Growth inhibition and coordinated physiological regulation of zebrafish (Danio rerio) embryos upon sublethal exposure to antidepressant amitriptyline

Amitriptyline is a tricyclic antidepressant used for decades. It is present at low detectable concentrations in the aquatic environment, but relative few studies have focused on its ecotoxicological effects on non-target aquatic animals. The present study conducted an acute toxicity test of waterborne amitriptyline exposure using zebrafish (Danio rerio) embryos 4 to 124h-post-fertilization. Time-dependent lethal concentrations were firstly determined and at mg/L levels. Effects of amitriptyline on zebrafish embryos were then evaluated under amitriptyline exposure at sublethal concentrations of 1, 10, 100ng/L, 1, 10, 100μg/L and 1mg/L. Our results showed that amitriptyline significantly reduced the hatching time and body length of embryos after exposure in a concentration-dependent manner. Our study also revealed that the exposure evoked a coordinated modulation of physiological and biochemical parameters in exposed zebrafish embryos, including alterations of adrenocorticotropic hormone (ACTH) level, oxidative stress and antioxidant parameters, as well as nitric oxide (NO) production and total nitric oxide synthase (TNOS) activity. A U-shaped concentration-dependent response curve was observed in ACTH level in response to amitriptyline exposure. However, both U-shaped and inversed U-shaped curves were indicated in the responses of antioxidant parameters, including total antioxidant capacity, antioxidant enzyme activities (catalase, superoxide dismutase and peroxidase), glutathione content and glutathione reductase activity. Correspondingly, hydroxyl radical formation and lipid peroxidation indices changed in similar U-shaped concentration-dependent patterns, which together the results of antioxidant parameters suggested induction of oxidative stress in embryos exposed to amitriptyline at high concentrations. Moreover, NO production and TNOS activity were both significantly affected by amitriptyline exposure. Notably, significant correlations between these measured parameters were revealed, which suggested a dynamic adaptation process and coordinated regulation of multiple physiological systems in fish embryos to amitriptyline treatment. Furthermore, our study demonstrated that the effective concentrations of amitriptyline for measured parameters in zebrafish embryos were as low as 10ng/L, and thus revealed the potential risk of amitriptyline and other antidepressants to aquatic life.