Fluoxetine and Curcumin Prevent the Alterations in Locomotor and Exploratory Activities and Social Interaction Elicited by Immunoinflammatory Activation in Zebrafish: Involvement of BDNF and Proinflammatory Cytokines.

The increase in proinflammatory cytokine expression causes behavioral changes consistent with sickness behavior, and this led to the suggestion that depression might be a psychoneuroimmunological phenomenon. Here, we evaluated the effects of the pretreatment with fluoxetine (10 mg/kg, i.p.) and curcumin (0.5 mg/kg, i.p.) on the immune response elicited by the inoculation of an Aeromonas hydrophila bacterin in zebrafish. Non-pretreated but A. hydrophila-inoculated and sham-inoculated groups of fish served as controls. The social preference, locomotor, exploratory activities, and cerebral expression of il1b, il6, tnfa, and bdnf mRNA were compared among the groups. Behavioral changes characteristic of sickness behavior and a significant increase in the expression of il1b and il6 cytokines were found in fish from the immunostimulated group. The behavioral alterations caused by the inflammatory process were different between males and females, which was coincident with the increased expression of cerebral BDNF. Fluoxetine and curcumin prevented the sickness behavior induced by A. hydrophila and the increased expression of proinflammatory cytokines. Our results point to the potential of zebrafish as a translational model in studies related to neuroinflammation and demonstrate for the first time the effects of fluoxetine and curcumin on zebrafish sickness behavior.

Investigation on the Anticonvulsant Potential of Luteolin and Micronized Luteolin in Adult Zebrafish (Danio rerio).

Epilepsy affects around 50 million people worldwide, and an important number of patients (30%) fail to respond to any available antiepileptic drug. Previous studies have shown that luteolin presents a promising potential as an anticonvulsant. On the other hand, different studies showed that luteolin does not promote anticonvulsant effects. Therefore, there is a lack of consensus about the use of luteolin for seizure control. Luteolin low bioavailability could be a limiting factor to obtain better results. Attractively, micronization technology has been applied to improve flavonoids bioavailability. Thus, the present study aimed to investigate the effects of luteolin on its raw form and micronized luteolin in a PTZ-induced seizure model in adult zebrafish (Danio rerio). Our results demonstrate that luteolin and micronized luteolin did not block PTZ-induced seizures in adult zebrafish. Also, luteolin and micronized luteolin did not provoke behavioral changes. Finally, our results show that 24 h after seizure occurrence, no changes were detected for p70S6Kb, interleukin 1ß, and caspase-3 transcript levels. Altogether, we failed to observe an anticonvulsant potential of luteolin in adult zebrafish, even in its micronized form. However, we recommend new studies to investigate luteolin benefits in epilepsy.

Micronized Resveratrol Shows Anticonvulsant Properties in Pentylenetetrazole-Induced Seizure Model in Adult Zebrafish.

Epilepsy affects 50 million people around the world, and the patients experience cognitive, psychological and social consequences. Despite the considerable quantity of antiepileptic drugs available, 30% of patients still suffer in seizure. Therefore, the advance in therapeutic alternatives is mandatory. Resveratrol has been attracting the attention of many researchers because of its pharmacological potential. However, despite its neuroprotective and anti-epileptic effects, clinical resveratrol use is impaired by its low bioavailability. Here, we applied the supercritical fluid micronization technology (SEDS) to overcome this deficit, and investigated the anticonvulsant potential of micronized resveratrol in a PTZ-induced seizure model in adult zebrafish (Danio rerio). SEDS permits obtaining significantly reduced particle size with a fine size distribution in comparison with the starting material. It can improve the pharmacotherapeutic efficacy. Our data showed that micronized resveratrol decreased the occurrence of the tonic-clonic seizure stage and slowed the development of the seizures in a similar manner of diazepam. Non-processed resveratrol was not able to protect the animals. Furthermore, diazepam decreased the locomotion and exploratory behavior. Differently from diazepam, the micronized resveratrol did not induce behavioral adverse events. In addition, our data showed that the PTZ-induced seizures increased the c-fos transcript levels following the neural excitability. However, the increase in c-fos levels was prevented by micronized resveratrol. In conclusion, our results demonstrate that the micronized resveratrol shows anticonvulsant effect, like the classical antiepileptic drug diazepam in a PTZ-induced seizure model. Excitingly, different from diazepam, micronized resveratrol did not provoke behavioral adverse events.

Erythromycin in the aquatic environment: deleterious effects on the initial development of zebrafish.

Due to the large volume of erythromycin continuously reaching waterbodies and its high persistence, this antibiotic drug has been detected in the aquatic environment at elevated concentrations. Although the problem of the presence of erythromycin in the environment is evident due to its influence in development of antimicrobial resistance, the toxicological consequences on non-target organisms remain to be determined. There are no apparent data on the impact of environmentally relevant concentrations of erythromycin on developing fish. Data on toxic effects during development are essential for evaluation of environmental risk to organisms. Therefore, the aim of this study was to investigate the effects of exposure to erythromycin on certain parameters including hatchability, survival rate, heart rate, and behavior in developing zebrafish. Zebrafish were exposed to a range of environmentally relevant concentrations of antibiotic (0.001, 0.01, 0.1, 1 µg/L) and one concentration 10-fold higher (10 µg/L). Exposure to erythromycin at 0.1 µg/L delayed hatching and decreased survival rate. Exposure to all tested concentrations increased heart rate. Further, exposure to erythromycin at 1 or 10 µg/L enhanced swimming activity. Our results indicated that erythromycin present in the aquatic environment might lead to disabling consequences in developing fish organisms and subsequently may result in ecological imbalance in the natural environment.

Exposure to the herbicide 2,4-dichlorophenoxyacetic acid impairs mitochondrial function, oxidative status, and behavior in adult zebrafish.

2,4-Dichlorophenoxyacetic acid (2,4-D) is one of the most commonly used herbicides worldwide. While the effects of 2,4-D in target organisms are well known, its consequences in nontarget organisms are not fully explained. Therefore, the purpose of this study was to investigate the effects of the herbicide on mitochondrial energy metabolism, oxidative status, and exploratory behavior in adult zebrafish. Animal exposure to 2,4-D increased cytochrome c oxidase and catalase activities and reduced SOD/CAT ratio, moreover, increased the total distance traveled and the number of crossings. Finally, animals exposed to 2,4-D spent more time in the upper zone of the tank and traveled a long distance in the upper zone. Overall, our results indicate the 2,4-D can provoke disabling effects in nontarget organisms. The obtained data showed that exposure to 2,4-D at environmentally relevant concentrations alters mitochondrial metabolism and antioxidant status and disturbs the zebrafish innate behavior.

Exposure to ractopamine hydrochloride induces changes in heart rate and behavior in zebrafish embryos and larvae.

Different veterinary drugs have been widely found in surface and groundwater, affecting non-target organisms. Ractopamine (RAC) is one of these drugs found in water bodies. It is a ß-adrenergic agonist used as a feed additive to modulate the metabolism, redirect nutrients from the adipose tissue towards muscles, and increase protein synthesis in swine, cattle, and turkeys. RAC shows toxicological potential, but there is no data about its impacts on the development of non-target organisms, such as zebrafish (Danio rerio). In this study, we evaluated the effect of the exposure to this feed additive on critical parameters (hatching, survival, spontaneous movement, heart rate, and exploratory and locomotor behavior) in zebrafish embryos and larvae. The animals were exposed to RAC hydrochloride at 0.1, 0.2, 0.85, 8.5, and 85 µg/L. Zebrafish exposed to the drug showed increased heart rate at all tested concentrations and alterations on locomotion and exploratory behavior at 85 µg/L. No changes were observed in the survival, hatching rate and spontaneous movement. Our results suggest that RAC present in the environment can induce disabling effects on non-target organisms and elicit an ecological imbalance by increasing the animals' vulnerability to predation due to greater visibility.

Micronized resveratrol shows promising effects in a seizure model in zebrafish and signalizes an important advance in epilepsy treatment.

Resveratrol is a natural non-flavonoid polyphenolic that has been emerging in epilepsy treatment. Despite its pharmacological properties, the poor bioavailability of resveratrol has been an important barrier that hinders its application as an anticonvulsant. The aim of this work was to improve resveratrol's anticonvulsant effects by micronizing this compound through supercritical fluid micronization technology, which promotes an increase of the particles' surface area and allows significantly reduced particle size to be obtained. We obtained commercial and micronized resveratrol and investigated the anticonvulsant effects of resveratrol as commercially found and micronized resveratrol in a pentylenetetrazole-induced seizure model in zebrafish (Danio rerio) larvae. Diazepam was used as the positive control. Also, animals had their locomotor and exploratory activity analyzed 24 h after the seizure occurrence. The occurrence of the tonic-clonic seizure stage was only prevented by diazepam and micronized resveratrol, unlike the non-processed compound. The seizure development was significantly slowed by diazepam and micronized resveratrol, while non-micronized resveratrol was not able to increase the latency of seizure stages. In addition, diazepam and micronized resveratrol prevented the deleterious effects of pentylenetetrazole-induced seizures on animals' locomotor and exploratory behaviour. Obtained data demonstrates that the micronization process potentiates the anticonvulsant effect of resveratrol. Micronized resveratrol achieved a similar effect to the classical drug diazepam, with the benefit that it may be a safe drug candidate to be used during the neurodevelopmental stage.

A mixture of fipronil and fungicides induces alterations on behavioral and oxidative stress parameters in zebrafish.

Pesticide commercial mixtures, including the insecticide fipronil and the fungicides pyraclostrobin and methyl-thiophanate, have been used in concomitant pest control, facilitating agricultural management. Their widespread use can lead to soil and water contamination and potentially induce damages in the ecosystem, producing toxic effects in non-target organisms. Despite their toxicological potential, their effects on behavioral and biochemical parameters are not well understood. Here we investigated the effects of the mixture of fipronil and fungicides (MFF) pyraclostrobin and methyl- thiophanate on behavioral and biochemical parameters of oxidative stress in adult zebrafish. Animals exposed to the highest MFF tested concentration showed a decrease in the total distance traveled and in the number of crossings in the different zones of the tank. Furthermore, animals exposed to highest MFF tested concentration spent more time in water surface. In addition, our data showed that the exposure to this preparation promoted a decrease in non-protein thiol content as well as in catalase activity. Finally, pesticide exposure induced an increase in the superoxide dismutase/catalase ratio. Our results indicate that alterations in behavioral and oxidative parameters are involved in MFF toxicity in zebrafish. The antioxidant mechanisms analyzed were altered in concentrations that did not affect zebrafish behavior. Therefore, the assessment of oxidative stress parameters in zebrafish brains could be very useful to detect the early effects of environmental exposure to the MFF.

Ractopamine hydrochloride induces behavioral alterations and oxidative status imbalance in zebrafish.

The occurrence of ractopamine (RAC) hydrochloride in water bodies is of significant concern due to its ecological impacts and toxicity to humans. RAC hydrochloride is a ß-adrenergic agonist drug used as a feed additive to (1) improve feed efficiency, (2) rate of weight gain, and (3) increase carcass leanness in animals raised for their meat. This drug is excreted by animals in urine and introduced into the environment affecting nontarget organisms including fish. In wastewater released from farms, RAC concentrations were detected from 0.124 µg/L to 30.1 µg/L, and in levels ranging from 1.3 × 10-5 to 5.4 × 10-4 µg/L in watersheds. The aim of this study was to examine the effects of exposure to RAC at 0.1, 0.2, 0.85, 8.5, or 85 µg/L dissolved in water on behavior and oxidative status in adult zebrafish. At 0.85 µg/L, RAC treatment increased exploratory behavior of zebrafish; while at 8.5 µg/L, decreased locomotor and exploratory activities were noted. With respect to oxidative stress biomarkers, results showed that RAC at 0.2 µg/L induced lipid peroxidation and elevated total thiol content in zebrafish brain. All drug tested concentrations produced a fall in nonprotein thiol content. Finally, RAC at 0.85, 8.5, or 85 µg/L increased catalase enzyme activity. Our results demonstrated that the exposure to RAC induced behavioral alterations and oxidative stress in zebrafish.