Impact of antiparasitic used in livestock: effects of ivermectin spiked sediment in Prochilodus lineatus, an inland fishery species of South America.

Ivermectin (IVM) is a widely used antiparasitic. Concerns have been raised about its environmental effects in the wetlands of Río de la Plata basin where cattle have been treated with IVM for years. This study investigated the sublethal effects of environmentally relevant IVM concentrations in sediments on the Neotropical fish Prochilodus lineatus. Juvenile P. lineatus were exposed to IVM-spiked sediments (2 and 20 µg/Kg) for 14 days, alongside a control sediment treatment without IVM. Biochemical and oxidative stress responses were assessed in brain, gills, and liver tissues, including lipid damage, glutathione levels, enzyme activities, and antioxidant competence. Muscle and brain acetylcholinesterase activity (AChE) and stable isotopes of 13C and 15N in muscle were also measured. The lowest IVM treatment resulted in an increase in brain lipid peroxidation, as measured by thiobarbituric acid reactive substances (TBARs), decreased levels of reduced glutathione (GSH) in gills and liver, increased catalase activity (CAT) in the liver, and decreased antioxidant capacity against peroxyl radicals (ACAP) in gills and liver. The highest IVM treatment significantly reduced GSH in the liver. Muscle (AChE) was decreased in both treatments. Multivariate analysis showed significant overall effects in the liver tissue, followed by gills and brain. These findings demonstrate the sublethal effects of IVM in P. lineatus, emphasizing the importance of considering sediment contamination and trophic habits in realistic exposure scenarios.

Persistent organic pollutants and chlorpyrifos in tissues of a histotrophic viviparous species, the Southern Eagle Ray Myliobatis goodei.

Elasmobranchs are good indicators of marine pollution as they accumulate pollutants from water and food, and occupy different trophic levels. Concentrations of organochlorine pesticides (OCPs), polychlorinated biphenyls (PCBs) and chlorpyrifos were quantified in muscle, liver, gonads, gills, and brain in both sexes and maturity stages of the Southern Eagle Ray, Myliobatis goodei, captured in Argentine coastal waters. Moreover, possible histological alterations in the liver and gonads were analyzed. Pollutant concentrations were pervasive across all tissues, with PCBs > OCPs > chlorpyrifos. Elevated pollutant levels were notably found in the liver and gills. We identified thirty-six PCB congeners in tissues, with low-chlorine congeners prevailing. Among OCPs, ∑DDT and ∑endosulfan were predominant. Females exhibited higher pollutant levels in most tissues compared to males, except in the gonads, and adults generally displayed elevated pollutant levels. Histological analysis revealed the presence of atretic follicles and melanomacrophages (MM). Continuous monitoring of pollutant levels, alongside their effects on physiological and ecological traits, is imperative for effective management and conservation efforts.

A biomarker approach to study the effects of polluted Brazilian urban reservoirs in a native fish.

Two of the largest water reservoirs in the Metropolitan Region of São Paulo, Brazil (MRSP), named Billings and Guarapiranga, are facing high levels of anthropic impact. This is evidenced by the presence of contaminants and pollutants, which are deteriorating their water quality. Therefore, this study evaluated antioxidant defense enzymes, lipoperoxidation and genotoxicity, in adult females of a native species, Astyanax altiparanae from the Guarapiranga and Billings reservoirs. The study also aimed to evaluate these biomarkers during two different periods of the year, the rainy (summer) and dry (winter) seasons. The oxidative stress was evaluated by the activity of enzymes such as glutathione peroxidase, glutathione S-transferases, superoxide dismutase, and catalase in the gills and liver, and the occurrence of lipoperoxidation was also evaluated in both organs. The genotoxicity was assessed by performing comet assay, micronucleus, and nuclear abnormality tests on blood samples. The results showed that fish from both reservoirs are subjected to oxidative stress and genotoxic damage, mainly during winter, but fish living in Billings showed greater alterations than fish from Guarapiranga. Likewise, the results of the principal component analysis suggested that caffeine, nitrogenous compounds, and some metals might be triggering these toxic effects in fish.

Physiological responses on the reproductive, metabolism and stress endpoints of Astyanax lacustris females (Teleostei: Characiformes) after diclofenac and ibuprofen exposure.

Diclofenac (DCF) and ibuprofen (IBU) are pharmaceutical compounds frequently detected in aquatic compartments worldwide. Several hazard effects including developmental abnormalities and redox balance impairment have been elucidated in aquatic species, but multiple endocrine evaluations are scarce. Therefore, the present study aimed to assess the disruptive physiological effects and toxicity of DCF and IBU isolated and combined, using females of the native freshwater teleost Astyanax lacustris. In regards to NSAIDs bioavailability, the results showed absence of degradation of IBU and DCF after 7 days of exposure. IBU LC50 for A. lacustris was 137 mgL-1 and females exposed to IBU isolated increased thyroxine (T4) concentration at 24 h and decreased after 96 h; DCF exposure decreased triiodothyronine (T3) concentration at 96 h. Circulating levels of 17ß-estradiol (E2), cortisol (F) and testosterone (T) were not affected by any treatment. HPG and HPI axis genes fshß, pomc and vtg were upregulated after 24 h of IBU exposure, and dio2 was downregulated in DCF fish exposed group after 96 h compared to the mixture. Protein concentration was reduced in muscle and increased in the liver by DCF and mixtures exposures at 24 h; while liver lipids were increased in the mixture groups after 96 h. The study point out the capacity of NSAIDs to affect endocrine endpoints in A. lacustris females and induce changes in energetic substrate content after acute exposure to isolated and mixed NSAIDs treatments. Lastly, the present investigation brings new insights into the toxicity and endocrine disruptive activity of NSAIDs in Latin America teleost species and the aquatic environment.

Integrative study of the reproductive biology and growth of Acestrorhynchus pantaneiro Menezes, 1992 (Characiformes, Acestrorhynchidae).

We studied the reproductive strategy, sexual system and growth of dientudo paraguayo Acestrorhynchus pantaneiro. After 2 years of monitoring in shallow areas of a floodplain lake from the lower Paraná basin (Argentina), it was evidenced that water temperature modulated gonadal maturation, but it was the river water level the synchronising stimulus that triggered spawning. This species exhibited a single annual breeding period from October to January, with most spawning activity in November. According to the von Bertalanffy growth curve, fish would reach autumn to winter months with LS of ~120 mm, already mature males. The first mature females were found at LS of 210 mm, becoming sexually mature between the second and third breeding seasons. This is the first integrative study that includes the body-length frequency distribution and sex differential size at first maturity and growth, and reports the presence of intersex gonads questioning its sexual pattern from gonochoristic to sequential hermaphrodite species. The sexual pattern, the multiple spawning behaviour and a medium to high absolute fecundity support the opportunistic and invasive behavior observed in previous contributions for this characiform species.

Latin America: Knowledge diversity for a sustainable future.

National and international collaboration and the incorporation of perspectives from outside academia are essential parts of a paradigm shift in science and are necessary for a sustainable future. The articles in this special series "Diversity of Knowledge for a Sustainable Future in Latin America" exemplify many of these aspects, showcasing the latest research and environmental policies for the protection of water, soil, and human health related to several pollutants, for example, microplastics, hydrocarbons, metals, and pesticides. Knowledge-built in an environment where communities are participants in this construction and not only subjects-helps promote understanding of the transdisciplinary nature of environmental issues provided by a diversity of traditional and nontraditional participants. Integr Environ Assess Manag 2023;19:613-614. © 2023 SETAC.

Neurotoxic, biotransformation, oxidative stress and genotoxic effects in Astyanax altiparanae (Teleostei, Characiformes) males exposed to environmentally relevant concentrations of diclofenac and/or caffeine.

The present study evaluated neurotoxic, biotransformation, genotoxic and antioxidant responses to relevant environmental concentrations of diclofenac (0.4 µg L-1) and caffeine (27.5 µg L-1), separate and combined, in adult males of the freshwater fish Astyanax altiparanae after a subchronic exposure (14 days). Fish exposed to diclofenac and caffeine, both separate and combined, revealed a neurotoxic effect through the inhibition of acetylcholinesterase activity in the muscle, while diclofenac alone and in combination caused cyclooxygenase inhibition. Caffeine alone produces genotoxicity on this species but, when combined with diclofenac, it potentiates hepatic lipoperoxidation and the inhibition of oxidative stress enzymes, while diclofenac alone or in combination produces a general inhibition of important enzymes. This study suggests that aquatic contamination produced by these pharmaceuticals has the potential to affect homeostasis and locomotion in A. altiparanae and compromise their immune system and general health.

Endocrine disruption in the sub Antarctic fish Patagonotothen tessellata (Perciformes, Notothenidae) from Beagle Channel associated to anthropogenic impact.

Situated in the sub-Antarctic region, Beagle Channel represents a unique marine ecosystem due to the connection between the Pacific and the Atlantic Oceans, and its proximity to the Antarctic Peninsula. Ushuaia city, the biggest settlement on the channel, exerts an increasing anthropogenic pressure by discharges of urban and industrial effluents. In the present work, we use Patagonotothen tessellata, one of the most abundant and widespread species in the channel, as a bioindicator species in order to evidence anthropic impact from Ushuaia Bay and surrounding areas. We first analyzed and characterized real time gene expression of androgen receptor, estrogen receptor and different forms of vitellogenin (VTG), under laboratory conditions. This was achieved by induction with estradiol of P. tessellata males. Then, the selected genes were used as biomarkers for an environmental biomonitoring study. Morphometric indices and circulating sex steroids (estradiol and testosterone) were also quantified in male fish collected from different sites. The qPCR analysis showed that vtgAb form is more inducible than vtgAa or vtgC forms after estrogen induction. The field survey revealed the up-regulation of vtgAb and the androgen receptor in fish from sites with higher anthropogenic influence. Sex steroids followed seasonal variations according to their reproductive cycle, with higher levels of estradiol and testosterone in winter and summer seasons. The use of biomarkers such as gene expression of VTG demonstrates that fish from Ushuaia Bay are likely to be exposed to endocrine disrupting compounds. To our knowledge, this research is the first attempt to assess the endocrine disruption associated to anthropic impact in a widespread fish of the Beagle Channel and contributes to a better understanding of the reproductive physiology of sub Antarctic ichthyofauna.

Ivermectin: A multilevel approach to evaluate effects in Prochilodus lineatus (Valenciennes, 1836) (Characiformes, Prochilodontidae), an inland fishery species.

Ivermectin (IVM) is one of the most widely used antiparasitics worldwide. It is a potent and effective drug for treatment and prevention of internal and external parasitic infections of livestock and humans. IVM is excreted unchanged in manure of treated animals. Thus, residues of IVM may reach aquatic systems, affecting non-target organisms such as fish. Although the presence of IVM in aquatic environments has been reported, a multilevel approach (from cellular to behavioral responses) is necessary to determine the health of exposed organisms and the environmental risks associated. The aim of the present study was to investigate the response of the Neotropical fish Prochilodus lineatus, one of the main target species of South American freshwater fisheries, exposed to environmental concentrations of IVM: low (0.5 µg L-1) and high (1.5 µg L-1). Behavioral responses were assessed in juvenile fish and included water column use, routine swimming, total distance travelled, total activity time and Maximum swimming speed achieved during the escape response. Biochemical/oxidative stress responses assessed included brain acetylcholinesterase (AChE), catalase (CAT) and glutathione S-transferase (GST) activities; total antioxidant competence against peroxyl radicals (ACAP) and lipid oxidative damage (TBARs). Hematological biomarker responses included blood glucose levels, hematocrit, hemoglobin concentration, mean corpuscular hemoglobin, mean corpuscular hemoglobin concentration, and mean corpuscular volume. Condition factor and hepatosomatic index were also calculated. The lowest IVM concentration caused a significant decrease in GST activity and maximum swimming speed during the escape response. Multivariate analysis with biochemical/stress and behavioral data revealed overall effects of IVM treatments. This multilevel analysis shows detrimental effects related to swimming behavior and predator avoidance which could affect population size and size-structure of P. lineatus. To our knowledge this is the first attempt to assess the effects of IVM on Neotropical fishes using an integrative approach based on biomarkers from different levels of biological organization.

Effects of Di-Isononyl Phthalate (DiNP) on Follicular Atresia in Zebrafish Ovary.

Di-isononyl phthalate (DiNP) is a plasticizer reported to elicit hormone-like activity and disrupt metabolism and reproduction in fish and other vertebrates. In general, phthalates have been used at high concentrations beyond reported environmental levels to assess their adverse effects on fish gonadal physiology. The present study exposed adult female zebrafish to a wide range of DiNP concentrations [0.42 µg L-1 (10-9 M), 4.2 µg L-1 (10-8 M), and 42 µg L-1 (10-7 M)] for 21 days. We evaluated gene expression profiles related to apoptosis, autophagy, and oxidative stress; DNA fragmentation (TUNEL assay: terminal deoxynucleotidyl transferase dUTP nick end labeling) and caspase activity (CAS3) were also examined. Exposure to 0.42 and 4.2 µg L-1 upregulated the genes coding for tnfa and baxa, sod1, prkaa1, respectively. CAS3 immunohistochemistry revealed a higher number of positive vitellogenic oocytes in ovaries exposed to 0.42 µg L-1. Subsequently, we examined the relationship between CAS3 signaling and DNA fragmentation. Accordingly, DNA fragmentation was observed in vitellogenic follicles of fish exposed to 0.42 and 4.2 µg L-1. Our results demonstrate that follicular atresia can occur after exposure to environmental levels of DiNP for 21 days, which may adversely affect the reproductive performance of female zebrafish in a non-monotonic manner.

Diclofenac and caffeine inhibit hepatic antioxidant enzymes in the freshwater fish Astyanax altiparanae (Teleostei: Characiformes).

Although concentrations of pharmaceutical compounds in aquatic ecosystems are low, they can cause toxic effects on organisms. The aim of this study was to evaluate the effects of diclofenac (DCF), a non-steroidal anti-inflammatory drug, and caffeine (CAF), a central nervous system stimulant, both alone or combined, in Astyanax altiparanae males under acute exposure (96 h), measuring neurotoxicity biomarkers, antioxidant response and damage at biochemical and cellular levels. DCF concentration in water, separated and combined, was 3.08 mg L-1 and that of CAF was 9.59 mg L-1. To assess neurotoxicity, brain and muscle acetylcholinesterase (AChE) activities were measured. To evaluate oxidative stress, the enzymatic activities of superoxide dismutase (SOD), glutathione peroxidase (GPx), catalase (CAT) and glutathione S-transferase (GST), as well as lipoperoxidation (LPO), were analyzed in liver and gills. Activity of hepatic cyclooxygenase (COX) was also evaluated. Genotoxicity was assessed in blood using comet assay and micronucleus test, as well as nuclear abnormalities. DCF and CAF, alone or combined, had neither effect on AChE activity, nor in the activity of SOD, CAT, GPx and GST in gills. In liver, DCF inhibited SOD and GPx activity, CAF inhibited CAT activity, the mixture inhibited SOD and GST activity; although only fish exposed to CAF showed increased hepatic LPO. Under these experimental conditions, no effect on COX activity was observed, nor cytotoxic and genotoxic damage. The most pronounced effects were caused by the drugs separately, since both compounds altered the enzymes, but only CAF triggered LPO, showing more harmful effects.

Endocrine disruptive action of diclofenac and caffeine on Astyanax altiparanae males (Teleostei: Characiformes: Characidae).

Diclofenac (DCF) and caffeine (CAF) are persistent pharmaceuticals that occur in mixtures in the aquatic ecosystems causing effects in the reproductive physiology of aquatic organisms. This study evaluated the physiological reproductive responses of Astyanax altiparanae males exposed to nominal concentrations of DCF (3.08 mg L-1) and CAF (9.59 mg L-1) separately and combined, for 96 h. The steroids profile, estrogenic biomarker vitellogenin (vtgA), testes and liver morphology, and also mortality of males were assessed. DCF and CAF degradation was 5% of the initial concentration for 24 h. The LC50 of the DCF and CAF were 30.8 mg L-1 and 95.9 mg L-1, respectively. Males exposed to DCF and CAF exhibited a reduction of 17ß-Estradiol (E2) concentration compared to control (CTL). Similarly, testosterone (T) was also reduced in the DCF treatment, but this response was not observed in 11-Ketotestosterone (11-KT). Males exposed to DCF + CAF combined did not exhibit differences in T, E2 and 11-KT steroids. The vtgA gene expression and the sperm concentration did not change among the treatments. Moreover, acute exposure revealed a hypertrophy of hepatocytes cells in the DCF and DCF + CAF treatments. In conclusion, DCF and CAF, isolated, exhibit an endocrine disruptive activity in A. altiparanae male, an opposite response observed with the mixture of both compounds that abolishes the endocrine disruptive effects. DCF seems to be more toxic for this species, altering also hepatocytes morphology.

Immunohistological Biomarkers of Toxicity by a Pharmaceutical Antidepressant in the Freshwater Cichlid Fish Cichlasoma dimerus (Teleostei, Cichliformes).

Melano-macrophage centers (MMCs) are nodular clusters of pigmented macrophages, implicated in homeostasis and destruction and recycling of endogenous and exogenous material. They can increase in size and/or frequency under environmental stress resulting in immunohistological biomarkers of water quality. Fluoxetine (FLX), a commonly prescribed antidepressant, can cause neuroendocrine, behavioral and reproductive alterations in teleost fish. In the present study, we analyzed the effects of a 2-week 50 µg/L FLX exposure on MMCs in histological sections of spleen and head-kidney (HK) of the cichlid fish Cichlasoma dimerus. In the spleen, FLX caused an increase in the area and a decrease in the number of MMCs. An increase in the proportion of the HK occupied by MMCs was observed in FLX-exposed fish, due to an increase in their number but not their area. The deposition rate of MMCs varies according to the hemolymphopoietic organ and would be the result of a differential response to FLX on homeostatic functions (elimination of cellular debris, iron processing and immune response).

Fluoxetine exposure disrupts food intake and energy storage in the cichlid fish Cichlasoma dimerus (Teleostei, Cichliformes).

Human pharmaceuticals are pollutants of special concern due to their widespread consumption over the last decades, their high persistence in the environment, and the reported alterations produced on non-target organism. The antidepressant fluoxetine (FLX) exerts its effect by inhibiting serotonin (5-HT) reuptake at the presynaptic membrane, thus increasing brain serotonergic activity. In vertebrates, there is a clear inverse relationship between hypothalamic 5-HT levels and food intake, therefore we hypothesized that FLX would inhibit food intake, and in consequence alter energy metabolism in freshwater fish. The aim of this study was to analyze the effect of FLX on feeding behavior and energy storage of the cichlid fish Cichlasoma dimerus. Adult fish were intraperitoneally injected daily with 2 or 20 µg.g-1 FLX or saline for a 5-day period, during which the 20 µg.g-1 FLX-injected fish exhibited a marked reduction in food intake, consistent with a decrease in total body weight and total hepatocyte area observed at the end of the experiment. Although not statistically significant, a marked 50% decrease in glycogen and lipid content and an increase in protein levels in liver was observed for the 20 µg.g-1 FLX dose. This was evidenced histochemically by a weak PAS positive reaction and an intense Coomasie Blue stain. Taken together, these results suggest that the SSRI antidepressant FLX produces an anorectic effect in adults of C. dimerus, which could alter normal physiological function and, in consequence, have a negative impact on fish growth, reproduction, and population success.

Bioaccumulation and Distribution Behavior of Endosulfan on a Cichlid Fish: Differences Between Exposure to the Active Ingredient and a Commercial Formulation.

Persistent organic pollutants reach aquatic ecosystems during application and can bioconcentrate/biomagnify because of their lipophilic nature. Toxicological studies focus almost exclusively on the active ingredients of pesticides, instead of commercial formulations, whose toxicity can differ as a result of nonspecified ingredients. The intensive use of endosulfan as a wide-ranging insecticide over the last few decades makes it one of the most frequently detected contaminants in the aquatic environment, even after it has been restricted worldwide. The aim of the present study was to evaluate the bioaccumulation and organ distribution of waterborne endosulfan in the freshwater fish Cichlasoma dimerus, comparing the active ingredient and a commercial formulation. Males were exposed to 0.7 µg/L endosulfan for 2 wk, which was quantified (gas chromatography with an electron capture detector) in the liver, testes, gills, brain, and muscle. The results suggest rapid metabolism of α-endosulfan and ß-endosulfan isomers to endosulfan sulfate (endosulfan-S) in tissues. Isomer levels were highest in gills, indicative of recent uptake. Levels of endosulfan-S were highest in liver and testes for the active ingredient and testes and brain for the commercial formulation. For the active ingredient, endosulfan-S levels showed a positive correlation with organ-lipid percentage. No correlation was evident for the commercial formulation, indicating that the presence of adjuvants alters endosulfan distribution because gills and liver showed a higher uptake and mobilization of ß-endosulfan. These differences in organ distribution may alter tissue-specific toxicity; therefore, additives cannot be considered inactive even if nontoxic. Environ Toxicol Chem 2020;39:604-611. © 2019 SETAC.

Endocrine and reproductive endpoints of Notothenia rossii and N. coriiceps: A baseline study for ecotoxicological monitoring in Antarctic waters.

Pollution threats Antarctica and scientists blame xenobiotics and anthropic activities. Yet little is known about their effect on Antarctic ichthyofauna. Accordingly, we investigated the endocrine system of male fish Notothenia rossii and N. coriiceps (Perciformes, Nototheniidae) collected during Austral summer. For N. rossii, hormone levels of estradiol, testosterone, and 11-ketotestosterone were higher in fish collected during March than those of January; whereas for N. coriiceps estradiol and androgens levels were higher and lower in March, respectively. Histological analysis of the testes showed an unrestricted lobular type with no pathological alterations. However, detection of vitellogenin-like in plasma and skin mucus were seen in 75% of N. coriiceps males and 7% of N. rossii males. This is the first report of mucus vitellogenin-like detection as a non-invasive biomarker of endocrine disruption in notothenioid males and settles a baseline for future studies of reproductive biology and endocrine disruption in Antarctic fishes.

Characterization of the gonadotropin-releasing hormone system in the Neotropical teleost, Steindachneridion parahybae during the annual reproductive cycle in captivity.

This study evaluated by immunohistochemical and Western blot methods, the distribution of two distinct gonadotropin-releasing hormones (GnRHs), corresponding to catfish GnRH (cfGnRH or GnRH1) and chicken-II GnRH (cGnRH-II or GnRH2), in Steindachneridion parahybae females in captivity, focusing these analyses on the reproductive cycle by semi-quantification of optical density (OD). Further, we found that the GnRH neuronal systems co-localized with their respective GnRH-associated peptides (GAPs). A group of neurons immunoreactive (ir) to GnRH1 were identified along the ventral region of the olfactory bulb (vOB) in the telencephalon (vTel) and in the main areas of the diencephalon (especially the medial basal hypothalamus, HBM), including fibers extending into the pituitary gland. In contrast, GnRH2 neurons were confined to the midbrain tegmentum, close to the ventricular surface, without projections to the pituitary gland. Moreover, a cfGAP (GnRH1)-specific band (9 kDa) was identified in the brain and pituitary gland, while a cGAP-II (GnRH2)-specific band (26 kDa) was observed only in the brain extract. During the reproductive cycle, GnRH1-ir presented greater OD values at the vitellogenic and regression stages than at the previtellogenic stage and after artificially induced to spawn. Larger GnRH2-ir neurons were observed during the reproductive cycle, but a higher OD was identified only in the regression stage compared with the other maturation stages. Finally, GnRH1 axons were found to be directed towards the pituitary, and this GnRH type, which is probably the hypophysiotropic form, can contribute to the reproductive dysfunction that occurs in S. parahybae females in captivity, whereas GnRH2 may act as a neuromodulator and/or neurotransmitter.

Effects of waterborne exposure to the antidepressant fluoxetine on swimming, shoaling and anxiety behaviours of the mosquitofish Gambusia holbrooki.

Chemical pollution from pharmaceuticals is increasingly recognized as a major hazard to the aquatic biota. Among the wide variety of pharmaceuticals, fluoxetine (FLX) is one of the most widely prescribed antidepressants, and therefore, it is frequently identified in the aquatic environment. As FLX is designed to alter human behaviour and many physiological pathways are conserved across vertebrates, this drug may affect the behaviour of fish living in FLX-polluted environments. Here, we exposed groups of female mosquitofish Gambusia holbrooki to waterborne FLX for 14 days, under semi-static conditions with daily renewal of test solutions. Following exposure, we conducted a set of behavioural assays in individual fish, aimed at assessing the effects of FLX on their locomotor activity and behavioural responses. We found that FLX impaired swimming behaviour at high concentrations (25 µg/L and 50 µg/L) but not at low concentrations close to environmental levels (1 µg/L and 5 µg/L). When swimming activity was assessed 5 min after transfer of the focal fish to the testing tank, 50 µg/L FLX was the only concentration showing significant effects. However, when the same trials were performed 24 h later, 25 µg/L FLX turned out to be an effect concentration in addition to 50 µg/L. Interestingly, these concentrations would elicit fish plasma concentrations comprised within the range of human therapeutic doses. When subjected to a light/dark preference test, fish showed tendency to remain less time in the dark area at high FLX concentrations, thus suggesting an anti-anxiety response. Shoaling behaviour was not affected by FLX exposure. Our study contributes to the growing body of literature evaluating the effects of FLX on animal behaviour. Regarding the experimental design used in behavioural testing, our findings suggest that focal fish should be subjected to long habituation periods, namely of at least a few hours, in order to better assess the effects of drug exposure.

Action of the Metalloproteinases in Gonadal Remodeling during Sex Reversal in the Sequential Hermaphroditism of the Teleostei Fish Synbranchus marmoratus (Synbranchiformes: Synbranchidae).

Teleostei present great plasticity regarding sex change. During sex reversal, the whole gonad including the germinal epithelium undergoes significant changes, remodeling, and neoformation. However, there is no information on the changes that occur within the interstitial compartment. Considering the lack of information, especially on the role played by metalloproteinases (MMPs) in fish gonadal remodeling, the aim of this study was to evaluate the action of MMPs on gonads of sex reversed females of Synbranchus marmoratus, a fresh water protogynic diandric fish. Gonads were processed for light microscopy and blood samples were used for the determination of plasma sex steroid levels. During sex reversal, degeneration of the ovaries occurred and were gradually replaced by the germinal tissue of the male. The action of the MMPs induces significant changes in the interstitial compartment, allowing the reorganization of germinal epithelium. Leydig cells also showed an important role in female to male reversion. The gonadal transition coincides with changes in circulating sex steroid levels throughout sex reversion. The action of the MMPs, in the gonadal remodeling, especially on the basement membrane, is essential for the establishment of a new functional germinal epithelium.