Morphological and molecular effects of cortisol and ACTH on zebrafish stage I and II follicles

Abstract

Oogenesis in zebrafish (Danio rerio) is controlled by the hypothalamus-pituitary-gonadal axis and reproductive hormones. In addition, an interference of stress hormones is known with reproductive biology. In the presented work, we aimed to explore the hypothesis that cortisol (Cort) and ACTH may affect early oogenesis in zebrafish, given the presence of the specific receptors for glucocorticoids and ACTH in the zebrafish ovary. Follicles at stages I and II were exposed in vitro to 1 mM Cort and ACTH for 48 h, then ultrastructural and molecular effects were analyzed. The comet assay demonstrated increased tail moments for Cort and ACTH treatment indicative of DNA damage. The mRNA expression of apoptotic genes (bax, bcl-2) was not altered by both treatments, but Cort increased significantly the expression of the ACTH receptor (mc2r). Cort stimulated the presence of the endoplasmic reticulum, predominantly at stage II, while ACTH induced a strong vacuolization. Viability of oocytes was not affected by both treatments and fluorescent staining (monodansylcadaverine/acridine orange) indicated a reduced quantity of autophagosomes for ACTH, and lower presence of nucleic acids in ooplasm for Cort and ACTH. Concluding, different responses were observed for stress hormones on early stages of zebrafish oocytes, which suggest a role for both hormones in the stress-mediated adverse effects on female gametogenesis. Reproduction (2015) 150 429-436

Maternal stress and fish reproduction: The role of cortisol revisited

“…The effect of cortisol on theca and granulosa cells during this period may lead to changes in glucocorticoid‐responsive genes, including 11β‐hydroxysteroid dehydrogenase (11β‐HSD2) (Lubzens et al., ); however, the extent of cortisol signalling and the downstream effects are far from clear. Exposure of stage II oocytes to cortisol caused an increase in MC2R; however, no change in GR was noted (Sousa et al., ). TEM micrographs of stage I follicles showed no morphological alterations with the addition of cortisol, whereas stage II exposure significantly stimulated the presence of the rough endoplasmic reticulum in the theca and granulosa cells, suggesting increased protein synthetic capacity for steroidogenesis (Sousa et al., ).…”

“…Exposure of stage II oocytes to cortisol caused an increase in MC2R; however, no change in GR was noted (Sousa et al., ). TEM micrographs of stage I follicles showed no morphological alterations with the addition of cortisol, whereas stage II exposure significantly stimulated the presence of the rough endoplasmic reticulum in the theca and granulosa cells, suggesting increased protein synthetic capacity for steroidogenesis (Sousa et al., ). Apart from cortisol‐mediated increases in protein synthesis, this steroid also appeared to increase nucleic acid instability in stage II follicles (Sousa et al., ), suggesting a role for this stress steroid in follicular apoptosis.…”

“…TEM micrographs of stage I follicles showed no morphological alterations with the addition of cortisol, whereas stage II exposure significantly stimulated the presence of the rough endoplasmic reticulum in the theca and granulosa cells, suggesting increased protein synthetic capacity for steroidogenesis (Sousa et al., ). Apart from cortisol‐mediated increases in protein synthesis, this steroid also appeared to increase nucleic acid instability in stage II follicles (Sousa et al., ), suggesting a role for this stress steroid in follicular apoptosis. As cortisol concentration used in the study was 1uM (320 ng/ml), and the follicles were exposed for 48 hr to this high concentration, the observed responses may be supraphysiological.…”

“…As cortisol concentration used in the study was 1uM (320 ng/ml), and the follicles were exposed for 48 hr to this high concentration, the observed responses may be supraphysiological. However, the results suggest that a high cortisol concentration in the ovarian follicles during early oogenesis may lead to abortion in both fish (Sousa et al., ) and mice (Yuan et al., ), as well as lead to ovarian atresia in fish (Clearwater & Pankhurst, ).…”

Zebrafish: A Versatile Animal Model for Fertility Research

“…The secretions of biological hormones such as CRF, ACTH, and cortisol under stress generally lead to impaired reproductive system through inhibition of the release of reproductive hormones and gametogenesis ( Table 1 ) [ 150 ]. In female zebrafish, the disruptive effects of ACTH and cortisol on gametogenesis and fertilization success have been illustrated through the identification of oocytes with DNA damage as well as reduced nucleic acid via disruption of protein synthesis [ 151 ]. Additionally, ACTH induces strong vacuolization in zebrafish ooplasm and similar condition was also observed in mammalian adrenal gland cells following exposure to ACTH [ 151 , 152 ].…”