Lipid peroxidation and antioxidant responses in zebrafish brain induced by Aphanizomenon flos-aquae DC-1 aphantoxins

Aphanizomenon flos-aquae is a cyanobacterium that is frequently encountered in eutrophic waters worldwide. It is source of neurotoxins known as aphantoxins or paralytic shellfish poisons (PSPs), which present a major threat to the environment and human health. The molecular mechanism of PSP action is known, however the in vivo effects of this neurotoxin on oxidative stress, lipid peroxidation and the antioxidant defense responses in zebrafish brain remain to be understood. Aphantoxins purified from a natural isolate of A. flos-aquae DC-1 were analyzed using high performance liquid chromatography. The major components of the toxins were gonyautoxins 1 and 5 (GTX1 and GTX5, 34.04% and 21.28%, respectively) and neosaxitoxin (neoSTX, 12.77%). Zebrafish (Danio rerio) were injected intraperitoneally with 7.73 μg/kg (low dose) and 11.13 μg/kg (high dose) of A. flos-aquae DC-1 aphantoxins. Oxidative stress, lipid peroxidation and antioxidant defense responses in the zebrafish brain were investigated at various timepoints at 1-24h post-exposure. Aphantoxin exposure was associated with significantly increased (>1-2 times) reactive oxygen species (ROS) and malondialdehyde (MDA) in zebrafish brain compared with the controls at 1-12h postexposure, suggestive of oxidative stress and lipid peroxidation. In contrast, reduced glutathione (GSH) levels in the zebrafish brain exposed to high or low doses of aphantoxins decreased by 44.88% and 41.33%, respectively, after 1-12h compared with the controls, suggesting that GSH participated in detoxification to ROS and MDA. Further analysis showed a significant increase in the activities of superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GPx) compared with the controls, suggesting elimination of oxidative stress by the antioxidant response in zebrafish brain. All these changes were dose and time dependent. These results suggested that aphantoxins or PSPs increased ROS and MDA and decreased GSH in zebrafish brain, and these changes induced oxidative stress. The increased activity of SOD, CAT and GPx demonstrated that these antioxidant enzymes could play important roles in eliminating excess ROS and MDA. These results also suggest that MDA, ROS, GSH and these three antioxidant enzymes in the brain of zebrafish may act as bioindicators for investigating A. flos-aquae DC-1 aphantoxins or PSPs and algal blooms in nature.