Acute toxicity of manganese in goldfish Carassius auratus is associated with oxidative stress and organ specific antioxidant responses

Abstract

Manganese is a relatively common, yet poorly studied element in freshwater ecosystems, where it can be significantly bioconcentrated. The knowledge about the mechanisms of Mn toxicity on fish health is still limited. The aim of the present study was to assess the potential induction of oxidative stress and the antioxidant response after a 96 h waterborne Mn-exposure (at 0.1 and 1 mM) in gill, kidney, liver and brain of goldfish (Carassius auratus). Mn 1 mM induced an increase of lipid hydroperoxides, superoxide dismutase (SOD) and glutathione peroxidase (GPx) activities in all tissues with the exception of SOD inhibition in the brain. Particular response of catalase (CAT) was indicated—its inhibition in the liver and kidney, but activation in the gill. Exposure to Mn 0.1 mM provoked most prominent changes in the liver and did not change the indexes in brain. These results strongly suggest that Mn exposure caused a generalized oxidative stress in the fish and revealed an organ specific antioxidant response involving a differential modulation of the SOD, CAT and GPx activities.

Introduction

At concentrations over the natural levels, heavy metals are a serious threat to the aquatic ecosystems due to their high toxicity, non-biodegradable and bioaccumulative properties (Velma and Tchounwou, 2010). Besides, metal pollution may not only lead to a disturbed integrity of ecosystems but also to a challenged human health through the food chain. Thus, monitoring and prevention of heavy metal contamination has attracted much attention. The use of fish species is highly recommended for this purpose. From the surrounding water, fish may absorb dissolved heavy metals that may accumulate in various tissues and organs and even be biomagnified (Abdullah et al., 2007), which makes it much of a significance for the biomonitoring. In addition, studies based on the responses to aquatic metal may help in the better understanding of the potential toxicological mechanism (Zhou et al., 2007).

Manganese is a relatively common, yet poorly studied element in freshwater ecosystems (Dittman and Buchwalter, 2010). Manganese is a naturally occurring element, comprising about 0.1% of the Earth's crust, which is found ubiquitously in the environment (Howe et al., 2004). Dissolved manganese in natural waters of anthropogenic sources/influences associated with metal mining and other industrial activities may reach very high concentrations (McNeely et al., 1979, Morillo and Usero, 2008). Manganese can be significantly bioconcentrated by aquatic biota at lower trophic levels (i.e., bioconcentration factors of 35–930 for fish have been estimated), but there are conflicting reports on whether biomagnification of manganese occurs (Howe et al., 2004).

Manganese is an essential micronutrient for fish but can be very toxic at concentrations above the optimal threshold level. Acute toxicity studies reported 96-h LC₅₀s for fish ranging from 2.4 mg/L for coho salmon (Oncorhynchus kisutch) to 3350 mg/L for Indian catfish (Heteropneustes fossilis) (Howe et al., 2004). The range of manganese concentration causing toxicity depends on fish species, life stage and ambient water chemistry (Fish, 2009). Manganese toxicity decreases with increasing water hardness (Stubblefield et al., 1997, Howe et al., 2004). Acid deposition causes elevated concentrations of metals like manganese in surface waters. Brown trout mortality in acid streams was explained by the pH and the concentration of labile inorganic manganese in the water, which correlated to the accumulation of manganese in the gill (Nyberg et al., 1995). In a study on the distribution of the ⁵⁴Mn radionuclide in the rainbow trout, Adam et al. (1997) classified the organs in two groups with different elimination kinetics. The first group consists of organs of penetration or transit, such as the skin, gills, kidneys, liver, primary and secondary gut and viscera, whereas the second group is made up of the receptor and storage organs and tissues such as the bone, head, fins and muscle. Participation of manganese in enzymatic activities of liver, pancreas and gastrointestinal mucosa may explain its high uptake in these tissues, whereas accumulation in the skeleton may be related to bone formation (Rouleau et al., 1995). A primary toxic action of manganese on brook charr, Sulvelinus fonfinalis, was the disruption of sodium regulation; plasma sodium decrease was inversely correlated with surface binding of manganese on the gill (Gonzalez et al., 1990). In the same way, Partridge and Lymbery (2009) showed in juvenile mulloway (Argyrosomus japonicus) that manganese accumulated in the gills, liver and muscle and produced alteration of carbohydrate metabolism and blood plasma ions such as potassium, sodium and chloride. Interestingly, Rouleau et al. (1995) demonstrated that the uptake of manganese by way of olfactory system favored its accumulation in the brain of brown trout and suggested that this route could be of significance for fish neurotoxicity. On the other hand, chronic studies reported a decrease in red blood cells and hematocrit due to internal hemorrhaging in tilapia (Wepener et al., 1992). Nonetheless, the knowledge about the impact of manganese on fish health is still limited.

A wide range of environmental pollutants, as heavy metals, are known to induce oxidative stress in aquatic animals including fish. The generation of reactive oxygen species (ROS) induced by heavy metals is commonly associated with cellular injures due to alterations in DNA, proteins and membranes (Leonard et al., 2004). To counteract the adverse effect of ROS, living organisms have a complex and effective antioxidant defense system comprising both enzymatic and non-enzymatic mechanisms such as superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx), ascorbate, reduced glutathione (GSH) and Vitamin E (Kelly et al., 1998). Variations in these antioxidant defenses can be very sensitive in revealing a pro-oxidant condition and have been used as biomarkers of oxidative stress in fish (Ahmad et al., 2006a, Ahmad et al., 2006b, Oliveira et al., 2008).

Despite the limited information on the biochemical mechanisms of manganese toxicity, there are evidences pointing to an association with the generation of ROS in mammals (Aschner et al., 2007). Manganese is involved in neurogenetic diseases through free radical formation (Chen et al., 2006) and the inactivation of antioxidant enzymes (Liccione and Maines, 1988, Desole et al., 1995). Furthermore, relatively short and ecologically relevant manganese exposures resulted in marked decreases in thiol concentrations scavenger in aquatic insects, which is especially alarming considering that manganese often co-occurs with metals and contaminants (Dittman and Buchwalter, 2010). However, the effect of manganese on the induction of oxidative stress and the antioxidant defenses has not been investigated thoroughly in fish. Depending on the concentration, manganese has been reported to have both protective and toxic effect by functioning either as a scavenger of superoxide radical or by inducing a ROS-dependent impairment of the antioxidant system (Nayak et al., 1999, Arabi and Alaeddini, 2005).

The present study was designed to investigate whether manganese exposure causes toxicity in fish by inducing oxidative stress. Taking into account a putative organ-specific toxicity, the effect of a short-term manganese exposure on the induction of oxidative damage and the antioxidant enzymes SOD, CAT and GPx was assessed in liver, kidney, gill and brain.