An in vitro ovarian explant culture system to examine sex change in a hermaphroditic fish

Alexander Goikoetxea; Erin L Damsteegt; Erica V Todd; Andrew McNaughton; Neil J Gemmell; P Mark Lokman

doi:10.7717/peerj.10323

An in vitro ovarian explant culture system to examine sex change in a hermaphroditic fish

PeerJ. 2020 Nov 11:8:e10323. doi: 10.7717/peerj.10323. eCollection 2020.

Authors

Alexander Goikoetxea¹, Erin L Damsteegt², Erica V Todd³, Andrew McNaughton¹, Neil J Gemmell¹, P Mark Lokman²

Affiliations

¹ Department of Anatomy, University of Otago, Dunedin, New Zealand.
² Department of Zoology, University of Otago, Dunedin, New Zealand.
³ School of Life and Environmental Science, Deakin University, Geelong, Australia.

Abstract

Many teleost fishes undergo natural sex change, and elucidating the physiological and molecular controls of this process offers unique opportunities not only to develop methods of controlling sex in aquaculture settings, but to better understand vertebrate sexual development more broadly. Induction of sex change in some sequentially hermaphroditic or gonochoristic fish can be achieved in vivo through social manipulation, inhibition of aromatase activity, or steroid treatment. However, the induction of sex change in vitro has been largely unexplored. In this study, we established an in vitro culture system for ovarian explants in serum-free medium for a model sequential hermaphrodite, the New Zealand spotty wrasse (Notolabrus celidotus). This culture technique enabled evaluating the effect of various treatments with 17β-estradiol (E₂), 11-ketotestosterone (11KT) or cortisol (CORT) on spotty wrasse ovarian architecture for 21 days. A quantitative approach to measuring the degree of ovarian atresia within histological images was also developed, using pixel-based machine learning software. Ovarian atresia likely due to culture was observed across all treatments including no-hormone controls, but was minimised with treatment of at least 10 ng/mL E₂. Neither 11KT nor CORT administration induced proliferation of spermatogonia (i.e., sex change) in the cultured ovaries indicating culture beyond 21 days may be needed to induce sex change in vitro. The in vitro gonadal culture and analysis systems established here enable future studies investigating the paracrine role of sex steroids, glucocorticoids and a variety of other factors during gonadal sex change in fish.

Keywords: Cortisol; Organ culture; Previtellogenic oocyte; Sex change; Spotty wrasse.

Grants and funding

Alexander Goikoetxea was supported by a scholarship from the Department of Anatomy, University of Otago. Erica V. Todd held a Health Sciences Career Postdoctoral Fellowship and Rutherford Postdoctoral Fellowship that supported this work. This work was further supported by a University of Otago Research Grant (ORG 0117-0318) to Neil J. Gemmell and Erica V. Todd, and a Marsden Grant (UOO1308) to Neil J. Gemmell and a Research Enhancement Grant (PBRF-ML63) to P. Mark Lokman. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.