the last treatment,
while Rajkumar et al. (1989) found estrus to occur only after
29 days. Commercially available melatonin implants maintain the melatonin release for 7090 days (Chemineau et
al., 1995). No comment on the availability and half-life of
melatonin during the present study could be made. It is
speculated that melatonin crystalline powder dissolved in
an oily base and given subcutaneous to does can induce
a fertile estrus in goats 12 months ahead of the natural
breeding season. The ease of administration and the lower
frequency of administration could be useful for out of season goat breeding.
Acknowledgement
The authors are highly thankful to the manager and
owners of Madhu farms for permission to conduct this
study.
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