1. 2 4 6 8 1 0
2 0
3 0
4 0
5 0
6 0
T im e (m in )
Avg.CumulativeDuration(s)
V e h ic le C o n tro l, N = 7
E E 2 , N = 7
L o w n a tu ra l, N = 6
H ig h n a tu ra l, N = 7
Gabrielle Mercado and Lilah Glazer
Effects of acute exposure to natural estrogen-
containing soy extract on the aggressive and courtship
behaviors in adult male zebrafish (Danio rerio)
Introduction
The focus of traditional sewage systems is the disinfection of waste water
before returning it back to the natural environment, while certain chemicals
are not eliminated. Wild fish populations are vulnerable to many contaminants
that are discharged from sewage systems and find their way into aquatic
habitats. Endocrine disruptors are environmental pollutants that adversely
affect hormonal systems within the body.
One such chemical is the synthetic estrogen 17α-ethinylestradiol (EE2), an
active component in oral contraceptives, which has been shown to exert
harmful effects on the health, embryonic development and reproductive
success of a number of fish species. In adult male zebrafish, acute exposure to
EE2 resulted in reduced aggressive and courtship behaviors1, indicating
impairment in competitive and reproductive success.
A global increase in the consumption of soy-containing supplements is
occurring due to advertisement of their potential health benefits. Soy contains
phytoestrogens which are a diverse group of naturally occurring compounds
that are structurally similar to estradiol, and may therefore potentially act as
endocrine disruptors with similar effects to those observed with EE2. Despite
intensive research on the harmful effects of synthetic estrogens, very little is
known on the impact of phytoestrogens.
In order to examine the potential effects of soy-derived phytoestrogens on fish
aggressive and courtship behaviors, we performed an acute, 5 hour, exposure
of adult male zebrafish to ethanol (vehicle control), 50 pg/mL EE2 (positive
control), and two concentrations of commercially purchased soy-extract;
0.8pg/mL (low concentration) and 4pg/mL (high concentration). We then
compared the aggressive2 and courtship behaviors of the vehicle control and all
three estrogen treatments of exposed fish.
Our study suggests that phytoestrogens present in soy extracts that are sold
commercially as dietary supplements, may cause a reduction in male zebrafish
aggression and courtship behavior. These results indicate possible negative
impacts on the competitive and reproductive success of male zebrafish which
may be similar in wild fish populations that are exposed to estrogen discharges
from sewage systems.
Experimental Approach & Timeline
Exposure of adult male zebrafish to phytoestrogen-containing soy
extract will negatively affect aggressive and courtship behaviors
Hypothesis
Exposure Groups:
2 µl/mL ethanol (vehicle control)
50 pg/ml 17α-ethinylestradiol (EE2; synthetic estrogen)
0.8 pg/mL soy extract (low natural)
4 pg/mL soy extract (high natural)
Experimental System:
EthoVision XT 10 (Noldus Information Technology,
The Netherlands)
Results
Conclusions References
Funding
This study was supported by the Woods Hole
Oceanographic Institution and Woods Hole
Partnership Education Program
Fig. 1- Average cumulative duration spent in aggressive zone
Fig. 2- Average cumulative duration spent in close zone
Fig. 3- Frequency of female association
Fig. 4- Average cumulative duration spent in female association
Here we show that acute exposure of adult male zebrafish to soy extract potentially results in altered
aggressive and courtship behaviors.
Our results indicate possible negative impacts on the reproductive and competitive success of male zebrafish.
These effects may be similar in other aquatic organisms in the natural environment if exposed to estrogen
discharges.
1. Colman et al, 2009. Aquatic Toxicology. 91: 346-354.
2. Gerlai et al, 2000. Pharmacology, Biochemistry and Behavior. 67: 773-782.
0 - 1 2 - 3 4 - 5 6 - 7 8 - 9 1 0 - 1 1
1 0
2 0
3 0
4 0
5 0
T im e (m in )
Avg.CumulativeDuration(s)
V e h ic le C o n tro l, N = 7
E E 2 , N = 7
L o w n a tu ra l, N = 6
H ig h n a tu ra l, N = 7
*
*
V e h ic le C o n tro l
E E 2
L o w N a tu ra l
H ig h N a tu ra l
0
1 0 0
2 0 0
3 0 0
Fem.Assoc.Frequency
*
*
There were no significant differences
between treatments in any of the time
points (Two-way ANOVA, P < 0.05).
A significant difference was found
between the low and high
concentrations of natural estrogen
during the second minute of the
trial (Two-way ANOVA, Tukey’s
multiple comparisons test, P <
0.05).
The frequency of female association was
reduced in males following estrogen
exposure. The difference between
vehicle control and high concentration
of soy extract is statistically significant
(One-way ANOVA, Tukey’s multiple
comparisons test, P < 0.05).
A significant reduction in the duration of female
association was observed in all exposed males in
the first ten minutes of the assay (indicated in
green box) (Two-way ANOVA, Tukey’s multiple
comparisons test, P < 0.05).