Understanding physiological responses and development of stress biomarker from tilapia treated with vitamin C during chronic stress induced by crowding

Experiments were conducted to investigate: i) the effects of vitamin C as a nutraceutical in modulating stress and ii) usability of expression of the genes adrenocorticotropic hormone (ACTH) receptor and corticotropin releasing hormone (CRH) as stress biomarkers. Tilapia (Oreochromis niloticus) fingerlings were obtained from a local fish farm and were acclimated in the new environment for four weeks. After the acclimation period, fish were maintained in two different conditions – controlled and stressed (by crowding; density \u3e 50 g/l). Within each condition, fish were further divided into two diet groups, each with two replicates, and were fed vitamin C supplemented (@1000 mg/kg feed) and vitamin C free feed, separately for fifteen days. Six fish from each group were sampled to assess the physiological parameters – plasma cortisol, blood glucose, plasma protein, packed cell volume, spleen somatic index, and condition factor. Fish in the stressed conditions showed significantly higher level (P\u3c0.05) of plasma cortisol and blood glucose indicating stress, both of which were reduced when fed vitamin C supplemented diet. Significantly higher (P\u3c0.05) hematocrit concentration was observed in the vitamin C fed group indicating better adaptation. To fulfill the second objective, reverse transcription-polymerase chain reactions (RT-PCR) were carried on mRNA collected from brain and head kidney tissues. In all samples, almost equal expression of β-actin, the housekeeping gene was observed. As for the two genes involved in stress response pathway, CRH expression was not detected in any of the experimental groups whereas ACTH receptors were clearly expressed in one of the stressed groups. Therefore, we suggest that instead of CRH, which may not be an ideal candidate, further research be performed to establish ACTH receptors as a genetic expression biomarker of stress