ELSEVIER 
BEHAVIORAL AND ENDOCRINE RESPONSES OF HAIR SHEEP EWES EXPOSED 
TO DIFFERENT MATING STIMULI AROUND ESTRUS 
R. W. Godfrey, = J. R. Collins and E.L Hensley 
University of the Virgin Islands, Agricultural Expedmant Station, St. Croix, US Virgin Islands 
Received f or publication: February 24, 2000 
Accepted: 0ctober 27, 2000 
ABSTRACT 
Hair sheep ewes were used to evaluate the influence of vadous levels of mating stimuli on 
the duration and timing of estrus and LH concentrations around estrus. Ewes were treated with 
PGF20¢ (15 mg, im) 10 d apart. At the time of the second PGF2c¢ treatment (Day 0) ewes were 
placed in groups and exposed to different types of mating stimuli. One group of ewes (n = 16) 
was exposed to an epididymectomized ram (RAM), a second group of ewes (n = 16) was 
exposed to an epididymectomized ram wearing an apron to prevent intromission (APRON) and 
a third group of ewes (n = 17) was exposed to an androgenized ovariectomized ewe (T-EWE). 
Jugular blood samples were collected from ewes at 6-h intervals through Day 5. Plasma was 
harvested and LH concentration was determined by RIA. The ewes were observed at 6-h 
intervals to detect estrus. A ewe was considered to be out of estrus when she no longer stood 
to be mounted by the teaser animal. There was no difference (P > 0.10) in the proportion of 
ewes expressing estrus (79.6%) or having an LH surge (85.7%) among the treatments. Neither 
the time to estrus nor the duration of estrus were different (P > 0.10) among APRON, RAM or 
T-EWE groups (41.6 + 3.8 vs 43.6 + 3.6 vs 46.1 + 3.6 h, respectively, and 26.5 + 2.2 vs 24.8 + 
2.3 vs 30.5 ± 2.2 h, respectively). The time to LH surge was similar (P > 0.10) among APRON, 
RAM and T-EWE groups (51.2 + 4.5 vs 51.2 + 4.7 vs 52.7 + 4.5 h, respectively). The 
magnitude of the LH surge was similar (P > 0.10) in the T-EWE, APRON and RAM ewes (99.7 
+ 4.9 vs 87.2 ± 4.9 vs 85.8 ± 5.0 ng/mL, respectively). The time from estrus to the LH surge 
was not different (P > 0.10) among APRON, RAM or T-EWE ewes (10.1 ± 2.2 vs 9.8 ± 2.3 vs 
11.6 ± 2.3 h, respectively). These results show that the expression and duration of estrus are 
not influenced by different types of mating stimuli in hair sheep ewes. In addition, the timing 
and the magnitude of LH release does not appear to be influenced by mating stimuli around the 
time of estrus. 
@ 2001 by Elsevier Scnence Inc 
Key words: sheep, estrus, LH, behavior, ram 
Acknowledgments 
Appreciation is expressed to Victor Callas, Jr. and Willie Gonzales for assisting with animal 
care and management, and Dr. J.E. Wheaton, University of Minnesota, for conducting the 
LH RIA. 
"Correspondence and reprint requests: RR 2 Box 10,000, Kingshill, St. Croix, US Virgin 
Islands 00850; email: rgodfre@mail.uvi.edu. 
Theriogenology 55:877-884, 2001 
0093-691X/01/$-see front matter 
O 2001 Elsevier Science Inc. 
PIh S0093-691X(01)00450-2 
878 
Theriogenology 
INTRODUCTION 
The ram effect is well documented in sheep and was shown to be useful for inducing estrous 
cyclicity in anestrous ewes (6, 16). The presence of the male around the time of estrus may 
also have some impact on the physiology of the female, although reports describing this effect 
in sheep are limited in number. The presence of a ram around estrus was shown to increase 
ovulation rate (8), to increase conception out of season (16) and to enhance P4 secretion (5). 
The presence of the boar at mating was shown to enhance LH levels in sows and mating with a 
stedle boar hastens ovulation (14). With the increasing use of estrus synchronization and 
artificial insemination (AI) in sheep the effect of the ram around the time of estrus is being 
investigated. When using synchronization and AI it may be beneficial, in some instances, to not 
have a ram with the ewes. The cost of producing and keeping a sterile ram for estrus detection 
may be prohibitive for some producers. The use of androgenized females for estrus detection 
is another option available. This project was designed to evaluate the effect of different types of 
mating stimuli around the time of a synchronized estrus on the behavior and endocrine 
response of hair sheep ewes in the tropics. 
MATERIALS AND METHODS 
Animals and Treatments 
Multiparous St. Croix White hair sheep ewes (n = 49) were given PGF2c¢ (15 mg im, 
Lutalyse®, Pharmacia & Upjohn Co., Kalamazoo, MI, USA) 10 d apart to synchronize estrus. 
At the time of the second PGF2cz treatment (Day 0) the ewes were placed in treatment groups. 
One group of ewes (n = 16) was exposed to an epididymectomized ram equipped with a 
marking hamess (RAM). The second group of ewes (n = 16) was exposed to an 
epididymectomized ram weadng an apron to prevent intromission (APRON). The apron was 
constructed using denim matedal attached to a modified ram marking harness. The apron 
completely covered the abdomen of the ram and prevented intromission when mounting ewes. 
Both rams had been used previously to detect estrus and possessed adequate libido. The third 
group of ewes (n = 17) was exposed to an ovadectomized androgenized ewe wearing a 
marking hamess. The ewe was given 100 mg of testosterone propionate in corn oil (25 mg/mL 
im) at 3-d intervals for 2 wk, and then once weekly for the duration of the study. The libido of 
the androgenlzed ewe was evaluated before use in the study by exposing the ewe to a group of 
ewes, some of which were in estrus, and observing which ewes were mounted. A ram of 
known libido was then exposed to the same ewes and observed for mounting. The 
androgenized ewe was able to identify the ewes in estrus with the same degree of accuracy as 
the ram. The 3 groups of ewes were kept in separate pens (6.1 m x 6.1 m) with an empty pen 
between each group. This prevented physical contact among animals in different groups but 
did not prevent visual or possible olfactory stimuli. While in the pen, the sheep were fed a 
pelleted ration and had ad libitum access to guineagrass hay, water and mineralized salt. 
Beginning on Day 0 each group of ewes was observed at 6-h intervals for mounting or 
crayon marks from the stimulus animal. The onset of estrus was determined as the time when 
a ewe was first observed to stand when mounted. The duration of estrus was determined as 
the time interval between the onset of estrus and when a ewe no longer stood to be mounted. 
Theriogenology 
879 
Blood Sampling and Hormone Assays 
Jugular vein blood samples were collected at 6-h intervals from all ewes beginning at the 
time of the second PGF2c¢ injection and ram introduction on Day 0 and continuing through 96 h. 
Plasma was harvested from all blood samples and stored at -20°C. Plasma LH was measured 
in all samples using a double-antibody RIA (2, 4). The intra- and inter-assay CV for the LH RIA 
were 4.4 and 11.2%, respectively. Sensitivity of the assay was 2.0 ng/mL. 
Statistical Analyses 
Data were analyzed using General Linear Models (GLM) procedures of SAS (13). 
Independent variables were the three treatment groups. Dependent variables analyzed were 
time to estrus, duration of estrus, time to preovulatory LH surge (maximal LH concentration) 
and magnitude of the LH surge. Preovulatory LH surges were identified in individual ewes after 
plotting their LH concentrations over time. Cumulative percentages of ewes in estrus in each 
treatment were analyzed using the CATMOD procedure using treatment, time after treatment 
and their interaction in the model (13). Results are expressed as least squares means + SEM. 
RESULTS 
The proportion of ewes exhibiting estrus was similar (P > 0.10) among ewes in the RAM, 
APRON and T-EWE groups (Table 1). The time to estrus after the second PGF2c¢ treatment 
and the duration of estrus were similar (P • 0.10) among the treatment groups. The cumulative 
percentage of ewes in estrus after the second PGF2c¢ was not different (P • 0.10) among 
treatments (Figure 1). 
Table 1. Time to onset of estrus, duration of estrus and magnitude of the LH surge in St. 
Croix White ewes exposed to vadous male stimuli 
Male Stimuli 
Control ram 
Apron ram 
Androgenized ewe 
Time to estrus (h) ° 
43.6 + 3.6 b 
41.6 + 3.8 
46.1 + 3.6 
Duration of estrus (h)" 
24.8 + 2.3 
26.5 + 2.2 
30.5 + 2.2 
Time to LH surge (h) = 
51.2 + 4.7 
51.2:1:4.5 
52.7 + 4.5 
Magnitude of LH surge (ng/mL) = 
85.8 + 5.0 
87.2 + 4.9 
99.7:1:4.9 
Proportion (%) of ewes 
Exhibiting estrus 
81.2 
81.2 
76.5 
Having an LH surge 
81.2 
87.5 
82.3 
Number of ewes in group 
16 
16 
17 
"Checked at 6-h intervals after second PGF2c¢ injection and stimulus animal introduction. 
b Least squares mean + SEM. 
c Measured in blood samples collected every 6 h after second PGF2~ injection and 
stimulus animal introduction. 
Neither the time to LH surge after introduction of mating stimulus animal (Day 0) nor the 
magnitude of the LH surge were different (P • 0.10) among the treatment groups (Table 1). The 
time of the LH surge ranged from 24 to 66 h in ewes exposed to the control ram, 18 to 78 h in 
ewes exposed to the androgenized ewe and 12 to 96 h in ewes exposed to the apron ram 
(Figure 2). In ewes exposed to the apron ram, 78.6% of ewes having an LH surge had it within 
880 
Theriogenology 
1 SD of the mean. In ewes exposed to the control ram, 69.2% of ewes having an LH surge had 
it within 1 SD of the mean. In ewes exposed to the androgenized ewe, 64.3% of ewes having 
an LH surge had it within 1 SD of the mean. 
¢R 
2 
._.R 
~e 
100- 
9O 
80 
70 
60 
5o 
40 
30 
20 
10 
0 
--II-- Control ram 
Apron ram 
Androgenized ewe 
I 
i 
I 
i 
i 
| 
I 
I 
I 
i 
I 
m 
I 
0 
12 
24 
36 
48 
60 
72 
Time alter PGF2c¢, h 
Figure 1. Cumulative percent of St. Croix White ewes exhibiting estrus after PGF2~¢ treatment 
and exposure to either a stedle ram, a stedle ram equipped with an apron or an 
androgenized ovadectomized ewe. 
Of the 39 ewes that exhibited estrus, 97.4% also had a detected LH surge, but only 30% of 
the 10 ewes that did not exhibit estrus had a detectable LH surge (P < 0.001). There was no 
difference (P > 0.10) among treatments in the proportion of ewes exhibiting estrus, having an 
LH surge, or exhibiting estrus and having an LH surge (Table 2). The proportion of ewes in the 
RAM, APRON and T-EWE groups that did not express estrus and had no detected LH surge 
was not different (P > 0.10). 
Table 2. Proportion of ewes in each treatment group that expressed estrus or had a 
detectable LH surge. 
Ewes that had: 
Estrus and 
Treatment 
LH surge 
Estrus only 
LH surge only 
Neither 
Control ram 
81.3' (13) 
0 (0) 
18.7 (3) 
0 (0) 
Apron ram 
81.3 (13) 
0 (0) 
12.5 (2) 
6.2 (1) 
Androgenized ewe 
70.7 (12) 
5.g (1) 
11.7 (2) 
11.7 (2) 
'Percentage of ewes in each group. The number in parenthesis is the actual number of 
ewes. 
Theriogenology 
881 
100 
.j 
8O 
E 
r- 
60 
,--I 
40 
2O 
0 
Control ram 
| 
• 
• 
• 
Ii 
i 
• 
ii 
• 
= 
i 
i 
120 
100 
80 
60 
4o 
20 
Apron ram 
a 
I 
120 
120 
t00 
6O 
,o 
20 
0 
I 
• 
• 
• 
I 
• 
l 
~V" 
I 
= 
0 
24 
48 
72 
96 
Time after PGF2~, h 
I 
120 
Figure 2. Concentration of LH in St. Croix White ewes after PGF2a treatment and exposure to 
either a stedle ram, a stedle ram equipped with an apron or an androgenized 
ovariectomized ewe. 
882 
Theriogenology 
DISCUSSION 
In the present study the vadous mating stimuli did not influence the timing of behavioral or 
endocrine events around estrus in the ewes. In a study by Romano (9) it was reported that 
does had a longer period of estrus when they were mounted but not served by a buck in 
comparison to does that were serviced by the buck at least once. In another study, Romano et 
al. (12) reported that does that were only mounted, but not serviced, during the first 12 h of 
estrus had a shorter duration of estrus than does serviced during this time period. Both of 
these studies disagree with the data being presented in the present study where there was no 
difference in the duration of estrus in the treatment groups (Table 1). The ram with the apron 
and the androgenized ewe were able to mount the ewes but there was no vaginal penetration 
and this is similar to the groups in the previously mentioned studies (9, 12) where the buck was 
only permitted to mount the does but not to service them. Both of these studies (9, 12) were 
performed dudng the natural breeding season of the goats, and since hair sheep in the tropics 
do not exhibit seasonal patterns of estrous cycles (1, 15) it is unlikely that season is responsible 
for the difference in results. In a subsequent study (10) it was found that service by the buck or 
mechanical stimulation of the cervix shortened the duration of estrus in comparison to natural 
service by the buck or the placement of seminal fluid in the cervical os. This implies that there 
is some influence of the physical contact of the penis with the vagina and cervix on the duration 
of estrus in goats. This outcome was not observed in the present study, although the duration 
of estrus was numerically lower in the ewes exposed to the control ram when compared to the 
ewes exposed to the androgenized ewe. If there is an effect of penile stimulation of the cervix 
on the duration of estrus in sheep, as there appears to be in goats, it was not detected in the 
present study. 
In the present study where there was no difference in the time to estrus after 
synchronization and "male" introduction among the treatment groups (Table 1). This is in 
contrast to the study of Mellado and Hernandez (7) where exposure of does to bucks resulted 
in a shorter time to estrus in comparison to does exposed to androgenized wethers or does 
during both the breeding and the nonbreeding seasons. In another study, it was reported that 
ewes synchronized dudng the breeding season had a shorter time to estrus after sponge 
removal when they were exposed to a ram at the time of sponge removal (11). 
There was also no effect of the mating stimuli used in the present study on the timing or 
magnitude of the preovulatory LH surge in the ewes (Table 2). It was reported that in the sow 
natural mating may prolong LH surge in comparison to AI (17). The duration of the LH surge in 
the present study was not evaluated, in part because of the frequency of sampling (6-h 
intervals). In a study by Husein et al. (5) the effect of having a ram present at the time of 
pessary removal and estrus on fertility to AI was evaluated, but the conception rate after AI was 
too low (5%) to make any conclusions. The authors did report that P4 concentrations at 12 to 
14 d after the synchronized estrus were higher in ewes exposed to teaser rams at the time of 
pessary removal than in ewes not exposed to rams. They suggested that the higher P4 
concentrations were due to enhanced LH secretion during the preovulatory period in response 
to the presence of the ram (5). Fertility to natural service could not be evaluated in the present 
study because of the nature of the treatments, and AI was not used. In addition, it is not known 
if the treatments in the present study had an influence on either the timing or frequency of 
ovulation or subsequent luteal function in the ewes since these responses were not monitored 
and P4 concentrations after estrus were not determined. 
Theriogenology 
883 
The proportion of ewes in each group that exhibited estrus or had an LH surge was not 
different in this study (Table 2 and Figure 2). These results are similar to those reported by 
Mellado and Hernandez (7) in goats where exposure of does to bucks, or androgenized 
wethers or ewes had no effect on the induction of estrus. The response to synchronization in 
the present study (Figure 1) was similar to that of a previous trial conducted in our laboratory 
(3). In a subsequent study when ewes were synchronized using PGF2cz, 100% of ewes were in 
estrus by 42 h after treatment (2). When PGF2c¢ was used on our research farm as part of our 
routine breeding program, the response rate was variable, but comparable to what is being 
reported in this study. 
Based on the results of this study, hair sheep ewes will express estrus and have a 
preovulatory LH surge when exposed to either natural mating stimuli (control ram) or mounting 
only (apron ram and androgenized ewe) after synchronization treatment with PGF2c¢. There 
was also no detectable influence on the timing or magnitude of the preovulatory LH surge 
among the mating stimuli treatments. It appears that the use of an androgenized female would 
be suitable for detecting estrus with no negative influence on estrus response or LH secretion in 
hair sheep. The effect of the type of animal (intact male or androgenized female) used to 
detect estrus on subsequent fertility, after either natural mating or AI, still needs to be 
investigated. 
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Theriogenology 
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