Sensitivity of different organs and tissues as biomarkers of oxidative stress in juvenile tambaqui (Colossoma macropomum) submitted to fasting.

The present study was conducted to evaluate the effects of fasting on responses of oxidative biomarkers and antioxidant defenses using different organs and tissues of Colossoma macropomum. The fish were divided into two groups: fed (control) and fasting (7 days). After 7 days, the fish were sampled for assessment of oxidative stress biomarkers (MDA-lipid peroxidation and PCO-protein carbonyl) and antioxidant defenses (SOD-superoxide dismutase; CAT-catalase; GPX-glutathione peroxidase; and GST-glutathione-S -transferase) in the liver, intestine, gills, muscle, brain, and plasma. The results showed an increase in MDA, PCO, SOD, and GPX concentrations in the liver and intestine of fasting fish. In contrast, in the branchial tissue, there was a reduction in the activity of SOD and CAT enzymes in fasting fish. There was also a reduction in CAT activity in the muscle of fasting fish, while in the brain, there were no changes in oxidative stress biomarkers. Plasma showed a relatively low antioxidant response. In conclusion, our results confirm that a 7-day fasting period induced tissue-specific antioxidant responses, but the increase in antioxidant responses was only for the SOD and GPX enzymes of the liver and intestine. Additionally, the liver and intestine were the most responsive tissues, whereas the plasma was the least sensitive to oxidative stress.

Spirulina (Arthrospira platensis) supplementation: Impact on growth, metabolism, and antioxidant status in zebrafish.

Zebrafish (Danio rerio) is a valuable model for biomedical research because of its mammalian genetic similarities, rapid reproduction, and low maintenance costs. However, further investigation is required regarding their nutritional requirements and standardized laboratory diets. This study evaluated the metabolic and growth responses of zebrafish juveniles fed on diets supplemented with spirulina, Arthrospira platensis (SP) at different levels for 77 days. Six diets with SP inclusion levels of 0%, 2%, 4%, 6%, 8%, and 10% (SP0-SP10) were formulated. A total of 300 zebrafish juveniles with an average initial weight of 0.113 ± 0.10 g (mean ± SD) were randomly distributed across six groups, with five replicates per group, each containing 10 animals. After 77 days, the SP6 group demonstrated significantly enhanced growth performance compared with the other supplementation levels. The condition factor was markedly higher in the SP6 and SP8 groups than in the SP0 group. No significant effects on total cholesterol levels were observed, but the SP4, SP6, and SP10 diets decreased triglyceride levels. Lipase activity was higher in the SP6 and SP8 groups than in the control group, whereas amylase activity showed no significant differences between treatments. Catalase and superoxide dismutase activities were significantly higher in the SP8 and SP10 groups than in the SP0 and SP2 groups. Glutathione S-transferase activity was higher in the SP6, SP8, and SP10 groups than in the SP0 group. In addition, SP inclusion in zebrafish diets improved female gonadal development. In conclusion, this study indicates that SP supplementation has substantial potential as a growth promoter, positively influencing lipid metabolism and antioxidant enzyme activity without affecting zebrafish survival.

Hematological, biochemical and oxidative responses induced by thermal shock in juvenile Tambaqui (Colossoma macropomum) and its hybrid Tambatinga (Colossoma macropomum x Piaractus brachypomus).

The effects of thermal shock on hematological, biochemical and antioxidant responses were evaluated in liver tissue of juvenile tambaqui (Colossoma macropomum) and tambatinga (♀ C. macropomum × â™‚ Piaractus brachypomus). Forty juveniles of tambaqui and 40 juveniles of tambatinga, of the same age and with an initial weight of 23.3 ± 6.7 g, were randomly distributed in eight 28L circular tanks. A tank (n = 10 fish) of tambaqui and a tank (n = 10 fish) of tambatinga were then used to obtain basal data. The other animals were subjected to thermal shock with sudden temperature reduction from 28 to 18 ºC. Blood and tissue were then collected after 1, 6 and 24 h from the onset of thermal shock. No mortality was observed during the experimental period. Thermal shock increased triglyceride levels after 24 h of stress for tambaqui and reduced values for tambatinga. There was an effect on plasma glucose only for fish group (P < 0.0001) and collection time (P < 0.0001) with a peak observed for the hybrid after 6 h. The interaction of factors for SOD indicated greater activity for tambatinga at the 6 h collection and lower at basal and 1 h collections. There was an interaction for CAT (P = 0.0020) with less activity for tambatinga at 1 h. However, thermal shock and hybridization did not influence GST and TBARS levels in liver tissue. Therefore, the results suggest that the hybrid, tambatinga, is more efficient at promoting adjustments of biochemical responses and antioxidant enzymes during thermal shock.

Obesity induction in adult zebrafish leads to negative reproduction and offspring effects.

Obesity is a transgenerational epigenetic metabolic disturbance. Although the diet-induced obese (DIO) zebrafish model is well established, reproductive parameters and changes in offspring have not yet been evaluated. Thus, the aim of this study was to evaluate possible changes in reproductive parameters, embryos and offspring (F1) generated by the reproduction of diet-induced obese males and females. The adult zebrafish were divided into two groups: one group receiving a balanced diet (control group) and the other group was overfed (DIO group) . The dietary protocol was maintained for 8 weeks. During this period, males and females in the same group were stimulated through a weekly reproduction protocol. To verify parental obesity, body weight, blood glucose, triglyceride, the hepatosomatic and gonadosomatic index and adipose tissue morphometry evaluations were carried out. Reproductive parameters were evaluated through ovarian and oocyte maturation stage, total spawning, fertility and fertilization index. To verify possible changes caused by parenteral obesity, all offspring were kept in separate groups in correspondence with their parents and were fed a control diet. Plasma glucose, triglycerides, mortality rate, hatching, and deformities were determined. After 8 weeks under the diet protocol, the DIO group exhibited characteristic obesity alterations, displaying significant increases in body mass and hepatosomatic and gonadosomatic indices, hyperglycemia and visceral and subcutaneous adipocyte hypertrophy. In addition, high mortality rates, morphologic deformities and high plasmatic glucose and triglyceride levels, with 100% mortality at 60 dpf, were observed for the offspring. Therefore, obesity induction in adults led to negative effects on their offspring, with a high occurrence of deformities and mortality.

Effect of melatonin supplementation to a cytoprotective medium on post-thawed Brycon orbignyanus sperm quality preserved during different freezing times.

The aim of the present study is to verify the viability of frozen B. orbignyanus sperm cells after freezing with dilution media containing different concentrations of the melatonin and after different freezing times. Semen from 15 males was collected and pooled as five pools from three random animals. Oocytes (100) from three females were separately used for fertilization. There were three treatments: (C) Control Medium: 90% of extender Beltsville thawing solution (5% concentration) + 10% methylglycol (MG); (M1) Control Medium + 1 mM melatonin; and (M2) Control Medium + 2 mM melatonin. Sperm samples were diluted in media at a final proportion of 1:4 [125 µl sperm (25% V/V) + 337.5 µl BTS (65% V/V) + 37.5 µl MG (10% V/V)]. Melatonin was added at final solution. Three Dry shipper freezing times were used: T1 (15 min), T2 (12 h) and T3 (24 h). The samples were transferred, stored in a cryobank and thawed in a water bath at 60 °C for 5 s and evaluated concerning viability, morphology and fertilization rate. B. orbignyanus semen frozen in M2 presented the highest fertilization rate (8.40 ± 2.54%). The highest vitality (85.2 ± 2.8%), motility (64.63 ± 8.3%), motility duration (84.22 ± 11.4 s) and progressive motility (17.01 ± 1.2%) rates were maintained for M2. The highest number of altered cells was observed in C (57.4 ± 5.9%). Melatonin at 2 mmol L-1 associated with the cryoprotectant methylglycol in cryopreservation could be used to improve a cryobank for endangered Brycon orbignyanus populations.

Effects of cooling rates on the quality of Prochilodus lineatus (Valenciennes, 1836) sperm.

This study aims to investigate the effect of different cooling rates on the semen cryopreservation of curimba (Prochilodus lineatus). Nineteen ejaculates were obtained from adults males and cryopreserved at 15°C/min (CR15), 30°C/min (CR30) (controlled temperature inside and outside straw, speed was stable during freezing) and direct freezing in liquid nitrogen vapour (~35.6°C/min) (CRNV). The straws were thawed and seminal parameters evaluated. DNA fragmentation through the comet assay was assessed. A fresh sperm sample was not frozen and used for analyses. Data were submitted to an analysis of variance (ANOVA), and means were compared by Scott-Knott test (p < 0.05) using the R Software. Mean motility percentage was 100%, and motility duration was 39.5 ± 5.7 s for the fresh sperm (subjective analysis); 58.9 ± 8.0% and 24.5 ± 5.7 s for CR15; 64.8 ± 4.8% and 26.5 ± 7.1 s for CR30; and 50.1 ± 16% and 25.7 ± 4.7 s for CRNV, respectively. Motility percentages were higher and equal between CR15 and CR30 compared to CRNV (p < 0.05). Some sperm motion kinetics, namely average path velocity (VAP) and straight line velocity (VAS), were higher for CR30 (p < 0.05), while curvilinear velocity (VCL) and velocity progression (PRO) were lower for CRNV (p < 0.05). Straightness (STR) and wobble (WOB) were the same among treatments (p > 0.05). Sperm morphology results indicated higher means for total morphological sperm alterations in CRNV. All cooling rates caused sperm DNA fragmentation, although CR30 provided a less harmful effect. This is the first report for cryopreserved P. lineatus sperm preserved under different controlled cooling rates. The cooling rate of 30°C/min is indicated for the cryopreservation of this fish sperm as it led to the lowest detrimental spermatozoa effects.

Milt cryopreservation for rheophilic fish threatened by extinction in the Rio Grande, Brazil.

BACKGROUND: Specific protocols for milt cryopreservation have been established for some freshwater fish species. However, cryopreservation reduces sperm quality, giving unsatisfactory results in reproduction. OBJECTIVE: The objective of this work was to evaluate the effect of different cryoprotectants on the quality of Prochilodus lineatus, Brycon orbignyanus and Piaractus mesopotamicus milt after cryopreservation. METHODS: The milt was diluted in different cryoprotectant solutions containing 10% methanol, dimethyl sulfoxide, glycerol, propylene glycol or ethylene glycol combined with the Beltsville Thawing Solution extender (5%), then placed in the vapour of a liquid nitrogen (LN) storage tank for 24 h, after which they were immersed in LN. After rewarming, the rate (%) and duration (s) of milt motility and abnormal morphology were evaluated. RESULTS: All of cryoprotectant solutions tested used maintained the viability of P. lineatus and P. mesopotamicus milt. However, in P. lineatus, glycerol ensured a lower percentage of abnormal morphology. In case of P. mesopotamicus, all of the cryoprotectant solutions tested may be used in the cryopreservation process, with the exception of those containing glycerol. CONCLUSION: For B. orbignyanus, cryoprotectant solutions containing methanol and ethylene glycol are recommended for use in the cryopreservation process, although they reduced the quality of sperm post-rewarming.

Metabolic effects of physical exercise on zebrafish (Danio rerio) fed a high-fat diet.

The present study aimed to establish zebrafish as an experimental model for investigations into obesity and physical exercise, as well as to assess the effects of these factors on metabolism. The experiment spanned twelve weeks, comprising a feeding trial during which the last four weeks incorporated a physical exercise protocol. This protocol involved placing fifteen animals in a five-liter aquarium, where they were subjected to swimming at an approximate speed of 0.08 m/s for 30 min daily. Throughout the experiment, histological analyses of visceral, subcutaneous, and hepatic adipose tissues were conducted, along with biochemical analyses of total cholesterol and its fractions, triglycerides, glucose, lactate, and alanine aminotransferase (ALT) levels. Additionally, oxidative stress markers, such as reactive oxygen species (ROS) levels, superoxide dismutase (SOD) activity, and catalase activity and the formation of thiobarbituric acid-reactive substances, were investigated. The results revealed that the group fed a high-fat diet exhibited an increase in ROS production and SOD activity. In contrast, the group administered the high-fat diet and subjected to physical exercise demonstrated a notable reduction in visceral adipocyte area, hepatic steatosis levels, ALT levels, and SOD activity. These findings indicate that physical exercise has a positive effect on obesity and oxidative stress in zebrafish, providing promising evidence for future investigations in this field.

Evaluation of the Anti-Inflammatory and Antioxidant Potential of Cymbopogon citratus Essential Oil in Zebrafish.

This study explored the protective capacity of the essential oil (EO) of Cymbopogon citratus against oxidative stress induced by hydrogen peroxide (H2O2) and the inflammatory potential in zebrafish. Using five concentrations of EO (0.39, 0.78, 1.56, 3.12, and 6.25 µg/mL) in the presence of 7.5 mM H2O2, we analyzed the effects on neutrophil migration, caudal fin regeneration, cellular apoptosis, production of reactive oxygen species (ROS), and activities of the antioxidant enzymes superoxide dismutase (SOD), catalase (CAT), and glutathione S-transferase (GST) after 96 h of exposure. A significant decrease in neutrophil migration was observed in all EO treatments compared to the control. Higher concentrations of EO (3.12 and 6.25 µg/mL) resulted in a significant decrease in caudal fin regeneration compared to the control. SOD activity was reduced at all EO concentrations, CAT activity significantly decreased at 3.12 µg/mL, and GST activity increased at 0.78 µg/mL and 1.56 µg/mL, compared to the control group. No significant changes in ROS production were detected. A reduction in cellular apoptosis was evident at all EO concentrations, suggesting that C. citratus EO exhibits anti-inflammatory properties, influences regenerative processes, and protects against oxidative stress and apoptosis.

Diet with different concentrations of lychee peel flour modulates oxidative stress parameters and antioxidant activity in zebrafish.

The agri-food industry generates substantial waste, leading to significant environmental impacts. Lychee (Litchi chinensis Sonnerat), which is rich in bioactive compounds in its peel, pulp, and seeds, offers an opportunity for waste use. This study aimed to evaluate the effects of supplementing a high-carbohydrate diet with varying levels of lychee peel flour on lipid metabolism biomarkers and oxidative stress in a zebrafish (Danio rerio) model. A total of 225 zebrafish, approximately four months old, were divided into five groups: control, high-carbohydrate (HC), HC2%, HC4%, and HC6%. The study did not find significant differences in the growth performance of zebrafish in any group. However, the HC6% group exhibited a significant decrease in glucose and triglyceride levels compared with the HC group. Furthermore, this group showed enhanced activities of the antioxidant enzymes catalase (CAT) and superoxide dismutase (SOD), along with reduced levels of malondialdehyde (MDA). Increased antioxidant activity was also evidenced by DPPH-, ABTS+, and ß-carotene/Linoleic acid assays in the HC6% group. A positive correlation was identified between SOD/CAT activity and in vitro antioxidant assays. These findings suggest that dietary supplementation with 6% lychee peel flour can significantly modulate glucose homeostasis, lipid metabolism, and antioxidant activity in zebrafish.

Acute ethanol exposure leads to long-term effects on memory, behavior, and transcriptional regulation in the zebrafish brain.

Alcohol consumption is associated with alterations in memory and learning processes in humans and animals. In this context, research models such as the zebrafish (Danio rerio) arise as key organisms in behavioral and molecular studies that attempt to clarify alterations in the Central Nervous System (CNS), like those related to alcohol use. Accordingly, we used the zebrafish as a model to evaluate the effects of ethanol on the learning and memory process, as well as its relationship with behavior and transcriptional regulation of lrfn2, lrrk2, grin1a, and bdnf genes in the brain. To this end, for the memory and learning evaluation, we conducted the Novel Object Recognition test (NOR); for behavior, the Novel Tank test; and for gene transcription, qPCR, after 2 h, 24 h, and 8 days of ethanol exposure. As a result, we noticed in the NOR that after 8 days of ethanol exposure, the control group spent more time exploring the novel object than when compared to 2 h post-exposure, indicating that naturally zebrafish remember familiar objects. In animals in the Treatment group, however, no object recognition behavior was observed, suggesting that alcohol affected the learning and memory processes of the animals and stimulated an anxiolytic effect in them. Regarding transcriptional regulation, 24 h after alcohol exposure, we found hyper-regulation of bdnf and, after 8 days, a hypo-regulation of lrfn2 and lrrk2. To conclude, we demonstrated that ethanol exposure may have influenced learning ability and memory formation in zebrafish, as well as behavior and regulation of gene transcription. These data are relevant for further understanding the application of zebrafish in research associated with ethanol consumption and behavior.

Melatonin Supplementation for the Cryopreservation of Canine Sperm.

Antioxidants can be used in sperm cryopreservation protocols to reduce oxidative stress that occurs due to the cryopreservation process. The aim of this study was to evaluate the effects of melatonin supplementation on quality and oxidative stress parameters in cryopreserved canine sperm. Eighteen sperm ejaculates were collected from 6 Frenchie Bulldog males (3 collections per male). Sperm motility parameters, membrane integrity, and sperm morphology were analyzed before the cryopreservation process. The extender used in cryopreservation was composed of Tris-egg yolk and ethylene glycol 5% was added as a cryoprotectant. The cryoprotective medium was supplemented with 1.0, 1.5, 2.0, 2.5, and 3.0 mM melatonin, and the control group (without melatonin). Post-thaw sperm was evaluated as described for fresh sperm and oxidative stress parameters (lipid peroxidation, catalase, and superoxide dismutase). Post-thaw sperm motility parameters, membrane integrity, sperm morphology, and oxidative stress parameters did not differ (p > 0.05) among the control group and samples supplemented with melatonin. The results of this study showed that melatonin supplementation had no positive or negative effect on the parameters evaluated. Thus, it is suggested that different concentrations of melatonin be tested to assess its effectiveness as an antioxidant in the cryopreservation process in canine sperm.

Ethanol Preference Leads to Alterations in Telomere Length, Mitochondria Copy Number, and Antioxidant Enzyme Activity in Zebrafish Brains.

BACKGROUND: The motivations for and effects of ethanol consumption vary considerably among individuals, and as such, a significant proportion of the population is prone to substance abuse and its negative consequences in the physical, social, and psychological spheres. In a biological context, the characterization of these phenotypes provides clues for understanding the neurological complexity associated with ethanol abuse behavior. Therefore, the objective of this research was to characterize four ethanol preference phenotypes described in zebrafish: Light, Heavy, Inflexible, and Negative Reinforcement. METHODS: To do this, we evaluated the telomere length, mtDNA copy number using real-time quantitative PCR (qPCR), and the activity of these antioxidant enzymes: catalase (CAT), superoxide dismutase (SOD), and glutathione peroxidase (GPx) in the brain, and the interactions between these biomarkers. Changes observed in these parameters were associated with ethanol consumption and alcohol abuse. RESULTS: The Heavy, Inflexible, and Negative Reinforcement phenotypes showed ethanol preference. This was particularly the case with the Inflexible phenotype, which was the group with the greatest ethanol preference. These three phenotypes showed telomere shortening as well as high SOD/CAT and/or GPx activities, while the Heavy phenotype also showed an increase in the mtDNA copy number. However, the Light phenotype, containing individuals without ethanol preference, did not demonstrate any changes in the analyzed parameters even after being exposed to the drug. Additionally, the PCA analysis showed a tendency to cluster the Light and Control groups differently from the other ethanol preference phenotypes. There was also a negative correlation between the results of the relative telomere length and SOD and CAT activity, providing further evidence of the biological relationship between these parameters. CONCLUSIONS: Our results showed differential molecular and biochemistry patterns in individuals with ethanol preference, suggesting that the molecular and biochemical basis of alcohol abuse behavior extends beyond its harmful physiological effects, but rather is correlated with preference phenotypes.

Oxytocin reduces the frequency of aggressive behaviours in male betta fish (Betta splendens).

The objective of this study was to evaluate the effect of visual isolation and the influence of oxytocin on the aggressive behaviour of male Betta splendens and produce an ethogram for analyses. In the first stage, the fish (n = 6) were kept for 60 days in a recirculation system, maintaining visual contact. After this period, the mirror test was applied. The same procedure was performed with the same individuals after 30 days of visual isolation. Visual isolation did no effect on the aggressive behaviour. Application of oxytocin (2.5 µg/g) for 1 h (twice) in this first stage was also conducted after the mirror test. The effect of exposure to oxytocin was evaluated by applying the mirror test 24 h after the second exposure. The second stage was conducted with another 10 individuals exposed to oxytocin (7.5 µg/g) for 1 h, after which the mirror test was performed. Exposure to oxytocin (2.5 µg/g) did not effect on combat-related behaviours. However, in the second stage influenced all behaviours related to combat and display, with an increase in the frequency of "rest" behaviour. The results indicate that exposure to a neuropeptide at a dose of 7.5 µg/g reduces the frequency of aggressive behaviour of male Betta splendens.

The toxicological effects of Eryngium foetidum extracts on zebrafish embryos and larvae depend on the type of extract, dose, and exposure time.

Eryngium foetidum is a herbaceous plant found in tropical and subtropical regions. In vivo pharmacological parameters show that leaf extracts of this plant have antioxidant, anti-inflammatory, antidiabetic, and antimicrobial activities due to their bioactive compounds such as flavonoids and phenols. Despite the evidence for several bioactivities of E. foetidum, information on its safety and tolerability is limited. The objective of this study was to assess the effect and concentration of different extracts of E. foetidum on the development of zebrafish (Danio rerio) embryos. To study the impact of aqueous (AE), ethanolic (EE), and methanolic (ME) extracts, the embryos were exposed to 0.625, 1.25, 2.5, 5, and 10 mg mL-1 for up to 120-h postfertilization to assess embryonic developmental toxicity and then to 0.039, 0.078, 0.156, 0.312, and 0.625 mg mL-1 to assess the antioxidant responses of the enzymes superoxide dismutase catalase, glutathione S-transferase (GST), and cell apoptosis. The results showed that, depending on the extraction solvent, concentration used, and exposure time, E. foetidum extracts caused mortality, altered the hatching time, and promoted changes in enzymatic activities. Delays in development and increased GST activity were found in all treatments. Apoptosis was not observed in any of the treatments. In conclusion, AE, EE, and ME concentrations above 0.625 mg mL-1 can cause adverse effects on the early stages of zebrafish development.

Melatonin administration attenuates acute stress by inducing sleep state in zebrafish (Danio rerio).

Melatonin plays a fundamental homeostatic role in basic biological functions, and an anti-stress role has been also proposed for this hormone. This study aimed to evaluate hormonal, enzymatic and behavioral parameters of zebrafish that received administration of melatonin and were submitted to acute stress. A total of 120 wild-type zebrafish were divided into five groups: naïve control (N), negative control group (Stress/C), positive control treated with diazepam (Stress/Diaz), treatment with melatonin at dose 1 (Stress/Melt. 1) and treatment with melatonin at dose 2 (Stress/Melt. 2). The exposure to treatments (diazepam or melatonin) was performed prior to the acute stress protocol, based on a chase by a fishing net during 5 min followed by exposure to the air for 1 min. The body cortisol levels were assessed, as well as oxidative stress (thiobarbituric acid reactive substances, reactive species of oxygen and antioxidant activity), and fish behavior (open field test). Melatonin was able to modulate acute stress effects on zebrafish by inhibiting cortisol increasing levels, reducing locomotor parameters, inducing a sleep state, reducing lipid peroxidation and stimulating antioxidant enzymatic activity.

Behavioral plasticity and gene regulation in the brain during an intermittent ethanol exposure in adult zebrafish population.

Ethanol consumption is correlated with different neurobiological and behavioral impairments. Acute and chronic exposure to this drug is associated with alterations in the regulation of the mesolimbic dopaminergic system as well as with transcriptional modulation of other receptors in the central nervous system and can unleash seeking behavior or behavioral adaptations and phenotypes such as loss of control, dependence and tolerance. In the present work, we characterized the chronological effects of acute and chronic intermittent exposure to ethanol (1% v/v) in an adult zebrafish population (Danio rerio). During sixteen days of ethanol exposure, we associated the neuromodulation of target genes (drd1, drd2, gabra2a, gabbr1a, gabbr1b) in the central nervous system with behavioral parameters, assessed by social preference, antipredatory capacity and anxiety-like analysis. Transcriptional and behavioral data were collected in days 0, 1, 4, 8, 12 and 16, after ethanol exposure. In days 1 and 4, ethanol exposure increased exploratory behavior regardless of the risk involved (less time spent close to conspecifics and lower avoidance reaction to predator). Along with the reduction of drd2, grin1a and gabra2a transcription seen in the same days, these results suggest an anxiolytic effect of acute ethanol exposure. Interestingly, in days 8, 12 and 16, an attenuation of the behavioral effects was observed. The social preference, antipredatory behavior, perception and exploration parameters were reconstituted. This behavioral re-establishment, accompanied by the increase in drd1, drd2 and gabbr1a transcription in the 8th day could be an indicative of an adaptation to chronic exposure to ethanol. The modulation of drd2 gene combined with the behavioral characterization observed in the study suggests this signalling pathway as a key participant in the phenotypic outcomes of a long-term chronic exposure to ethanol. Lastly, our results reaffirm the ethanol deleterious impacts in perception, ability to respond to adverse stimuli and in anxiety-like behavior.

Inhibition of Lrrk2 reduces ethanol preference in a model of acute exposure in zebrafish.

Due to its multifactorial and yet to be fully understood origin, ethanol addiction is a field that still requires studies for the elucidation of novel genes and pathways that potentially influence the establishment and maintenance of addiction-like phenotypes. In this context, the present study aimed to evaluate the role of the LRRK2 pathway in the modulation of ethanol preference behavior in Zebrafish (Danio rerio). Using the behavioral Conditioned Place Preference (CPP) paradigm, we accessed the preference of animals for ethanol. Next, we evaluated the transcriptional regulation of the gene lrrk2 and the receptors drd1, drd2, grin1a, gria2a, and gabbr1b in the zebrafish brain. Additionally, we used a selective inhibitor of Lrrk2 (GNE-0877) to assess the role of this gene in the preference behavior. Our results revealed four distinct ethanol preference phenotypes (Light, Heavy, Negative Reinforcement, and Inflexible), each showing different transcriptional regulation patterns of the drd1, drd2, grin1a, gria2a, and gabbr1b receptors. We showed that the lrrk2 gene was hyperregulated only in the brains of the animals with the Inflexible phenotype. Most importantly, we showed, for the first time in the context of preference for ethanol, that treatment with the GNE-0877 inhibitor modulates the transcription of the target receptor genes and reduces the preference for ethanol in the animals of the Inflexible group. This result corroborates the hypothesis that the LRRK2 pathway is involved in the inflexible preference for ethanol behavior. Lastly, we identified a possible pharmacological target for the treatment of abusive preference behavior for ethanol.

Profile identification of canicross practitioners and their dogs in Brazil / Identificação do perfil dos praticantes de canicross e seus cães no Brasil / Identificación del perfil de los practicantes de canicross y sus perros en Brasil

The study analyzed the profile of Canicross practitioners in Brazil. A semi- structured questionnaire consisting of 33 questions was used: seven related to the handler and 26 to the dogs. Results: Of the total handlers, 53.3% are women, 64% have practiced the sport for at least 5 years and 52.3% trained with a personal trainer. Of the dogs, 62.6% were females, 60% were between one and four years old and the Mixed Breeds were the majority with 34.6%; as for the health of the animals, 80% of the handlers stated that they did not carry out veterinary monitoring aiming at the Canicross. 6.6% of the animals had sports-related injuries and 80% of these cases received veterinary treatment. It was clear that handlers know strategies to prevent and / or treat injuries caused by the sport, however this is not reflected in dogs. The unprecedented nature of the work will help handlers, veterinarians and sports lovers to draw up protocols so that dogs can practice Canicross safely.

O estudo analisou o perfil dos praticantes de Canicross no Brasil. Foi utili- zado um questionário semiestruturado composto por 33 questões, sendo sete relacionadas ao condutor e 26 aos cães. Resultados: Do total de manipuladores, 53,3% são mulheres, 64% praticam o esporte há pelo menos 5 anos e 52,3% treinam com personal trainer. Dos cães, 62,6% eram fêmeas, 60% tinham entre um e quatro anos e os sem raça definida eram a maioria com 34,6%; quanto à saúde dos animais, 80% dos tratadores afirmaram não realizar acompanhamento veterinário visando o Canicross. 6,6% dos animais tiveram lesões esportivas e 80% desses casos receberam tratamento veterinário. Ficou claro que os tratadores conhecem estratégias para prevenir e/ou tratar lesões causadas pelo esporte, porém isso não se reflete nos cães. O caráter inédito do trabalho ajudará tratadores, veterinários e esportistas a traçarem protocolos para que os cães possam praticar Cani- cross com segurança.

El estudio analizó el perfil de los practicantes de Canicross en Brasil. Se utilizó un cuestionario semiestructurado compuesto por 33 preguntas, siendo siete relaci- onadas con el conductor y 26 con los perros. Resultados: Del total de manejadores, 53,3% son mu-lheres, 64% practican el deporte hace por lo menos 5 años y 52,3% entrenan con trai-ner personal. De los perros, 62,6% eran hembras, 60% tenían entre uno y cuatro años y los sin raza definida eran la mayoría con 34,6%; en cuanto a la salud de los animales, 80% de los manejadores afirmaron no realizar acompañamiento veterinario con vistas al Canicross. El 6,6% de los animales presentaron lesiones deportivas y el 80% de estos casos recibieron tratamiento veterinario. Quedó claro que los cuidadores conocen estra- tegias para prevenir y/o tratar lesiones causadas por el deporte, sin embargo esto no se refleja en los perros. El carácter inédito del trabajo aju-dará a cuidadores, veterinarios y deportistas a trazar protocolos para que los perros puedan practicar Canicross con segu- ridad.

Complexes between methyltestosterone and ß-cyclodextrin for application in aquaculture production.

The inclusion complexes between 17-α-methyltestosterone (MT) and ß-cyclodextrin (bCD) were prepared and characterized in dissolution and solid phase. The complex promoted a sixfold increment in solubility of the hormone. It has a limited solubility and stoichiometry of 2:1 (bCD:MT) determined by DSC, NMR and solubility experiments, the association constant Ka=2846Lmol-1 and complex fraction of 76% (assessed by DOSY-NMR, in (1:3) DMSO/D2O). The association constant obtained in water by the solubility isotherms is 7540Lmol-1. 2D-ROESY experiments indicate the intermolecular orientation (complete inclusion of the hormone in the cavity). Simulations by molecular dynamics agreed with the formation of the inclusion complex 2:1. Release tests showed the slower release for the complexes, with 50% for lyophilization and 56% for malaxation. These results clearly demonstrate the complexation of MT in bCD, which formulations are promising for further applications involving this steroid in aquaculture, both for sexual reversal and in technologies of hormone in water sequestration.