Cadmium and Benzo[a]pyrene are two toxicants of great environmental importance given their frequency and ability to cause extensive toxicity in aquatic organisms including fish. There is evidence that fish can modulate their respective uptake rate during simultaneous exposures, albeit the mechanism behind this is poorly understood. The present study aimed to examine this interaction by exposing adult zebrafish to either 89.3 nM Cd, 4.25 nM BaP or a combination of the two for 72 hrs prior to examining the uptake rate of either toxicant via short-term exposures (3-6 hrs) to radiotracers (109Cd and 14C-BaP). Our results showed that Cd uptake rate increased significantly in the gills when animals were pre-exposed to both toxicants simultaneously, resulting in an increased maximum uptake rate (Jmax). The increased Cd uptake rate did not correspond to increased expression of gill Cd transporters such as the epithelium calcium channel (ECaC) or the divalent metal transporter 1 (DMT1). Furthermore, BaP uptake rate increased significantly at the whole-body level when animals were exposed to both 5.03 nM 14C-BaP and 89.3 nM Cd concurrently. Additionally, we ran a time-course and observed BaP uptake rate is highest in the 6-12 hrs following the beginning of the exposure. Our results provide evidence that the increased bioaccumulation of Cd and BaP observed during co-exposures is at least in part due to an increase in uptake rate and is driven by separate mechanisms.

An in vitro study was carried out to examine the impact of UV exposure on metal-dissolved humic material (M-DHM) complexes in aqueous systems at different pH. Complexation reactions of dissolved M (Cu, Ni, and Cd) with DHM increased with the increasing pH of the solution. Kinetically inert M-DHM complexes dominated at higher pH in the test solutions. Exposure to UV radiation did affect the chemical speciation of M-DHM complexes at different pH of the systems. The overall observation suggests that exposure to increasing UV radiation increased the lability, mobility, and bioavailability of M-DHM complexes in aquatic environments. The dissociation rate constant of Cu-DHM was found to be slower than Ni-DHM and Cd-DHM complexes (both before and after UV exposure). At a higher pH range, Cd-DHM complexes dissociated after exposure to UV radiation and a part of this dissociated Cd precipitated out from the system. No change in the lability of the produced Cu-DHM and Ni-DHM complexes after UV radiation exposure was observed. They did not appear to form new kinetically inert complexes even after 12 h of exposure. The outcome of this research has important global implications. The results of this study helped to understand DHM leachability from soil and its effect on dissolved metal concentrations in the Northern Hemisphere water bodies. The results of this study also facilitated to comprehend the fate of M-DHM complexes at photic depths (where pH changes are accompanied by high UV radiation exposure) in tropical marine/freshwater systems during summer.

Complex mixtures of unknown contaminants present a challenge to identify toxicological risks without using large numbers of animals and labor-intensive screens of all organs. This study examined soil extracts from a legacy-contaminated pesticide packaging and blending site. HepG2 cytotoxicity was used as an initial screen of 18 soil samples; then, three extracts (A, B and C) from different locations at the study site were used for testing in animals. The first two extracts were identified as the most toxic in vitro, and the latter extract obtained from a location further from these two toxic sampling sites. Then, target organ toxicities were identified following biweekly oral gavage for one month of three soil extracts (0.1% in polyethylene glycol or PEG) compared to vehicle control in male Sprague–Dawley rats (n = 9–10/group). Exposure to extract A significantly increased neutrophils and lymphocytes compared to control. In contrast, all extracts increased plasma α−2 macroglobulin and caused mild-to-moderate lymphocytic proliferation within the spleen white pulp, all indicative of inflammation. Rats exposed to all soil extracts exhibited acute tubular necrosis. Cholinesterase activity was significantly reduced in plasma, but not brain, after exposure to extract A compared to control. Increased hepatic ethoxyresorufin-o-deethylase activity compared to control was observed following exposure to extracts A and B. Exposure to soil extract C in rats showed a prolonged QTc interval in electrocardiography as well as increased brain lipid peroxidation. Candidate contaminants are organochlorine, organophosphate/carbamate pesticides or metabolites. Overall, HepG2 cytotoxicity did not successfully predict the neurotoxicity and cardiotoxicity observed with extract C but was more successful with suspected hydrocarbon toxicities in extracts A and B. Caution should be taken when extrapolating the observation of no effects from in vitro cell culture to in vivo toxicity, and better cell culture lines or assays should be explored.

Individual prey often exhibit consistent behavioural differences in responses to risk. Here, we assess whether such behavioural consistency is linked to morphological changes that are known to result in differences in vulnerability to predators. Some fishes increase their depth-to-length ratio when under increased risk of predation, thereby reducing their risk to gape-limited predators. However, the development of these defences is limited by available resources. We asked whether behavioural tendencies associated with shelter seeking and activity are linked to differences in growth patterns. Common carp classified as 'active/non-sheltering' or 'passive/sheltering' based on their movement patterns and shelter use, showed consistency in behaviour over a 10-month period with active/non-sheltering fish developing a greater depth to length ratio than passive/sheltering fish. The effectiveness of anti-predator strategies in animals is an integrative function of both behaviour and morphology. Our works highlights a fascinating link between behavioural consistency and the development of adaptive morphologies.

Anthropogenic activities over the past several decades have depleted stratospheric ozone, resulting in a global increase in ultraviolet radiation (UVR). Much of the negative effects of UVR in aquatic systems is minimized by dissolved organic carbon (DOC) which is known to attenuate UVR across the water column. The skin of many fishes contains large epidermal club cells (ECCs) that are known to play a role in innate immune responses and also release chemical alarm cues that warn other fishes of danger. This study investigated the effects of in vivo UVR exposure to fathead minnows (Pimephales promelas), under the influence of two sources of DOC: Sigma Aldrich humic acid, a coal based commercial source of DOC and Luther Marsh natural organic matter, a terrigenous source of DOC. Specifically, we examined ECC investment and physiological stress responses and found that fish exposed to high UVR, in the presence of either source of DOC, had higher ECC investment than fish exposed to high UVR only. Similarly, exposure to high UVR under either source of DOC, reduced cortisol levels relative to that in the high UVR only treatment. This indicates that DOC protects fish from physiological stress associated with UVR exposure and helps maintain production of ECC under conditions of UVR exposure.

The combined use of a competing ligand exchange (CLE) method and a diffusive gradient in thin films (DGT) technique in a quasi-labile system provides a better understanding of dynamic metal (Cu and Ni) complexes in the presence of humic substances of different origins. The CLE and DGT techniques provide total labile (dynamic) metal complexes (Cu and Ni) and their dissociation rate constants in environmental systems. DGT was found to estimate lower concentrations of labile metal complexes than CLE. These discrepancies were caused by diffusion controlled metal flux (towards the binding resin gel) in the diffusive gel of DGT. The interactions of Cu and Ni with humic acids are stronger than their interactions with fulvic acid and natural organic matter. Changes in the lability of Ni and Cu complexes (complexed with humic substances of different origins) with the changing analytical detection window indicate that the complexes of these metals were formed with different binding sites with diverse binding energies in the humic substances. The combination of these two techniques was found to be very useful in determining diffusion coefficients of labile metal-humate complexes in quasi-labile systems. The values of diffusion coefficients of labile Ni and Cu complexes determined in this study are in good agreement with limited results from the literature. This finding is novel and can be very useful in further improving our understanding of the metal-humate interactions in natural environments.

Anthropogenic activities have dramatically increased over the past decades, with the consequence that many organisms are simultaneously exposed to multiple stressors. Understanding how organisms respond to these stressors is a key focus for scientists from many disciplines. Here we investigated the interactive effects of two stressors, UV radiation (UVR) and cadmium (Cd) exposure on a common freshwater fish, fathead minnow (Pimephales promelas). UVR is known to influence the density of epidermal club cells (ECCs), which are not only a key component of the innate immune system of fishes, but are also the source of chemical alarm cues that serve to warn other fishes of nearby predators. In contrast, Cd impairs the physiological stress response and ability of fish to respond to alarm cues. We used an integrative approach to examine physiological stress response as well as investment in ECCs. Fish exposed to UVR had higher levels of cortisol than non-exposed controls, but Cd reduced cortisol levels substantially for fish exposed to UVR. Fish exposed to UVR, either in the presence or absence of Cd, showed consistent decreases in ECC investment compared to non-exposed controls. Despite differences in ECC number, there was no difference in the potency of alarm cues prepared from the skin of UVR and Cd exposed or non-exposed fish indicating that UVR and Cd exposure combined may have little influence on chemically-mediated predator-prey interactions.

Many prey fishes possess large club cells in their epidermis. The role of these cells has garnered considerable attention from evolutionary ecologists. These cells likely form part of the innate immune system of fishes, however, they also have an alarm function, releasing chemical cues that serve to warn nearby conspecifics of danger. Experiments aimed at understanding the selection pressures leading to the evolution of these cells have been hampered by a surprisingly large intraspecific variation in epidermal club cell (ECC) investment. The goal of our current work was to explore the magnitude and nature of this variation in ECC investment. In a field survey, we documented large differences in ECC investment both within and between several populations of minnows. We then tested whether we could experimentally reduce variation in mean ECC number by raising fish under standard laboratory conditions for 4 weeks. Fish from different populations responded very differently to being held under standard laboratory conditions; some populations showed an increase in ECC investment while others remained unchanged. More importantly, we found some evidence that we could reduce within population variation in ECC investment through time, but could not reduce among-population variation in mean ECC investment. Given the large variation we observed in wild fish and our limited ability to converge mean cell number by holding the fish under standard conditions, we caution that future studies may be hard pressed to find subtle effects of various experimental manipulations; this will make elucidating the selection pressures leading to the evolution of the cells challenging.

Recent anthropogenic activities have caused deleterious effects to the stratospheric ozone layer, resulting in a global increase in the level of ultraviolet radiation (UVR). Understanding the way that organisms respond to such stressors is key to predicting the effects of anthropogenic activities on aquatic ecosystems and the species that inhabit them. The epidermal layer of the skin of fishes is not keratinized and acts as the primary interface between the fish and its environment. The skin of many species of fishes contains large epidermal club cells (ECCs) that are known to release chemicals (alarm cues) serving to warn other fishes of danger. However, the alarm role of the cells is likely secondary to their role in the immune system. Recent research suggests that ECCs in the epidermis may play a role in protecting the fish from damage caused by UVR. In the present study, we examined the effects of in vivo exposure to UVR on fathead minnows (Pimephales promelas), specifically investigating ECC investment, physiological stress responses, and alarm cue production. We found that fish exposed to UVR showed an increase in cortisol levels and a substantive decrease in ECC investment compared to non-exposed controls. Unexpectedly, our subsequent analysis of the behavioural response of fish to alarm cues revealed no difference in the potency of the cues prepared from the skin of UV-exposed or non-exposed minnows. Our results indicate that, although nonlethal, UVR exposure may lead to secondary mortality by altering the fish immune system, although this same exposure may have little influence on chemically-mediated predatorprey interactions. (c) 2012 The Linnean Society of London, Biological Journal of the Linnean Society, 2012, 105, 832841.

For prey species that rely on learning to recognize their predators, natural selection should favour individuals able to learn as early as possible. The earliest point at which individuals can gather information about the identity of their potential predators is during the embryonic stage. Indeed, recent experiments have demonstrated that amphibians can learn to recognize predators prior to hatching. Here, we conditioned woodfrog embryos to recognize predatory salamander cues either in the morning or in the evening, and subsequently exposed the two-week-old tadpoles to salamander cues either in the morning or in the evening, and recorded the intensity of their antipredator behaviour. The data indicate that amphibians learn to recognize potential predators while still in the egg, and also learn the temporal component of this information, which they use later in life, to adjust the intensity of their antipredator responses throughout the day.