Testosterone synthesis in the female songbird brain

https://doi.org/10.1016/j.yhbeh.2020.104716Get rights and content

Highlights

  • Estradiol (E2) acutely increases in the female songbird auditory cortex during song.

  • Testosterone (T) rises in response to song when its conversion to E2 is blocked.

  • This rise in T only happens in the left hemisphere and does not happen in the blood.

  • This rise is abolished when the production of T is locally blocked.

  • The female brain thus produces T in response to socially-relevant stimuli.

Abstract

Decades of work have established the brain as a source of steroid hormones, termed ‘neurosteroids’. The neurosteroid neuroestradiol is produced in discrete brain areas and influences cognition, sensory processing, reproduction, neurotransmission, and disease. A prevailing research focus on neuroestradiol has essentially ignored whether its immediate synthesis precursor - the androgen testosterone - is also dynamically regulated within the brain. Testosterone itself can rapidly influence neurophysiology and behavior, and there is indirect evidence that the female brain may synthesize significant quantities of testosterone to regulate cognition, reproduction, and behavior. In songbirds, acoustic communication is regulated by neuroestrogens. Neuroestrogens are rapidly synthetized in the caudomedial nidopallium (NCM) of the auditory cortex of zebra finches in response to song and can influence auditory processing and song discrimination. Here, we examined the in vivo dynamics of NCM levels of the neuroestrogen synthesis precursor, testosterone. Unlike estradiol, testosterone did not appear to fluctuate in the female NCM during song exposure. However, a substantial song-induced elevation of testosterone was revealed in the left hemisphere NCM of females when local aromatization (i.e., conversion to estrogens) was locally blocked. This elevation was eliminated when local androgen synthesis was concomitantly blocked. Further, no parallel elevation was observed in the circulation in response to song playback, consistent with a local, neural origin of testosterone synthesis. To our knowledge, this study provides the first direct demonstration that testosterone fluctuates rapidly in the brain in response to socially-relevant environmental stimuli. Our findings suggest therefore that locally-derived ‘neuroandrogens’ can dynamically influence brain function and behavior.

Significance statement

This study demonstrates that androgen synthesis occurs rapidly in vivo in the brain in response to social cues, in a lateralized manner. Specifically, testosterone synthesis occurs within the left secondary auditory cortex when female zebra finches hear male song. Therefore, testosterone could act as a neuromodulator to rapidly shape sensory processing. Androgens have been linked to functions such as the control of female libido, and many steroidal drugs used for contraception, anti-cancer treatments, and sexual dysfunction likely influence the brain synthesis and action of testosterone. The current findings therefore establish a clear role for androgen synthesis in the female brain with implications for understanding neural circuit function and behavior in animals, including humans.

Introduction

Studies on rapid regulation of neurosteroids have focused almost exclusively on the potent estrogen 17β-estradiol (E2). E2 is produced via the local aromatization of testosterone within many brain regions, and influences major functions and behaviors including social communication (Ball and Balthazart, 2010; Balthazart et al., 2009; Garcia-Segura, 2008; Hull and Dominguez, 2015; Yoder and Vicario, 2012). The local dynamics and acute actions of E2 on social communication are most evident in songbirds.

One region of songbird secondary auditory cortex (caudomedial nidopallium; NCM) is enriched with estrogen receptors and aromatase (Metzdorf et al., 1999; Yoder and Vicario, 2012). Both male and female zebra finches exhibit a rapid elevation in NCM E2 in social and auditory contexts (Remage-Healey et al., 2008, Remage-Healey et al., 2012). Increases in local E2 also directly enhance the auditory-evoked activity of NCM neurons (Remage-Healey et al., 2010; Remage-Healey and Joshi, 2012; Tremere et al., 2009; Tremere and Pinaud, 2011), while infusion of fadrozole, a potent and selective aromatase inhibitor, causes acute decreases in the auditory-evoked activity of NCM neurons and the behavioral preference for familiar songs (Remage-Healey et al., 2010). Therefore, local E2 acts as a neuromodulator to directly influence both local neuronal response properties and downstream sensorimotor integration (Remage-Healey et al., 2013; Vahaba and Remage-Healey, 2018). The rapid fluctuations in local E2 levels in the songbird brain predict that upstream precursors like testosterone also exhibit dynamic regulation.

Theoretically, song-induced elevations in NCM E2 can depend on three related mechanisms, including (a) a rapid increase of aromatase activity, (b) a rise of the precursor testosterone in the plasma and diffusion into the brain and/or (c) a direct local testosterone synthesis independent from the periphery. Rapid changes in aromatase activity have been well demonstrated in the avian brain (Balthazart et al., 2006; Cornil and Bournonville, 2017) including the telencephalon of zebra finches (Remage-Healey et al., 2009). Instead, this study focused specifically on the substrate fluctuation/synthesis hypotheses. This current work was conducted using female zebra finches for three reasons. First, females have lower peripheral testosterone levels than do males (Kabelik et al., 2011; Prior et al., 2014). Second, the key enzymes responsible for testosterone synthesis, 3β-hydroxysteroid-dehydrogenase (3β-HSD) and 17β-hydroxysteroid-dehydrogenase (17β-HSD), are expressed and active in the female NCM, exceeding that in males (London et al., 2003, London et al., 2006; Soma et al., 2004; Tomaszycki and Dzubur, 2013). Third, this high androgenic enzymatic activity is rapidly regulated during stress in females but not males (Soma et al., 2004). Therefore, testosterone synthesis in the female NCM could have a functional behavioral role in addition to providing a substrate for fast conversion to estrogens.

Section snippets

Animals

Females zebra finches (adults > 105 dph) were used for this study. Protocols were approved by the Institutional Animal Care and Use Committee at the University of Massachusetts. All animals were raised from our colony or came from a commercial supplier.

General procedures

Three main experiments were performed using in vivo microdialysis or blood measurements for steroids according to previously published protocols (Remage-Healey et al., 2008, Remage-Healey et al., 2012). All animals were housed in acoustically

Validations of the testosterone assay on NCM dialysates

Testosterone was measured in NCM dialysates per previously published protocols (Remage-Healey et al., 2008). Here we used sequential assays of both steroids in the same samples and drugs that can interfere with these measurements, five validation steps were thus necessary.

First, the fidelity of both the testosterone-only and the sequential assays were compared. Increasing concentrations of testosterone were added to control aCSF or to the mix that came from the previous E2 assay. This mix

Discussion

To our knowledge, this study provides the first evidence that testosterone is locally synthesized in the brain, in vivo. Our findings also clarify rapid neuroestrogen fluctuations in the forebrain. Neuroestrogens can fluctuate in several species, including within the hippocampus and hypothalamus (Kenealy et al., 2013; Sato and Woolley, 2016). Our findings replicated a song induced-elevation in local neuroestrogen levels that was previously observed in the NCM of male (Remage-Healey et al., 2008

Conclusions

Our findings demonstrate that testosterone is actively synthetized in the female brain and suggest that neuroandrogens might thus regulate brain function and behavior. These data focus attention on reproduction and cognition for neuroandrogen actions (Davison and Davis, 2003; Pluchino et al., 2015). It is also now important to evaluate the translational implications of androgen synthesis in the female brain, since steroidal drugs are used by women for sexual desire disorder and anti-cancer

Declaration of competing interest

None.

Acknowledgements

This work was supported by National Institutes of Health - Grant R01NS082179 (to LRH). CdB was supported by a Belgian American Education Foundation (BAEF) fellowship.

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    Present address: GIGA Neuroscience, University of Liege, Liege, B-4000, Belgium.

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