Testosterone production, sexually dimorphic morphology, and digit ratio in the dark-eyed junco

This article is based on the March 2013 paper entitled “Testosterone production, sexually dimorphic morphology, and digit ratio in the dark-eyed junco,” the authors are K.E. Cain, C.M. Bergeon Burns, and E.D. Ketterson, and it is found in Vol. 24 of Behavioral Ecology pages 462-469.


A dark-eyed junco (Junco hyemalis) image from: http://www.arkive.org/dark-eyed-junco/junco-hyemalis/


Hormones produced during development may have an effect on morphological, behavioral, and physiological characteristics in adults (Trainor and Marler 2001), partly because of the hypothalamic-pituitary-gonadal (HPG) axis (Pfannkuche et al. 2011).  In this HPG axis, signals are sent to the hypothalamus, inducing more gonadotropin-releasing hormone (GnRH) to be released (Burns et al. 2014).  GnRH activates secretions of gonadotropins, such as luteinizing hormone (LH), from the anterior pituitary gland, which subsequently acts at the gonad, promoting the production and release of steroids, including testosterone from the gonads (Burns et al. 2014).


The hypothalamic-pituitary-gonadal (HPG) axis in males (left) and females (right).  The system regulates development, reproduction, and aging.  It relies on complicated negative feedback loops.  Click on images to enlarge.  These two images are from: http://geekymedics.com/2011/02/27/how-the-gonadal-axis-works/

The gonadal steroid testosterone has extensive effects on many morphological, physiological, and behavioral features throughout the brain and beyond it, bolstering characteristics, such as sperm manufacture, aggression, sexual behaviors, and decorations (Burns et al. 2014).  Appropriately, how large the gonad is and levels of testosterone usually vacillate seasonally along with these phenotypes (Burns et al. 2014).

The ratio of the length of digit 2 to digit 4 (2D:4D) has been demonstrated to mirror the ratios of estrogen:androgens sustained during development (Zheng and Cohn 2011).  2D:4D corresponds to hormone level because there are variations in androgen and estrogen receptor quantity in the digits, and steroids act to turn on those receptors, so whether digit growth is activated or hindered is affected (Zheng and Cohn 2011).

GnRH challenges, in which subjects are given a typical amount of exogenous GnRH, basically bypass the brain so downstream ability to secrete androgens can be quantified (Burns et al. 2014).

Question: How do testosterone levels affect hormonal phenotype and physical features of adult male and female dark-eyed juncos?  This is an important question because dark-eyed juncos can be used as a model organism to explain human hormonal systems.

Objectives of Study:

  • To investigate any correlations between individual differences in endogenous hormone exposure and adult phenotype, using 2D:4D as a substitute for total developmental hormone exposure:
    • To probe the relationship between male 2D:4D and 2 endogenous aspects of adult hormonal phenotype:

1. Testosterone levels in reaction to a physiological activation of the HPG axis (a standardized injection of GnRH)

2. Testosterone levels before this test

    • To study the relationships between 2D:4D and sexually dimorphic features of body size (wing, tail, and tarsus length) in males and females.


1. Endogenous testosterone production and 2D:4D:

There was no correlation between 2D:4D and testosterone prior to the challenge.

There was a significant negative correlation between postchallenge testosterone and 2D:4D (Fig. 1).

Figure 1

Figure 1 (from Cain et al. 2013). Scatterplot for showing the relationship between 2D:4D and the level of testosterone made in reaction to physiological challenge with GnRH. Curved lines are 95% confidence interval.

2. Dimorphic morphology and 2D:4D:

In males, the correlation between 2D:4D and morphology was solidly reliant on how old the bird was.  Among males that were second-year or older, tail, wing, and tarsus measurements were all positively correlated with 2D:4D (Fig. 2.).  On the other hand, among males that were first-year, the correlations between 2D:4D and morphology were all negative (Fig. 2).

All females were second-year or older and displayed comparable correlations between 2D:4D and morphology with the second-year or older males.  Female 2D:4D was positively correlated with tarsus, wing, and tail length, but only the tarsus results were significant (Fig. 2).

Figure 2f

Figure 2 (from Cain et al. 2013). Scatterplots with regression lines showing the age- and sex-specific relationships between 2D:4D and phenotypic features. The left panel illustrates male wing length and 2D:4D. Closed diamonds and solid line refer to second-year or older males; open squares and dashed line refer to first-year males. The right panel represents female 2D:4D and tarsus length.


1. Endogenous hormone production:

The results show that exposing the junco to large amounts of androgens, or small amounts of estrogens, leads to lessened HPG responsiveness in adults.

A weakness of the study was that the authors were unsuccessful in finding a correlation between 2D:4D and testosterone before the HPG challenge.  Even though natural differences in developmental exposure to hormones may change individual capacity to make testosterone in reaction to activation of the HPG axis, the effect that such differences have on testosterone concentration when activation is not present is weakened.

2. Digit ratios and sexually dimorphic structures:

Higher levels of testosterone during ontogeny (developmental history of an organism) can influence the later expression of sexually specific adornments, or sexually dimorphic characteristics.  Other studies have found connections between 2D:4D and sexually dimorphic features, though some have discovered positive correlations, and others negative, so there are contradictory findings.

A weakness of this investigation was that only second-year or older females were examined (it would have been more comprehensive if they also included first-year females in their experiment for comparison).


Their results provide evidence that there is a possibility that endogenous differences in the developmental hormone environment can have long-lasting effects on adult hormonal phenotype and physical morphology.  Unraveling how endogenous individual differences in hormonal exposure during ontogeny moderates adult phenotype is a significant objective for scientists with a desire to comprehend the evolution of hormonally affected phenotypes and sexually dimorphic characteristics and behavior.


Burns CMB, Rosvall KA, Hahn TP, Demas GE, Ketterson ED. 2014. Examining sources of variation in HPG axis function among individuals and populations of the dark-eyed junco. Horm Behav. 65:179-187.

Cain KE, Burns CMB, Ketterson ED. 2013. Testosterone production, sexually dimorphic morphology, and digit ratio in the dark-eyed junco. Behavioral Ecology. 24:462-469.

Pfannkuche KA, Gahr M, Weites IM, Riedstra B, Wolf C, Groothuis TGG. 2011. Examining a pathway for hormone mediated maternal effects—yolk testosterone affects androgen receptor expression and endogenous testosterone production in young chicks (Gallus gallus domesticus). Gen Comp Endocrinol. 172:487–493.

Trainor B, Marler C. 2001. Testosterone, paternal behavior, and aggression in monogamous California mouse (Peromyscus californicus). Horm Behav. 40:32–42.

Zheng Z, Cohn MJ. 2011. Developmental basis of sexually dimorphic digit ratios. Proc Natl Acad Sci USA. 108:16289–16294.

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