Behavioral
Ecology
The official journal of the
ISBE
International Society for Behavioral Ecology
Behavioral Ecology (2014), 25(4), 685–699. doi:10.1093/beheco/aru019
Invited Review
Male-to-female testosterone ratios, dimorphism,
and life history—what does it really tell us?
Wolfgang Goymanna and John C. Wingfieldb
aAbteilung für Verhaltensneurobiologie, Max-Planck-Institut für Ornithologie, Eberhard-Gwinner-Straße
6a, D-82319 Seewiesen, Germany and bDepartment of Neurobiology, Physiology and Behavior,
University of California, 294 Briggs Hall, Davis, Davis, CA 95616, USA
Received 21 August 2013; revised 15 January 2014; accepted 19 January 2014; Advance Access publication 21 February 2014.
Testosterone is a key hormone for the development of secondary sexual characters and dimorphisms in behavior and morphology of
male vertebrates. Because females often express detectable levels of testosterone, testosterone has been suggested to also play a
role in the modulation of secondary sexual traits in females. Previous comparative analyses in birds and fish demonstrated a relationship between male-to-female testosterone ratios and the degree of sexual dimorphism. Furthermore, female maximum testosterone
was related to mating system and coloniality. Here, we reevaluate these previous ideas using phylogenetic analyses and effect size
measures for the relationship between birds’ male-to-female maximum testosterone levels. Further, we investigate the seasonal androgen response of female birds (the difference from baseline to maximum testosterone), which in males is strongly related to mating
system. We could not confirm a relationship between male-to-female testosterone, maximum female testosterone, or the seasonal
androgen response of females with any life-history parameter. We conclude that the expectation that testosterone regulates traits in
females in a similar manner as in males should be reconsidered. This expectation may be partially due to hormone manipulation studies using pharmacological doses of testosterone that had similar effects in females than in males but may be of limited importance for
the physiological range of testosterone concentrations occurring within ecological and evolutionary contexts. Thus, the assumption
that circulating testosterone should covary with ecologically relevant secondary sexual traits in females may be misleading: selection
pressures on females differ from those on males and females may regulate behavior differently.
Key words: aggression, androgen, aves, body size, female, latitude, mating system, plumage.
Introduction
The sex steroid testosterone is a key hormone player with regard
to the development of some secondary sexual characters or
ornaments in males (including morphological, physiological,
and behavioral traits; e.g., Lincoln et al. 1972; Harding 1981;
Balthazart 1983; Wingfield et al. 1990, 2006; Wingfield and
Silverin 2002; Oliveira 2004; Hirschenhauser and Oliveira 2006;
Hau 2007; Fusani 2008b). Although secondary sexual characters
are typically more developed in males, females of many species
also develop secondary sexual characteristics including weaponry,
ornamentation, and aggressive behavior (West-Eberhard 1983;
Andersson 1994; Kraaijeveld et al. 2007). Because female vertebrates often express detectable levels of testosterone and exhibit
seasonal cycles of testosterone, several authors have suggested that
it may also play a role in the modulation of secondary sexual traits
including aggressive behavior in female vertebrates (reviewed, e.g.,
by Staub and de Beer 1997; Ketterson et al. 2005; Rosval 2013).
Address correspondence to W. Goymann. E-mail: [email protected].
© The Author 2014. Published by Oxford University Press on behalf of
the International Society for Behavioral Ecology. All rights reserved. For
permissions, please e-mail: [email protected]
The evolution of a trait can be influenced by correlations between
the effects of genes on male and female characters, and selection acting on one sex may produce a correlated response in the other sex
(Lande 1980; Lande and Arnold 1983). This idea goes back to Darwin
(1871) and in this context, it has been suggested that testosterone concentrations in females could be a genetically correlated response to
selection on testosterone in males (Ketterson et al. 2005; Møller et al.
2005; Mank 2007) and/or vice versa (Ketterson et al. 2005).
Several comparative studies in birds have already attempted to
relate levels of testosterone in females to their life history, ecology,
and to those of conspecific males. With the exception of one study
in actinopterygiian fish (Mank 2007), we are not aware of similar
studies in other vertebrate groups (but see Staub and de Beer 1997
for a review on the role of testosterone in female vertebrates and
a recent review on “atypical” mammals by French et al. 2013).
Three avian studies correlated the ratio of mean maximum levels
of testosterone of male and female birds either with a dimorphism
index (combining differences in body size, plumage, and territorial
aggression between males and females; Wingfield 1994; Wingfield
et al. 2000) or with dimorphisms in plumage, territorial aggression,
686
and breeding density separately (Ketterson et al. 2005). These studies assumed that sexual dimorphism could be an indicator of the
degree of within-sex competition, that is, the larger sex, the sex
with the more brightly colored plumage, or the sex that shows a
higher degree of territorial aggression is experiencing a higher
degree of within-sex competition than the smaller, the less brightly
colored or the less aggressive sex, and that the degree of this competition would be reflected in testosterone concentrations. Indeed,
the ratio of mean maximum testosterone concentrations of male
and female birds was significantly related to the combined dimorphism index (Wingfield 1994; Wingfield et al. 2000) or to plumage
dimorphism (Ketterson et al. 2005): when dimorphism was low, the
ratio of male-to-female testosterone concentrations was lower than
when the dimorphism was high. Also, the greater the dimorphism,
the more variable was the ratio of male-to-female testosterone.
In addition, Ketterson et al. (2005) found that female maximum testosterone concentrations were related to mating system, with females
of socially monogamous mating systems expressing higher levels of testosterone than females of polygynous or polyandrous mating systems.
A fourth avian study investigated the slope of male and female
mean maxima of testosterone in a selected subset of birds and
found that females of colonial species had relatively higher levels of
testosterone compared with females of noncolonial species (Møller
et al. 2005). Both Møller et al. (2005) and Ketterson et al. (2005)
concluded that current comparative evidence is consistent with
both correlated evolution of testosterone concentrations in males
and females and with selection acting on each sex separately.
These previous comparative studies (and testosterone manipulation studies) have been crucial in shaping current thinking about
the role of testosterone in female birds. But for several reasons, it
is timely to reevaluate these previous ideas and discuss them in the
light of new analyses.
First, there are several methodological reasons to reanalyze
the data. Different species cannot be considered independent
data points and hence, any comparison of species needs to take
phylogeny into account (Felsenstein 1985). Only the study by
Møller et al. (2005) controlled for phylogeny, but this study was
limited in scope as it focused on the covariation of male and
female testosterone and its relationship to coloniality. Also, stateof-the art meta-analyses (e.g., Arnqvist and Wooster 1995) typically use effect sizes such as Cohen’s d (Cumming and Finch 2001;
Nakagawa and Cuthill 2007), which take the sampling variance
of the trait in question into account. As mentioned, the studies
by Wingfield (1994), Wingfield et al. (2000), and Ketterson et al.
(2005) used the ratios of male-to-female maximum testosterone
concentrations, which have the disadvantage that they do not take
sampling variance into account and thus may give a biased estimate of differences between males and females. Further, because
captivity may have a large effect on plasma testosterone levels
(e.g., Calisi and Bentley 2009), a comparison that includes only
data from free-living wild birds may be more consistent than
including data from captive birds (all previous studies). Finally,
the dimorphism indices of previous studies unfortunately did
not distinguish whether a trait was more prominently expressed
in females or males. However, for the evaluation of a dimorphic
trait, it is conceptually important to distinguish whether either the
male or the female expresses a more brightly colored plumage
or a larger body size (i.e., it is important to have more brightly
colored or larger females on the one end of the scale and more
brightly colored or larger males on the other end of the scale; see
Methods for details).
Behavioral Ecology
Second, there are important conceptual reasons concerning the
mechanisms of hormonal action that should make us reconsider
the relationship between male and female testosterone within an
ecological and evolutionary framework. From a proximate perspective, testosterone manipulation studies in females have nourished
the assumption that testosterone has similar effects and functions in
females as in males. But is this really the case? Classical studies in
endocrinology are concerned with basic mechanisms of hormone
action. To study effects of testosterone in males, one would, for
example, remove the testes and investigate the changes in behavior,
physiology, and morphology (e.g., Berthold 1849). Then, one would
attempt to restore the trait in question by administering testosterone (see reviews in Balthazart and Adkins-Regan 2002; AdkinsRegan 2005; Ball and Balthazart 2008). In such studies, females
are often used as a “blank slate” to study the organizational and
activational effects that testosterone has in males (see, e.g., the classic study on organizational and activational effects of sex steroids
on male and female copulation behavior in guinea pigs by Phoenix
et al. 1959, or studies on organizational and activational effects
of steroids on song or courtship behavior in passerine songbirds,
e.g., Leonard 1939; Gurney and Konishi 1980; Nottebohm 1980;
Hausberger et al. 1995; Fusani et al. 2001. 2003. Day et al. 2007).
If females develop male-like traits after administration of testosterone, one would conclude that the trait is testosterone dependent.
However, the hormone dosages used in these studies are typically
in a pharmacological range: a “normal” female would never experience such concentrations of testosterone under natural and ecologically relevant circumstances. We believe that this rather drastic
hormone manipulation approach is well justified within a purely
mechanistic perspective (demonstrating that sometimes even fully
sexually differentiated females have the capacity to develop malelike traits). But when adopted in a one-to-one manner in physiological or behavioral ecology to study hormone dependency of
traits under more natural conditions, this approach may become
problematic (see also Rosval 2013). In fact, females may use different mechanisms to generate male-like traits. For example, studies
of sex-role-reversed bird species in which females are more competitive than males have failed to demonstrate an important role
of high circulating levels of testosterone in the regulation of malelike traits in females (Rissman and Wingfield 1984; Fivizzani and
Oring 1986; Fivizzani et al. 1986. 1990. Oring et al. 1988; GrattoTrevor et al. 1990; Goymann and Wingfield 2004; Goymann et al.
2008). Furthermore, a recent study on the song of the forest weaver
(Ploceus bicolor) has convincingly demonstrated that males and
females use very different mechanisms to express the same kind of
behavior, that is, brain areas that are involved in the control of song
are much larger in males, but females show a higher expression of
genes relevant for the control of song than males (Gahr et al. 2008).
Thus, the rather common assumption that testosterone may have
similar functions in female birds as in males is based on relatively
weak evidence.
Third, there are important conceptual reasons to reconsider
also the ultimate relationship between male and female testosterone within an ecological and evolutionary framework. Previous
comparisons of male and female testosterone concentrations and
their association with ecologically relevant traits were based on
the implicit assumption that the evolution of ornamental traits in
females is related directly to sexual selection (representing the predominant explanation for ornamental traits in males; Darwin 1871;
Andersson 1994) or is indirect and evolved as a correlated response
to selection acting on males (Darwin 1871). But females may
Goymann and Wingfield • Male-to-female testosterone ratios, dimorphism, and life history
compete for resources and develop secondary sexual characters also
outside a mating context and thus these traits may be shaped by
evolutionary forces outside the traditional concept of sexual selection (West-Eberhard 1983; Tobias et al. 2012). Thus, the implicit
assumption that secondary sexual characters of females exclusively
evolved in a sexual selection context and may thus be related to
testosterone could be misleading as well.
Finally, to our knowledge, no study so far has tried to relate the
magnitude of the change in testosterone from the seasonal breeding baseline to the breeding maximum concentrations in females
to mating system. In male birds, this so-called seasonal androgen
response (sensu Goymann et al. 2007) shows a very robust relationship with mating system. This relationship was initially described
by Wingfield et al. (1990) and later confirmed for birds (but not
other vertebrate groups) by a comparative study (Hirschenhauser
and Oliveira 2006).
Because birds are the taxonomic group for which most information regarding plasma testosterone concentrations of free-living
males and females are available, we decided to base our analysis
on this taxon, just as most of the previous studies did. We are convinced, however, that the results will be of general relevance also
for other vertebrate groups. Using a coherent, phylogenetically controlled set of plasma testosterone data from 51 free-living wild bird
species, we here ask whether the standardized difference in mean
maximum testosterone concentrations between female and male
birds (the standardized effect size) is related to dimorphisms in body
size and plumage coloration, to the mating system, to the latitudinal distribution of birds (because latitude has an effect in males,
see Garamszegi et al. 2005), and to coloniality. Furthermore, we
investigated whether maximum levels of testosterone in females
relate to any of these parameters and whether the seasonal androgen response of females is related to mating system. In summary,
we demonstrate that maximum male-to-female testosterone is not
related to measures of dimorphism or any of the other parameters and argue that the expectation that physiological levels of
testosterone may have similar effects and functions in female birds
as in males may have been misleading and should probably be
reconsidered.
Methods
We reviewed the existing literature (until December 2012) on
plasma testosterone levels reported for males and females of the
same species taking into account the information provided by previous comparative studies (Wingfield 1994; Wingfield et al. 2000;
Ketterson et al. 2005; Møller et al. 2005) and a search in ISI Web
of Knowledge using the search terms “testoster*” and “femal*”
within the taxonomic group of “aves.” Unlike these previous studies, we only considered data from 51 species of free-living, wild
birds, because captivity may affect plasma testosterone levels (Calisi
and Bentley 2009). To calculate mean peak levels of testosterone of
males and females, we included only studies providing several measures (i.e., a seasonal profile) of plasma testosterone for both sexes
and a sample size of at least 3 individuals for the time of peak testosterone concentrations. The mean peak level of testosterone was
defined as the highest average concentration of testosterone (and
the respective standard errors) reported in any of the early breeding substages (territory establishment, prebreeding, nest-building,
and egg-laying) in actively breeding birds of the respective species.
A similar approach was used for baseline testosterone data from
females measured during the parental phase, that is, assuming that
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testosterone levels were basal when females were feeding nestlings
(following Wingfield et al. 1990). For cooperative breeders, only
data from breeding pairs and not helpers were considered.
A general problem of comparative studies of absolute hormone
levels is that hormone concentrations were determined in different
laboratories. This is only a minor issue for the following reasons.
First, for the male-to-female comparison, we used standardized
effect sizes (Cohen’s d) of the difference in male and female maximum levels of testosterone. A similar approach was used for the
seasonal androgen response of females. Hence, in these 2 comparisons, absolute differences in hormone levels are no longer relevant
(at least when males and females have been measured in the same
lab, which is the case in most studies). Second, a previous study
did not find any pattern in testosterone that would be explained by
lab (Goymann et al. 2004). Nevertheless, for the current study, we
decided to include only data gathered by radioimmunoassay (RIA)
for the following reason. Several studies have measured plasma
testosterone levels of female birds using an Assay Designs (Ann
Arbor, MI, #901-065) enzyme immunoassay (EIA) for testosterone.
Testosterone levels of female birds measured with this EIA seem
consistently higher than those determined by RIA. For example,
the testosterone levels of female white-throated sparrows (Zonotrichia
albicollis) measured with this EIA (Swett and Breuner 2008) were
consistently higher than those of the same species measured with
RIA (Spinney et al. 2006). Males of the tan-striped morph of
white-throated sparrows had relatively similar mean maximum levels of testosterone with 3 ng/mL (EIA; Swett and Breuner 2008)
and 2.5 ng/mL (RIA; Spinney et al. 2006). In contrast, females of
the tan-striped morph had about 1.7 ng/mL testosterone with EIA
(Swett and Breuner 2008), but only 0.4 ng/mL with RIA (Spinney
et al. 2006). Thus, because the EIA appears to overestimate testosterone concentrations specifically in females, the relationship
between male-to-female testosterone may become distorted. Hence,
we decided to include only testosterone data measured after sample
extraction with RIA. Also, we did not include testosterone metabolite data from fecal or dropping assays because such assays may
not allow a sound comparison of females and males (see Goymann
2005, 2012, for a discussion of this methodological problem).
Information about mating system, body size, plumage dimorphism, absolute latitude, and coloniality was extracted from the
original papers and additional information using bird handbooks
(Cramp 1977; del Hoyo et al. 1992; Poole 2005). Similar to previous studies (Wingfield 1994; Wingfield et al. 2000; Ketterson
et al. 2005), we related differences in testosterone concentration of
females and males to differences in body size and plumage dimorphism, as well as to mating system (social monogamy, polygyny,
classical polyandry, and cooperative breeding) and distinguished
whether the species is a colonial breeder or not. With regard to size,
the previous studies used a scale from 1 to 3 with 1 = sexes of similar size (80–100% overlap), 2 = 5–80% overlap between sexes, and
3 = no overlap between the sexes. Obviously the studies did not
distinguish whether males are larger or females are larger. Instead
of a scale, we used body mass data extracted from Lislevand et al.
(2007) and calculated the proportion of female body mass relative to male body mass and used this proportion as a covariate in
the analysis. With regard to plumage, we scored plumage from −1
(female more brightly colored than male), 0 (female and male with
similar plumage) to +1 (male more brightly colored than female). In
addition, we also included absolute latitude because previous studies have found that mean maximum testosterone concentrations of
male birds can depend on absolute latitude (Goymann et al. 2004;
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Garamszegi et al. 2008). The compiled data set with the respective
references can be found in Supplementary Tables S1 and S2.
The phylogeny used was derived from Hackett et al. (2008) using
MESQUITE (Maddison and Maddison 2007) with arbitrarily ultrametricized branch lengths (Supplementary Figure S1). Effect sizes
and their 95% confidence intervals (CIs) between male and female
testosterone concentrations and between seasonal baseline and seasonal maxima of testosterone concentrations in females were calculated with the program ESCIdelta (Cumming and Finch 2001).
The sampling variance of the effect size was defined as V = [(+95%
CI − 95% CI)/4]2. Data were analyzed using a Markov Chain
Monte Carlo algorithm (MCMCglmm R package) (Hadfield 2010)
in R 2.15 (R Development Core Team) following the instructions
for phylogenetic analyses of continuous and categorical characters
(Hadfield and Nakagawa 2010). We ran several “animal models”
with the effect size difference between maximum testosterone concentrations in males and maximum testosterone concentrations in
females as a dependent variable and either body size dimorphism,
plumage dimorphism, mating system, absolute latitude, or coloniality as separate independent variables or a combined model with
body size dimorphism, plumage dimorphism, mating system, and
absolute latitude as independent variables. When combining all
measures in one model, plumage dimorphism was entered as a continuous variable. Similar “animal models” were used to test for an
effect of these parameters on female maximum testosterone concentrations and the effect size difference between maximum and breeding baseline concentrations of testosterone (during the parental
phase) in females in relation to mating system. In addition, we tested
for a correlation of mean maximum testosterone concentrations of
males and females. In all models, we controlled for phylogeny (using
the random~animal function implemented in MCMCglmm) and
measurement error variance (using the mev function implemented
in MCMCglmm to control for sampling variance in the effect sizes
or mean maximum testosterone concentrations). All models were
run in 1 000 000 iterations with a burn-in period of 60 000 iterations
and a thinning interval of 100. Autocorrelation of the posterior distribution was checked using the autocorr function implemented in
MCMCglmm and model fit was compared using the deviance information criterion (DIC). Meaningful estimates for categorical predictors were extracted by rerunning all models without intercept and
reporting the respective posterior means and their confidence limits
(following Schielzeth 2011).
To test for publication bias, that is, the likelihood that studies
reporting statistically significant effects are more likely to be published than studies with nonsignificant effects, we applied Egger’s
regression (Egger et al. 1997) to the meta-analytic residuals
(Nakagawa and Santos 2012) plotted against their original measurement error variance using the model of the male–female comparison with the lowest DIC value (model no. 5, latitude only) and
the model in which we tested the effect size difference of baseline
and maximum testosterone in females with respect to mating system. If the intercept of the Egger’s regression does not significantly
differ from zero, then it can be concluded that there is little evidence for publication bias (Egger et al. 1997).
Results
The mean maximum levels of testosterone in male and female bird
species were slightly, but significantly, correlated when controlling
for phylogeny and sampling variance (MCMCglmm, P = 0.0378;
Figure 1).
Behavioral Ecology
Figure 1
Mean maximum levels of testosterone in male and female birds are
significantly correlated (P = 0.0378). Data points represent means ± 95%
CI. The regression line is y = 0.110x + 0.292.
The magnitude of the difference between maximum testosterone
concentrations in male and female birds (effect size Cohen’s d) was
not related to coloniality (Figure 2a), mating system (Figure 2b),
plumage dimorphism (Figure 2c), body size dimorphism (Figure 2d),
or latitude (Figure 2e and Table 1) when controlling for phylogeny
and sampling variance. Also, an overall model including mating
system, plumage dimorphism, body size dimorphism, and latitude
did not explain a significant proportion of the variance in the data
(Table 1). In addition, because the previous studies by Wingfield
(1994), Wingfield et al. (2000), and Ketterson et al. (2005) used
ratios instead of effect sizes, we investigated whether using ratios
in a phylogenetic study would give similar results to those reported
by these previous authors. However, a phylogenetic analysis of the
male-to-female ratio in maximum testosterone from free-living
birds also did not confirm these previous results (Supplementary
Table S3), with the exception that there was a trend for absolute
latitude, which was probably driven by testosterone concentrations of males and had nothing to do with females (see below and
Garamszegi et al. 2008). The intercept from the Egger’s regression
to test for publication bias was statistically significant (b0 = 3.298,
95% CI = 1.991–4.748, Supplementary Figure S2a). This result
can be interpreted in 2 ways: Either there is heterogeneity among
the studies of our data set, which is caused by unmeasured variables, or the data set is skewed due to publication bias (Egger et al.
1997). Heterogeneity cannot be tested, but publication bias can be
assessed by using the trim and fill test, a method to estimate the
number of studies that are missing from a meta-analysis and the
effect that these studies might have on the outcome of the metaanalysis (Duval and Tweedie 2000). We used the R package meta
(Schwarzer 2010) and applied its implemented trim and fill method
to the meta-analytic residuals and the corresponding measurement
variance. The trim and fill test added 13 additional data points to
the original 50 data points and gave an estimate of −0.459 (94%
CI = −0.759 to −0.159; Supplementary Figure S2b). Close inspection of Supplementary Figure S2b indicates that the publication
bias is due to the fact that there are very few studies with a negative effect size, that is, there are only very few bird species in which
Goymann and Wingfield • Male-to-female testosterone ratios, dimorphism, and life history
689
Table 1
Relationship of the standardized difference between male and female maximum testosterone concentrations (Cohen’s d) with
coloniality, mating system, plumage and size dimorphism, and latitude
Factor/variable
Posterior mean
−95% CI
+95% CI
P value (MCMC)
Model 1: effect size ~ coloniality, random = ~animal, mev = effect size variance
Colonial (intercept)
1.677
0.276
3.010
Noncolonial
1.517
0.334
2.683
0.769
Model 2: effect size ~ mating system, random = ~animal, mev = effect size variance
Monogamy (intercept)
1.621
0.451
2.829
Cooperative
0.602
−1.344
2.481
0.270
Polyandry
1.796
0.119
3.789
0.857
Polygyny
1.863
0.328
3.437
0.693
Model 3: effect size ~ plumage dimorphism, random = ~animal, mev = effect size variance
Similar (intercept)
1.457
0.368
2.567
Female brighter
1.662
−0.719
4.153
0.858
Male brighter
1.923
0.566
3.450
0.401
Model 4: effect size ~ body size dimorphism, random = ~animal, mev = effect size variance
Intercept
1.240
−1.400
3.757
Body size dimorphism
0.304
−2.095
2.671
0.799
Model 5: effect size ~ latitude, random = ~animal, mev = sampling variance
Intercept
0.911
−0.640
2.374
Latitude
0.016
−0.011
0.042
0.246
Model 6: effect size ~ mating system + body size + plumage dimorphism + latitude, random = ~animal, mev = effect size variance
Monogamy (intercept)
1.005
−2.532
4.742
Cooperative
0.213
−3.592
3.908
0.404
Polyandry
1.422
−1.406
4.472
0.709
Polygyny
1.003
−3.471
5.432
0.981
Body size dimorphism
0.043
−3.383
3.601
0.968
Plumage dimorphism
0.308
−2.005
2.535
0.622
Latitude
0.013
−0.018
0.043
0.413
DIC
168.8
171.2
171.8
169.9
169.1
178.5
Meaningful posterior means for categorical variables (and CIs) were extracted by rerunning all models without intercept; P values refer to differences of
categorical variables with respect to the reference category (intercept). Posterior means of continuous variables refer to the slope of the relationship.
females have significantly higher levels of testosterone than males.
This “bias” is obviously driven by the fact that males naturally have
higher levels of testosterone than females in most species. A true
publication bias would indicate that studies reporting significantly
higher levels of testosterone in females than in males would be less
likely to be published than studies that report higher levels in males
than in females (Supplementary Figure S2).
The magnitude of the difference between maximum testosterone
concentrations in male and female birds could be related to either
particularly high or low levels in males or particularly high or low levels in females. For example, the effect size could be small because both
females and males have very high levels of testosterone, or a similar
effect size could emerge because both sexes have very low levels (see
also Ketterson et al. 2005). We thus looked at whether mean maximum testosterone concentrations of females per se are related to any
of the variables in question. Mean maximum levels of testosterone
in female birds did not show any relationship with mating system,
plumage dimorphism, body size dimorphism, coloniality, or absolute
latitude (Figure 3 and Table 2), when controlling for phylogeny and
sampling variance in female maximum testosterone concentrations.
Finally, in contrast to the strong relationship in males (Wingfield
et al. 1990; Hirschenhauser and Oliveira 2006; Goymann et al. 2007),
the effect size difference between breeding season baseline levels of
testosterone and seasonal maxima of testosterone in female birds was
not related to mating system (Figure 4 and Table 3). The intercept of
the Egger’s regression to test for publication bias was also statistically
significant (b0 = 3.779, 95% CI = 2.437–5.315, Supplementary Figure
S3a). Again, also this result can be interpreted in 2 ways: Either there
is heterogeneity among the studies of our data set, which is caused by
unmeasured variables, or the data set is skewed due to publication bias
(Egger et al. 1997). Again, we assessed publication bias by using the
trim and fill test (Duval and Tweedie 2000) using the R package meta
(Schwarzer 2010) to the meta-analytic residuals and the corresponding measurement variance. In this case, the trim and fill test added
12 additional data points to the original 44 data points and gave an
estimate of −0.411 (94% CI = −0.686 to −0.136; Supplementary
Figure S3b). Close inspection of Supplementary Figure S3b indicates
that the “publication bias” is due to the fact that baseline testosterone
concentrations are by definition always lower (or equal) during the
parental phase than during the period of peak concentrations of this
hormone (Supplementary Figure S3).
Discussion
In this study, we used effect sizes and phylogeny to study the relationship between male and female maximum levels of plasma
testosterone measured in free-living birds. The results confirmed earlier reports that mean maximum testosterone levels
of male and female birds are correlated (Ketterson et al. 2005;
Møller et al. 2005), albeit with a very shallow slope in the relationship (Figure 1). However, we could not confirm any of the
other relationships described in earlier reports: there was no indication that the standardized difference (i.e., effect size) of maleto-female maximum testosterone concentrations was related to
mating system, plumage and body size dimorphisms, coloniality,
or—previously untested—absolute latitude. Also, mean maximum
concentrations of testosterone in females per se did not significantly covary with any of these variables. Finally, unlike in male
birds (Wingfield et al. 1990; Hirschenhauser and Oliveira 2006;
Goymann et al. 2007; Goymann 2009), mating system was not
related to the seasonal increase of testosterone in females. Thus,
the relationship of male-to-female testosterone did not seem to
690
Behavioral Ecology
Figure 2
Effect size differences of male-to-female testosterone were not related to (a) coloniality, (b) mating system, (c) plumage dimorphism, (d) body size dimorphism,
and (e) latitude (see Table 1 for statistics). Data are presented as posterior means (±95% CI) corrected for phylogeny and jittered individual data points.
be a good predictor of sex differences in intra- and intersexual
competition and female testosterone concentrations did not seem
to relate to differences in female life history. Because our study
only considered testosterone in adult males and females, it did not
address organizing effects that testosterone may have during ontogenetic development.
Female testosterone—a correlated response to
selection on testosterone in males?
Testosterone concentrations of male and female birds were weakly
correlated when controlling for phylogeny. But does this imply that
female testosterone is a correlated response to selection (and expression) of testosterone in males, or—taking a female perspective (Zuk
Goymann and Wingfield • Male-to-female testosterone ratios, dimorphism, and life history
2002; Gowaty 2003)—that male testosterone concentrations could
be constrained by selection on low concentrations of this hormone
in females (see also Ketterson et al. 2005)? Considering the shallow
slope of the relationship and the large variance between and even
within species (Figure 1), both ideas seem unlikely, that is, there seem
to be some degrees of freedom to avoid a strong constraint. Given
that testes—the main tissue producing testosterone in males—do
not exist in females and given further that the production, metabolism, and action of testosterone is influenced not just by 1 gene, but
by a magnitude of processes (including the expression of production and metabolizing enzymes, receptors, and cofactors; see e.g.,
Hau 2007), it would be surprising if testosterone concentrations of
females would be constrained by selection on males or vice versa.
Furthermore, testosterone and androstenedione are precursor molecules of estradiol, a central hormone of female reproductive physiology (Bentley 1998). Thus, all females produce testosterone and/or
androstenedione as a first step and then convert them to estradiol
691
(in fact, also in males, many behavioral effects of testosterone are
mediated through local conversion to estrogens). It is hard to conceive that females would be constrained in converting testosterone
into one of the major sex steroids involved in female reproduction.
One of the clearest arguments against the importance of correlated selection with regard to sex steroids is sexual differentiation,
a coordinated process that, among other processes, leads to differences in the ratio of estradiol and testosterone between females
and males triggering differences in primary and secondary sexual
traits without altering other organs or traits (reviewed by AdkinsRegan 2008). Although selection on secondary sexual characteristics indeed can lead to correlated responses in the other sex (e.g.,
Harrison 1953; Wilkinson 1993; Price and Birch 1996; Chenoweth
and Blows 2003), usually such correlated responses are weak (but
see Dale et al. 2007 for a potentially strong one). Clutton-Brock
(2009) has pointed out that “comparative studies indicate that sexlinked modification of the expression of ornaments (or other sexual
traits) is more common than sex-linked inheritance of ornament
genes” (see also Amundsen 2000; Wiens 2001; Pointer et al. 2013).
This is particularly true for hormonal pathways, which may show
large differences even among closely related species (Hau 2007;
Adkins-Regan 2008). In this context, it is interesting that in fish, the
correlation between female and male testosterone has a steep slope
(Mank 2007), suggesting a strong connection between female and
male testosterone. Given that the magnitude of testosterone expression is similar in female and male fish (Mank 2007), and given that
male fish “invented” 11-keto-testosterone that fulfills many of the
functions that testosterone does in other vertebrate groups, it is difficult to interpret these findings on testosterone or relate them to
those of other vertebrates.
Mating system and testosterone
Figure 3
Female maximum testosterone concentrations were not related to coloniality
(see Table 2 for statistics). Data are presented as posterior means (±95% CI)
corrected for phylogeny and jittered individual data points.
Arguably, the sample sizes for mating systems other than social
monogamy were small (but not smaller than Ketterson et al. 2005,
who came to different conclusions) and may prevent a definite
answer, but the data did not even show the slightest trend. Even
though the sample for nonmonogamous birds was limited, we
Table 2
Relationship of female maximum testosterone concentrations with coloniality, mating system, plumage and size dimorphism, and
latitude
Factor/variable
Posterior mean
−95% CI
+95% CI
Model 1: female testo ~ coloniality, random = ~animal, mev = (SEM female testo)2
Colonial (intercept)
0.639
0.161
1.114
Noncoloniality
0.368
−0.048
0.782
Model 2: female testo ~ mating system, random = ~animal, mev = (SEM female testo)2
Monogamy (intercept)
0.514
0.060
0.974
Cooperative
0.299
−0.378
0.973
Polyandry
0.305
−0.385
1.001
Polygyny
0.376
−0.186
0.950
2
Model 3: female testo ~ plumage dimorphism, random = ~animal, mev = (SEM female testo)
Similar (intercept)
0.481
0.052
0.901
Female brighter
0.243
−0.617
1.041
Male brighter
0.352
−0.159
0.885
Model 4: female testo ~ body size dimorphism, random = ~animal, mev = (SEM female testo)2
Intercept
0.471
0.057
0.858
Body size dimorphism
0.385
−0.399
1.219
Model 5: female testo ~ latitude, random = ~animal, mev = (SEM female testo)2
Intercept
0.162
−0.372
0.681
Latitude
0.007
−0.002
0.016
P value (MCMC)
0.144
0.480
0.495
0.516
0.528
0.463
0.359
0.103
DIC
51.9
60.6
57.2
56.7
55.1
Meaningful posterior means for categorical variables (and their CIs) were extracted by rerunning all models without intercept; P values refer to differences of
categorical variables with respect to the reference category (intercept variable). Posterior means of continuous variables refer to the slope of the relationship.
Behavioral Ecology
692
wonder whether an absence of a relationship between male-tofemale testosterone and mating system really is surprising. Previous
studies have shown that peak testosterone concentrations of male
birds do not vary with mating system (Wingfield et al. 1990;
Beletsky et al. 1995; Hirschenhauser et al. 2003; Goymann et al.
2004; Garamszegi et al. 2005; see also summary in Goymann
2010). Hence, any effect related to mating system would need to
be driven by differences in maximum testosterone concentrations
of females. But if peak testosterone does not vary as a function
of mating system in males—the sex in which the hormone arguably plays a role in competition—why should it do so in females?
Indeed, also maximum testosterone in females was not related to
mating system, when controlling for phylogeny. A number of studies in classically polyandrous birds has demonstrated that maximum testosterone concentrations in females that fiercely compete
among each other and often have a more brightly colored plumage
than males are similar or even lower than those of species in which
females do not compete for access to mates (Rissman and Wingfield
1984; Fivizzani et al. 1986; Fivizzani and Oring 1986; Oring et al.
1988; Gratto-Trevor et al. 1990; Goymann and Wingfield 2004).
Thus, in a context in which female ornamentation clearly evolved
within the sexual selection paradigm, testosterone concentrations
of females are not related to competition. However, why should
we expect that maximum testosterone should reflect the degree of
competition in females, if maximum testosterone is not related to
Figure 4
The effect size of the seasonal androgen response (the difference between
seasonal baseline levels of testosterone and the seasonal peak) of female
birds was not related to mating system (see Table 3 for statistics). Data
are presented as posterior means (±95% CI) corrected for phylogeny and
jittered individual data points.
degree of competition in males (see above)? A closer look at the significant result of mating system in Ketterson et al. (2005) suggests
that the significantly higher levels of testosterone in socially monogamous females were mainly driven by high maximum levels of testosterone (>1 ng/mL) in female penguins, albatrosses, the dark-eyed
junco, and the Lapland longspur, thus highlighting the importance
of controlling for phylogeny (and asking the question why females
of these species express such high levels of testosterone?).
What about the seasonal dynamics of testosterone and mating
system? In males, the seasonal increase in testosterone is related to
mating system with males from socially monogamous mating systems showing a larger increase than polygynous males (Wingfield
et al. 1990; Hirschenhauser and Oliveira 2006; Goymann et al.
2007). It has been argued that socially monogamous males show
this very dynamic response because high levels of testosterone may
interfere with parental care (Wingfield et al. 1990). In female birds,
the seasonal difference in testosterone from breeding baseline to
peak levels was not related to mating system. Again, low sample
sizes for mating systems other than social monogamy may preclude
definitive answers, but also here, there was not even a trend in the
data (but a large scatter in the socially monogamous group). The
absence of a relationship of the seasonal change in testosterone
with mating system in female birds may not be surprising, though,
because peak testosterone levels in females are usually much lower
than in males and it is thus unlikely that such low levels would
interfere with parental care. Furthermore, in male birds, the testes represent a reliable source of testosterone until they regress at
the end of the breeding season. In female birds, gonadal testosterone mainly stems from developing follicles that produce sex steroids including testosterone. After finishing egg-laying, the gonadal
release of steroids drops (Johnson 1998; Johnson and Woods 2007;
Williams 2012). Even though primordial and primary follicles may
continue to produce steroids (Johnson and Woods 2007), it is likely
that after egg-laying, females do not have the capacity to modulate gonadal testosterone in a similar manner as males (see also
Jawor et al. 2007 for evidence in dark-eyed juncos). Also in other
vertebrate groups, the female gonad typically releases sex steroids
in a less predictable manner than that of males (e.g., Crews 1998;
Ferin 1998; Khan and Thomas 1998). Thus, it is an open question
whether the hypothalamic–pituitary–gonadal axis is involved in the
regulation of short-term changes in testosterone in female birds
and females of other vertebrates. It is well possible that nongonadal
sources such as the adrenal gland or the brain could be involved.
Plumage and body size dimorphism and
testosterone
The phylogenetic effect size analysis could not confirm earlier data
suggesting that male-to-female maximum testosterone was significantly related to a dimorphism index (aggression, body size, and
plumage; Wingfield 1994; Wingfield et al. 2000) or dimorphism in
Table 3
Effect size of the seasonal androgen response of females in relation to mating system
Factor/variable
Posterior mean
−95% CI
Model: eff_fem testo ~ mating system, random = ~animal, mev = Var eff_fem testo
Monogamy (intercept)
1.034
0.353
Cooperative
1.534
0.106
Polyandry
0.872
−0.453
Polygyny
1.160
0.110
+95% CI
P value (MCMC)
1.662
2.825
2.236
2.271
0.450
0.801
0.780
DIC
121.4
Goymann and Wingfield • Male-to-female testosterone ratios, dimorphism, and life history
plumage only (Ketterson et al. 2005). We are fully aware that plumage and the other sexually dimorphic traits served as a proxy for
sex differences in competition, but again we wonder whether the
absence of a relationship between male-to-female testosterone and
plumage dimorphism as a measure for competition is really surprising (even though sample size—in particular for more brightly
colored females than males—was again limited, thus possibly preventing a more definite conclusion). From an ultimate perspective,
plumage is a complicated trait and sexual selection may not be the
only reason for the development of a brightly colored plumage.
In particular, the plumage of monomorphic brightly colored and
social species may be under selection for social interactions within
the group rather than under sexual selection for courtship within
a pair (West-Eberhard 1983). In contrast to birds, dichromatism,
elongated fins, breeding tubercles, and mating calls in ray-finned
fish seem to be unambiguously the result of sexual selection and
indeed are related to high levels of 11-keto-testosterone and testosterone in this other vertebrate taxon (Mank 2007).
On a mechanistic level, the dependence of a brightly colored
plumage on sex steroids is also debated and may differ depending on the species and molting pattern (e.g., Witschi 1961; Lindsay
et al. 2009, see also Table 4 highlighting the magnitude of control mechanisms of sexual dimorphisms). Postnuptial molt in males
typically occurs when androgen levels are low (e.g., Nolan et al.
1992; Goymann et al. 2006) and can be suppressed by high levels
of these hormones (Hahn et al. 1992; Nolan et al. 1992; Kimball
and Ligon 1999; Stoehr and Hill 2001; but see Laucht et al. 2011
and McGlothlin et al. 2008 for evidence that breeding season levels of testosterone may correlate with plumage characteristics). Less
is known about the influence of androgens on prenuptial molt in
males, but there is some evidence that prenuptial molt may indeed
be regulated by plasma androgens (e.g., Witschi 1961; Kimball
and Ligon 1999; Peters et al. 2000; Day et al. 2006; Lindsay et al.
2009). Recent evidence suggests that differences in plasma androgen concentrations during prenuptial molt may be even responsible
for individual variance in plumage traits including melanin-based
and structural colors (Bókony et al. 2008; Lindsay et al. 2009). In
Struthioniformes, Galliformes, and Anseriformes a brightly malelike colored plumage develops in the absence of estrogens, whereas
a female-like dull plumage develops in the presence of estrogens
(Kimball and Ligon 1999). Similar mechanisms may operate in
693
some passerines (Perlut 2008). Thus, in males of different species,
sexually dimorphic plumage traits are regulated by different mechanisms, probably preventing a simple connection of plumage traits
to plasma testosterone and sexual selection in males. In comparison, much less is known how elaborate plumage traits are regulated
in females. Certainly, females seem to be able to exploit strategies
other than plasma testosterone concentrations: In sex-role-reversed
red-necked phalaropes (Phalaropus tricolor), females express a more
brightly colored breeding plumage than males, but plasma testosterone concentrations of females are far lower than those of males
(Gratto-Trevor et al. 1990). Female phalaropes express higher concentrations of androgen conversion enzymes in the skin than males,
suggesting that local conversion of sex steroids in the skin could
be involved in the regulation of sexually dimorphic plumage in
this species (Fivizzani et al. 1990). Hence, a relationship between
sex-role-reversed plumage traits and plasma androgens may not
necessarily be expected in phalaropes and other species in which
females have a more brightly colored plumage than males (but see
Muck and Goymann 2011 for an example where testosterone correlates with elaborate female plumage traits). Thus, the influence
of androgens depends on the molting pattern (pre- and/or postnuptial molt) and can be regulated locally, preventing a generalization of the relationship between plumage dimorphism and plasma
testosterone.
Similarly, is the absence of a relationship between male-tofemale testosterone and body size dimorphism surprising? Once
more, sexual dimorphism in body size is a complicated trait, it may
have evolved for many reasons and it may be controlled by different
mechanisms (including organizing effects of steroids during development, see also Table 4 for an overview of potential mechanisms).
Hence, larger body size is not necessarily a trait related to increased
competition for mates, which could then be reflected in plasma testosterone profiles, if the magnitude of testosterone would be indicative of competition. For example, sexual selection may favor large
or small body size in males: strong sexual competition for mating
partners may favor large-bodied males, whereas in sexual selection for male display agility, females may select even smaller males
(Andersson 1994; Székely et al. 2009; but see Dale et al. 2007). But
sexual selection is not the only factor shaping differences in body
size: sexual size dimorphism may have evolved for other reasons,
that is, resource division or sexual fecundity (reviewed by Andersson
Table 4
Potential control mechanisms of sexual dimorphism in birds
Trait
Ontogenetic/genetic
Epigenetic
Hormonal
Body size and shape
Sex-specific brain structures
Song control system, social brain network, parental and aggression circuits, etc.
Sex-specific color
Plumage, skin, eye iris, eye ring, feet, tarsi, etc.
Sex-specific reproductive organs
Oviduct, deferent duct, cloacal protuberance, etc.
Territorial behavior
Multipurpose territory, lek position, display territory, nest site, mate guarding, etc.
Sexual behavior
Pair bonding, courtship, copulatory behavior, etc.
Parental behavior
Nest-building, nest maintenance, incubation, brooding, feeding offspring, escort, parental
aggression, etc.
Parental structures, physiology
Brood patch, crop sac structure, mouth pouches, water carrying, crop-milk, stomach oil, etc.
Yes
Yes
Yes
?
Yes
Yes
Yes
Yes
Yes
Yes
?
Yes
Yes
Yes
Yes
Yes
Yes
Yes
Yes
Yes
Yes
Yes
?
Yes
694
1994; Blanckenhorn 2000; Székely et al. 2009). In particular, in
classical polyandrous bird species, it is not clear whether females
are larger than males because of stronger intrasexual competition
between females or because of the fecundity advantages that a
larger female may possess. Thus, body size difference between the
sexes is not a universal indicator of increased competition in the
larger sex and, for this simple reason, may not be indicative for the
relationship of plasma testosterone levels.
Coloniality, latitude, and testosterone
Møller et al. (2005) found that females of colonial species had
significantly higher levels of testosterone compared with females
of noncolonial species. Møller et al. (2005) used a phylogeny but
included data from species held in captivity. Also, the current data
set comprises a larger number of noncolonial species than the initial analysis of Møller et al. (2005). Similar to the mating system
effect in the Ketterson et al. (2005), all species in the group of colonial birds in which females have particularly high levels of testosterone are either penguins or albatrosses, suggesting that something
is special about this group of birds, when it comes to testosterone
concentrations in females. Thus, the effect that Møller et al. (2005)
found may not have been related so much to coloniality as such,
but to the fact that female penguins and albatrosses that nest colonially, express relatively high levels of testosterone compared with
females of other colonially nesting species.
Previous comparative analyses have demonstrated a latitudinal effect on testosterone with a trend that males of high latitude
species express higher maximum testosterone concentrations than
lower latitude males (Goymann et al. 2004; Garamszegi et al.
2008), although there are many factors that modify this pattern, in
particular, the length of the breeding season: the shorter the breeding season, the higher the peak in testosterone also in low latitude
birds (Goymann et al. 2004; Hau et al. 2008). The present data
suggest that a similar effect does not exist in females.
A role of testosterone in the expression of
female secondary sexual traits?
These results open the question whether there is evidence for a role
of plasma testosterone in the expression of female birds’ secondary sexual traits? No doubt, females express detectable levels of
androgens (such as testosterone), androgen-metabolizing enzymes,
and androgen receptors, and androgens definitely play a role in
the regulation of traits in females (see Introduction and, e.g., the
review in Staub and de Beer 1997). However, the common assumption that plasma levels of testosterone play a similar role in females
as in males, that is, that the hormone is involved in the expression
of secondary sexual traits involved in sexual or social competition,
may have been overrated due to the effects found in testosterone
manipulation studies (see Introduction). Unfortunately, we are still
not good at dosing hormone implants within a physiological range:
the concentration levels that hormone implants initially generate typically by far exceed the possible physiological levels that are
present in the circulation of birds (particularly during the first days
after implantation, which is rarely checked; see e.g., the studies of
Edler et al. 2011 and Fusani 2008a). Such supraphysiological levels
of testosterone are problematic if interpreted within an ecological
and evolutionary framework. Tail elongation in widowbirds may
serve as an example to illustrate the problem: In his famous study
on the effect of the elongation of tails in long-tailed widowbirds
(Euplectes progne), Andersson (1982) added about 50% to the average
Behavioral Ecology
tail of 50 cm and found that males with such elongated tails were
more successful in recruiting females onto their territories than control males. In contrast, the doses that are commonly used to manipulate testosterone may lead to a short-term increase in testosterone
concentrations (i.e., for a few days) that may be 10–20 times higher
than average levels experienced by female birds (Fusani 2008a;
Quispe-Valdez R, Goymann W, unpublished data). This represents
the equivalent of a long-tailed widowbird tail that would be 5–10
m long. We doubt that such an over-exaggerated long tail would
allow drawing any firm conclusion about sexual or natural selection
(some may find this a lame comparison because as hormone receptors become saturated any additional elevation in hormone levels
may be neutral. However, hormone dynamics can be complicated,
e.g., Hews and Moore 1997; Adkins-Regan 2005, p. 37f; thus, the
comparison may not be so lame at all).
Indeed, there are some studies that looked at the relationship
between aggressive behavior and natural testosterone levels in freeliving females. Testosterone levels of females sampled after experimental encounters with female decoys increased in some instances
(Desjardins et al. 2006; Gill et al. 2007; Ross and French 2011), but
not in others (Elekonich 2000; Davis and Marler 2003; Rubenstein
and Wikelski 2005; Jawor et al. 2006; Navara et al. 2006; Goymann
et al. 2008). Although the adaptive significance of changes in testosterone during behavioral interactions is still unclear, the presence
or absence of a change in testosterone concentrations in female
birds depends on the species and the behavioral context (see also
Gill et al. 2007). Experimental studies have shown that—in female
birds—testosterone implants may induce song and increase aggression in a similar manner as in males of some species (reviewed by
Staub and de Beer 1997; Fusani et al. 2003; Ketterson et al. 2005;
Sandell 2007). Yet, as discussed above, the testosterone levels generated by such implants typically by far exceed the levels present in the
circulation of unmanipulated birds and may be of limited meaning in an ecological and evolutionary context. In female European
robins (Erithacus rubecula), treatment with an antiandrogen did not
affect territorial aggression (Kriner and Schwabl 1991). A recent
study in dark-eyed juncos (Junco hyemalis) suggests that the endogenous capacity to produce testosterone weakly correlates with the
latency of aggressive behavior toward a same-sex-simulated territorial intrusion in females (Cain and Ketterson 2012). Currently, this
study (in combination with unpublished data from blackbirds Turdus
merula; Miranda C, Goymann W, Partecke J, unpublished data)
probably presents the best evidence for a potential role of plasma
testosterone in the control of aggressive behavior in a female bird
within an ecologically relevant context. Studies in mammals suggest
that aggression may be the default state in females and hence does
not need to be activated by hormones. Rather, steroid hormones
may be involved in suppressing aggression during the fertile phase
of females (e.g., Floody 1983). In female mice, progesterone modulates aggression induced by estradiol and testosterone (Albert et al.
1992). Also in black coucals (Centropus grillii), a sex-role-reversed
bird species in which females compete for territories, progesterone
modulates aggression (Goymann et al. 2008). However, this may
not be completely independent of androgens, as female black coucals express more androgen receptors in parts of the brain that are
responsible for social and agonistic behavior than males (Voigt and
Goymann 2007). In female mountain spiny lizards (Sceloporus jarrovi),
estradiol may play the main role in modulating aggressive behavior
(Woodley and Moore 1999a, 1999b). In spotted hyenas (Crocuta crocuta), where females are more aggressive than males, there is mixed
evidence about the importance of testosterone in mediating female
Goymann and Wingfield • Male-to-female testosterone ratios, dimorphism, and life history
dominance during ontogeny: Dloniak et al. (2006) found a significant correlation of fecal androgen metabolites of mothers during
late gestation with the aggression of cubs at 2–6 months of age,
but East et al. (2009) convincingly demonstrated that competitive
ability of offspring at adulthood was best explained by postnatal
maternal behavioral support rather than exposure to maternal
hormone concentrations during gestation. At present, there is little
evidence that female dominance and aggression is activated by testosterone in adult spotted hyenas (see e.g., Goymann et al. 2001).
Also in ring-tailed lemurs (Lemur catta), females are dominant over
males. Agonistic interactions and levels of sex steroids peak during the breeding season, but a causal association between levels
of testosterone and aggression has also not been established (von
Engelhardt et al. 2000; Drea 2007). French et al. (2013) list further
examples of “atypical” mammals in which females are either more
aggressive than or dominant over males, but a clear role of androgens as mediators of “atypical” behavior remains to be established.
695
Current evidence suggests that, in mammals, androgens are more
likely to play an organizational role in this respect rather than an
activational role (see examples in French et al. 2013). Thus, overall, there is limited evidence that changes in physiological levels of
plasma testosterone have a strong impact on aggression in female
vertebrates (see also Staub and de Beer 1997). If androgens play
a role in the modulation of female morphology or behavior, their
regulation may occur on the target tissue level, that is, the local sensitivity to testosterone may be enhanced rather than plasma levels
of testosterone (e.g., Voigt and Goymann 2007).
Future challenges for comparative studies in
hormone physiology
Neural and endocrine regulatory mechanisms are highly conserved
across vertebrates (Wingfield 2005). Nonetheless, there are diverse
ways in which this system can be adjusted to customize responses
of the individual, male or female. Hormones serve as the signaling
Figure 5
Potential control mechanisms for avian plumage and skin color in birds. There are 3 major components to control mechanisms: the regulation of hormone
secretion from the hypothalamic–pituitary–gonadal axis (left-hand part of the figure), transport of hormones such as testosterone or estradiol in the blood
(lines in red), and the mechanisms associated with action of the hormone in the target cell, in this case, a skin or feather follicle cell (central part of the figure).
The net result is the regulation of plumage and skin traits (right-hand part of the figure). This 3-part system of control of hormone secretion, transport,
and effects on target organs is an important concept because it provides many points of potential regulatory mechanisms. The secretion component on the
left summarizes how sensory information is transduced through neurotransmitter, for example, glutamate (NMDA) and neuroendocrine secretions such as
gonadotropin-releasing hormone (GnRH) and gonadotropin-inhibiting hormone (GnIH) into release of gonadotropins luteinizing hormone (LH), folliclestimulating hormone (FSH—not shown), and melanocyte-stimulating hormone (MSH) from the anterior pituitary into the blood. LH circulates to the gonad
where it acts on cells that express steroidogenic enzymes to stimulate secretion of sex steroids such as testosterone (T) or estradiol (E2) that are in turn
released into the blood. Sex steroids have negative on neuroendocrine and pituitary secretions (not shown). In birds, circulating T and E2 bind weakly to
corticosterone-binding globulin (CBG) before entering a target cell. Once T has entered a target cell, it has 4 potential fates. First, T can bind directly to
the androgen receptor (AR), a member of the type 1 genomic receptors that become gene transcription factors once they are bound to T. Second, T can be
converted to E2 by the enzyme aromatase (aro). E2 can then bind to either estrogen receptor alpha (Erα) or estrogen receptor beta (Erβ) both of which are
genomic receptors that regulate gene transcription, but different genes from those regulated by AR. Third, T can be converted to 5alpha-dihydrotestosterone
(5α-DHT) that also binds to AR and cannot be aromatized, thus enhancing the AR gene transcription pathway. Fourth, T can be converted to 5betadihydrotestosterone (5β-DHT) that binds to no known receptors and also cannot be aromatized, indicating a deactivation shunt. A complex system of
corepressors and coactivators of genomic steroid receptor action are also known. The end result is regulatory action in the skin or feather follicle cell. Even
in species in which the gonads do not affect plumage dimorphism, it is possible that sex steroids may play a role, as they can be produced locally: steroid
metabolizing enzymes are present in skin, see for example, Schlinger et al. (1989).
Behavioral Ecology
696
molecules and circulate in the blood. So far, comparative studies
in hormone physiology have focused on these circulating hormone
concentrations and related these hormone levels to the life history,
the environment, or other traits of interest. The above examples for
a potential role of testosterone in the regulation of traits in females
made clear that in the future, it will be important to implement the
concept that there is more than one way that a hormone can control
a trait. Basically, there are 3 major components to endocrine control
systems (Wingfield 2005; Hau and Wingfield 2011). First is the regulation of hormone secretion from perception of the environmental
stimulus and transduction by the brain, which results in release of
neuroendocrine signals from the hypothalamus. This triggers the
release of tropic hormones from the anterior pituitary gland and, in
turn, peripheral endocrine secretions that regulate morphological,
physiological, and behavioral responses (see Figure 5 for an example). Second, the transport of many hormones (e.g., steroids and
thyroid hormones) involves binding to carrier proteins (e.g., Breuner
and Orchinik 2002). Finally, once the hormone signal arrives at a
target cell, there are multiple fates of that hormone that can have
profound influences on the type of response (e.g., Wingfield 2005).
In some cases, the target cell may express enzymes that deactivate the hormone before it can interact with a receptor or change
it to another form that may interact with a very different receptor
(Figure 5). All of these components of the perception–transduction–response axes and the resultant cascade of hormone actions
are sites of regulation of how an individual can respond to signals
from the physical and social environments. Furthermore, combinations of regulatory points enable highly diverse ways for individuals
to respond to similar environmental cues. These may be the bases
of species and sex differences in several species studied. Future techniques will have to be developed to assess all of these components
simultaneously and implement them in comparative studies.
Conclusions
Our analysis has shown that relating sexually dimorphic traits to
testosterone in birds is problematic because such traits can evolve
for reasons other than sexual selection and may not necessarily be
controlled by sex steroids. Thus, for a proper comparison, it would
be necessary to know whether plumage and body size dimorphisms
are related to sexual selection and whether the molt of a species
is under androgenic control. Further, pharmacological manipulations of testosterone that lead to the expression of male-like traits
in females have generated the assumption that secondary sexual
traits are regulated in a similar manner in females and males.
However, to draw firm conclusions about the role of testosterone
in the expression of secondary sexual traits in females within an
ecophysiological and evolutionary context, it will be essential to
manipulate testosterone within the physiological range and during
various stages of ontogenetic development.
Overall, the relationship of male-to-female testosterone may not
be a good predictor of sex differences in intra- and intersexual competition. Females are likely to use other means to regulate aggression or sexually dimorphic traits such as a colorful plumage. The
regulation may occur at other stages of the perception–transduction–response system of hormones or females may rely on completely different mechanisms than males (see Gahr et al. 2008 for an
illustrative example). The discussion above shows that meta-analyses
of hormone levels face the challenge that the action of hormones is
regulated at different times and at different levels (Wingfield 2012)
and that sexually dimorphic traits can be modulated by interactions
of physiology, genetics, and the environment (Table 4). The degree
of regulation at each of these different levels may differ between
species, between the sexes, and even between individuals (giving rise
to different phenotypes). This very much complicates meta-analyses
that focus on just one time point or level (i.e., the level of hormone
secretion during adulthood in the current case), but we are on the
verge of new technologies that will allow us to address the various
levels of regulatory mechanisms (e.g., Pavey et al. 2012).
Supplementary Material
Supplementary material can be found at http://www.beheco.
oxfordjournals.org/
Funding
J.C.W. is grateful for support from grant number IOS-0750540 from
the National Science Foundation and the Endowed Professorship in
Physiology from the University of California, Davis. W.G. is grateful for funding from M. Gahr and the Max-Planck Gesellschaft.
This article is dedicated to W. Wickler on occasion of his 82nd birthday. We
would like to thank L. Fusani, M. Hau, K. Hirschenhauser, E. Ketterson,
and 2 anonymous referees who critically read previous versions of this
manuscript and whose comments greatly improved these earlier drafts.
Furthermore, we would like to thank S. Nakagawa for suggesting the use
of Egger’s regression test and practical help in implementing it to the
MCMCglmm models.
Forum editor: Shinichi Nakagawa
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