Abstract
The survival of an animal relies vitally on its ability to respond appropriately towards a stressor. Activation of the hypothalamic pituitary interrenal axis (HPI), also known as the stress axis, is a key element of the stress response in teleost fish. Elevated levels of cortisol have been associated with interrupted growth and development, reproduction and immune system functions. Thus, the ability to manipulate cortisol levels can be a beneficial tool for breeding and maintaining healthy fish stocks in aquaculture, and for basic science aimed at understanding stress responses. Adrenal cortisol synthesis is increased in response to binding of adrenocorticotropic hormone (ACTH) to the melanocortin 2 receptor (Mc2r).
In the catadromous New Zealand short fin eel (Anguilla australis), non-migrants display higher plasma cortisol concentrations than migrants. Conversely, the key sex steroid hormone, 11-ketotestosterone (11-KT), was in higher levels in migrants that also display more advanced gonadal development. It was hypothesised that the difference in plasma cortisol between non-migrants and migrants would be reflected by increased activation of the HPI axis. This was measured as changes in gene expression of mc2r, as well as the genes encoding steroidogenic enzymes that are involved in the synthesis of cortisol (Chapter 2). In this thesis, differences in plasma cortisol and 11-KT levels measured by radioimmunoassay between non-migrants and migrants were confirmed. Real-time quantitative PCR showed that the gene expression of mc2r, star (steroidogenic acute regulatory protein), cyp11a1 (Cholesterol side-chain cleavage enzyme) and cyp11b (11β-hydroxylase) between non-migrants and migrants mirrored the trends of plasma 11-ketotestosterone. This suggests there is higher activity of the HPI axis in migrant eels despite having lower plasma cortisol levels than that of non-migrant eels.
In addition, the effects of blockage of Mc2r were examined in A. australis in vivo, using a novel ACTH antagonist (ACTH-X; Chapter 3) contained in cholesterol-cellulose implants. Increasing the concentration of ACTH-X reduced the stress response to a standardized stressor on Day 1 through the reduction of plasma cortisol, and the down-regulation in gene expression of cyp11a1 and cyp11b, but not in mc2r and star. In contrast, suppression of the HPI axis was not observed on Day 3. High pressure liquid chromatography analysis on the release-rates of ACTH-X implants in vitro revealed the absence of ACTH-X on Day 3 and subsequent days. Thus, this may explain the absence of any significant trends on Day 3 that were initially found on Day 1.
Lastly, the expression of mc2r and several other melanocortin receptors (MCRs) were examined in different tissues of A. australis to identify their distribution. In addition to mc2r, mc4r was also expressed in the interrenal tissues. This study demonstrates the potential value of ACTH-X as a new tool to suppress cortisol levels, so as to further understand the regulation of the stress response and develop means to limit the negative effects of stress on animal well-being and production. This study also opens further avenues for future research which could provide significant implications for aquaculture.