Tamoxifen activates hypothalamic l-dopa synthesis to stimulate ovarian estrogen production in chicken

https://doi.org/10.1016/j.bbrc.2018.01.182Get rights and content

Highlights

  • Blocking estrogen action by tamoxifen resulted in elevated estrogen levels.

  • Tamoxifen could activate hypothalamus dopaminergic signaling.

  • l-dopa could stimulate ovarian estrogen synthesis.

Abstract

Estrogen is the primary sex hormone responsible for the development and modulation of the female reproductive system in all vertebrates including avian species. The actions of estrogen are mediated by the estrogen receptor, which could be modulated by the selective estrogen receptor modulator tamoxifen (TAM). In this study, we administered TAM into the actively laying chicken to investigate the ovarian and hypothalamic responses to the estrogen action blockage. The laying was disrupted and the development of the pre-ovulatory hierarchical follicles was arrested. However, the TAM treatment caused an increase of estrogen level in both serum and ovary. Among the main estrogen targeted tissues, the hypothalamus showed specific dopaminergic activation as indicated by gene expression analysis. In the ovary, l-dopa, the precursor of dopamine, could stimulate the estrogen synthesis in undifferentiated follicles but not in the differentiated pre-ovulatory follicles. Thus, we established a feedback loop links ovarian estrogen production with hypothalamic l-dopa synthesis and we propose that the dopamine is involved in estrogen action to regulate the ovarian follicle development and ovulation.

Introduction

Avian species are an important comparative vertebrate model for the research in endocrinology and reproductive biology. The chicken is also one of the most important agricultural animals, producing 1.1 trillion eggs per year [1]. Thus, studying the physiology of chicken reproduction could benefit both the understanding of basic reproductive biology and the improvement of food industry.

As in other vertebrates, the female avian reproductive systems are primarily regulated by the hypothalamus-pituitary-gonadal axis [2]. The hypothalamus produces gonadotropin-releasing hormone (GnRH) that stimulates pituitary production of follicle-stimulating hormone (FSH) and luteinizing hormone (LH), which in turn direct the ovarian function and reproductive activity. Ovarian derived steroids, predominantly estrogen and progesterone, return to the central nervous system through the bloodstream and provide a feedback to the hypothalamic GnRH production and release. It is well known that in birds, estrogen plays a central role in the female reproduction by stimulating ovary and follicle development [3,4]. The effects of estrogen in the target cells are mediated by estrogen receptors (ER), which are expressed not only in the hypothalamus and pituitary but also in ovary and oviduct of chicken. Once activated by estrogen, the ER is able to translocate into the nucleus and bind to DNA to regulate the activity of different genes [5].

Tamoxifen (TAM) is a synthetic estrogen antagonist that could inhibit the transcription of target genes by binding to ER and has been widely used in the therapy for cancer patients with ER-positive tumours [6]. Previous studies in birds have confirmed that TAM could bind to chicken ER with high affinity and limit the transcription of estrogen-induced genes [[7], [8], [9]]. Therefore, TAM was frequently employed to investigate the roles of ER in different chicken tissues by loss-of-function means.

Here we show that TAM treatment in reproductive active chicken resulted in the disruption of laying, which may mainly attribute to the lowered FSH and LH. In contrast, the estrogen levels increased in both ovary and serum. Genome-wide transcriptome analysis of hypothalamus revealed that the hypothalamic l-dopa synthesis was enhanced by TAM treatment. Furthermore, we provide novel evidence that l-dopa could stimulate estrogen production in the undifferentiated but not differentiated follicles in the ovary. We reason that blocking ER leads to enhanced hypothalamic l-dopa synthesis, which in turn stimulates estrogen production in the ovary.

Section snippets

Animals and reagents

Sexually mature (older than 23 weeks) Hy-Line Brown hens were collected from a local research farm affiliated with Taishan University. All animals had free access to water and feed. The hens were housed in separate cages under a daily light period of 14 h and the laying events were recorded by checking the cage every 2 h during the light period to determine the regular laying sequence. For the tamoxifen (TAM) experiment, the control group (n = 6) animals were treated subcutaneously with vehicle

Tamoxifen treatment blocks ovulation and inhibits follicle development

Daily treatment of laying hens with tamoxifen (TAM) caused a gradual decrease of the egg-laying rate from the first day of the experiment. On day 6, all the animals in the TAM treated group stopped laying whereas the control animals continue to lay normally (Fig. 1A). All the animals were sacrificed on day 7 of the experiment. The normal mature chicken ovary was mainly composed of the stroma tissue containing small white and small yellow follicles (undifferentiated follicles) and hierarchical

Discussion

In chicken, the tamoxifen (TAM), are very potent antagonists of estrogen action and manifest very little agonistic activity compared to their action in other species [14]. Therefore, it has been frequently used as loss-of-function means to investigate estrogen action in birds [[15], [16], [17]]. In consistent with previous study, treatment of hens with TAM resulted in cessation of laying and reduction in the number and weight of the hierarchical follicles [18,19]. Our data indicate that this

Acknowledgements

This study was supported by the National Natural Science Foundation of China (31301974) and the Natural Science Foundation of Shandong Province (ZR2013CQ002).

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  • 1

    These authors contributed equally to this work.

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