Molecular Neurobiology

, Volume 56, Issue 1, pp 394–405 | Cite as

Role of HPA and the HPG Axis Interaction in Testosterone-Mediated Learned Helpless Behavior

  • Birgit Ludwig
  • Bhaskar Roy
  • Yogesh DwivediEmail author


Affective disorders show sex-specific differences in prevalence, symptoms, and complications. One hypothesis for this discrepancy is the interaction between the hypothalamic-pituitary-adrenal (HPA) axis and hypothalamic-pituitary-gonadal (HPG) axis. The present study investigates the influence of androgen on the behavioral phenotype and explores how it interacts with HPA axis genes. Gonadectomized (GDX) and GDX rats treated with testosterone propionate (T) were tested for learned helplessness (LH) behavior and compared with tested controls (TC). Prefrontal cortex was used for analyses of HPG- axis-related genes (androgen receptor, (Ar); estrogen receptor-β (Er-β)) and HPA axis-related genes (corticotropin-releasing hormone, (Crh); glucocorticoid receptor, (Nr3c1); corticotropin-releasing hormone receptor 1, (Crhr1); corticotropin-releasing hormone receptor 2, (Crhr2); FK506 binding protein 5, (Fkbp5)). Promoter-specific CpG methylation in the Crh gene was determined by bisulfite sequencing. Chromatin immunoprecipitation (ChIP) assay was used for determining ER-β binding on the proximal promoter region of Crh gene. Serum testosterone levels confirmed a testosterone-depleted GDX group, a group with supraphysiological levels of testosterone (T) and another group with physiological levels of testosterone (control (C)). Unlike GDX rats, T group exhibited significantly higher LH score when compared with any other group. Crh and Fkbp5 genes were significantly upregulated in GDX group compared with controls, whereas Er-β showed a significant downregulation in the same group. Methylation analysis showed no significant differences in-between groups. ChIP assay was unable to determine a significant change in ER-β binding but revealed a notable contrast in Crh promoter occupancy between T and GDX groups. Altogether, the present study reveals an increased susceptibility to depression-like behavior due to chronic supraphysiological level of androgen via HPA axis inhibition.


Hypothalamic-pituitary-adrenal axis Hypothalamic-pituitary-gonadal axis Stress Depression Testosterone Androgen receptor 



Hypothalamic-pituitary-adrenal axis


Hypothalamic-pituitary-gonadal axis




Testosterone propionate


Learned helplessness


Tested controls


Androgen receptor


Estrogen receptor-β


Corticotropin-releasing hormone


Glucocorticoid receptor


Corticotropin-releasing hormone receptor


FK506 binding protein 5


Chromatin immunoprecipitation


Prefrontal cortex


Postnatal day


Enzyme-linked immunosorbent assay


Escape test


Inescapable shock




Quantitative polymerase chain reaction


Glyceraldehyde 3-phosphate dehydrogenase


Complementary DNA


Phosphate buffered saline


Proteasomal inhibitors


Revolution per minute


Tris-ethylenediaminetetraacetic acid


Lithium chloride


Forced swim test


Anabolic androgenic steroid


5-Alpha-androstane 3beta,17beta diol


Estrogen receptor elements


cAMP-response elements


Activator protein-1


Periventricular nucleus


Funding Information

The research was supported by grants from the National Institute of Mental Health (R01MH082802, 1R01MH101890, R01MH100616, 1R01MH107183-01) to Dr. Dwivedi. The sponsoring agency had no role in study design, collection, analysis, interpretation of data, and in the writing of the manuscript.

Compliance with Ethical Standards

Research Involving Animals

This research involves the use of animals. The study was approved by the Institutional Animal Care and Use Committee of the University of Alabama at Birmingham.

Conflict of Interest

The authors declare that they have no conflict of interest.

Supplementary material

12035_2018_1085_MOESM1_ESM.png (24 kb)
Fig. S1 A schematic overview representing the groupwise distribution of rats under the present study. A total of 30 Long Evans rats were randomly assigned to treatments (GDX = 10 rats having undergone gonadectomy, T = 10 rats having undergone gonadectomy and daily supraphysiological testosterone injections, C = 10 naive controls). For behavioral testing, all 30 rats were further randomly assigned to be restrained only (tested controls (TC)) or restrained and exposed to inescapable shocks (IS). (PNG 23 kb)
12035_2018_1085_MOESM2_ESM.docx (24 kb)
Fig. S2 Methylation of Crh gene promoter. a The sequence represents 420 bp upstream region of rat Crh promoter with CpG sites highlighted, indicating their position relative to transcription start site (right arrow). b The checker board represents methylation status of individual CpG site as identified from each animal considering individual groups, all methylated cytosine (C) not converted thymine (T) are presented with double-plus symbol. Whereas, identification of both C and T peaks is represented with plus symbol, this indicates potentially incomplete bisulfite conversion. (DOCX 24 kb)
12035_2018_1085_MOESM3_ESM.docx (17 kb)
Table S1 (DOCX 17 kb)


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Authors and Affiliations

  1. 1.Department of Psychiatry and Behavioral NeurobiologyUniversity of Alabama at BirminghamBirminghamUSA

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