Cannabinoid system of dorsomedial telencephalon modulates behavioral responses to noxious stimulation in the fish Leporinus macrocephalus.

Fish dorsomedial telencephalon has been considered a pallial region homologous to mammals amygdala, being considered a possible substrate for nociception modulation in this animal group. The present study aimed to evaluate the participation of the cannabinoid system of Dm telencephalon on nociception modulation in the fish Leporinus macrocephalus. We demonstrated that cannabidiol microinjection in Dm telecephalon inhibits the behavioral nociceptive response to the subcutaneous injection of 3% formaldehyde, and this antinociception is blocked by previous treatment with AM251 microinjection. Furthermore, AM251 microinjection in Dm prior to restraint stress also blockades the stress-induced antinociception. These results reinforce the hypothesis that this pallial telencephalic structure has a pivotal role in nociception modulation in fish.

Acute fluoxetine treatment increases aggressiveness in juvenile matrinxã (Brycon amazonicus).

Fluoxetine (FLX) is a selective serotonin (5-HT) reuptake inhibitor known for its effects modifying aggressiveness, personality traits, and anxiety-like behaviors. The aim of the present study was to evaluate the influence of the acute treatment, by immersion, with FLX on aggressive behavior of resident Brycon amazonicus fish. Fish pretreated with FLX presented an increase in aggressiveness, evidenced by the increase on the number of bites and chases against the intruder and a decrease in latency for the first attack, when compared to control fish. Together with previous studies, these results show the complexity of the neural modulation of the aggressive behavior in fish by 5-HTergic system.

Social challenge increases cortisol and hypothalamic monoamine levels in matrinxã (Brycon amazonicus).

The neural circuitry for social behavior and aggression appears to be evolutionarily conserved across the vertebrate subphylum and involves a complex neural network that includes the hypothalamus as a key structure. In the present study, we evaluated the changes in monoamine levels in the hypothalamus and on serum cortisol and plasma glucose of resident matrinxã (Brycon amazonicus) submitted to a social challenge (introduction of an intruder in their territory). The fight promoted a significant increase in hypothalamic 5-HT, NA and DA levels and on the metabolites 5-HIAA and DOPAC, and decreased 5-HIAA/5-HT and DOPAC/DA ratios in resident fish. Furthermore, an increase in serum cortisol and plasma glucose was also observed after the fight. Resident fish presented a high aggressiveness even with increased 5-HT levels in the hypothalamus. The alteration in hypothalamic monoaminergic activity of matrinxã suggests that this diencephalic region is involved in aggression and stress modulation in fish; however, it does not exclude the participation of other brain areas not tested here.

GABAA-benzodiazepine receptors in the dorsomedial (Dm) telencephalon modulate restraint-induced antinociception in the fish Leporinus macrocephalus.

The possibility that fish experience pain has been denied based on the absence of the neural substrates to support this "experience". In this context, the identification of brain regions involved in nociception modulation could provide important insights regarding the processing of nociceptive information in fish. Our study evaluated the participation of the GABAA-benzodiazepine receptor in the dorsomedial (Dm) telencephalon in restraint-induced antinociception in the fish Leporinus macrocephalus through the microinjection of the anxiolytic drug midazolam. The microinjection of midazolam in the Dm did not alter the nocifensive response; however, this drug did block the inhibition of the nocifensive response to formaldehyde promoted by restraint stress. The fish that received midazolam (40nmol) microinjection prior to restraint (3 or 5min), followed by subcutaneous injection with formaldehyde presented a higher distance traveled than the fish that received saline microinjection. This effect might reflect the specific action of midazolam on benzodiazepine receptors in the Dm telencephalon, as pre-treatment with flumazenil, a benzodiazepine receptor antagonist, inhibited the effects of this drug. In the present study, we present the first evidence demonstrating a role for the dorsomedial telencephalic region in the modulation of stress-induced antinociception in fish, revealing new perspectives in the understanding of nociceptive information processing in this group.

Acute administration of a cannabinoid CB1 receptor antagonist impairs stress-induced antinociception in fish.

This study evaluated the influence of the pre-treatment with AM251 (a cannabinoid type I receptor (CB1) selective antagonist) on the stress-induced antinociception promoted by restraint in the fish Leporinus macrocephalus. The application of 3 and 5 min of restraint stress promoted an inhibition of the behavioural response to the subcutaneous injection of 3% formaldehyde (increase in locomotor activity), suggesting the activation of an antinociceptive system. The acute intraperitoneal administration of AM251 (3 mg·kg(-1)) impaired this antinociceptive response induced by 3 and 5 min of restraint stress. The fish treated with AM251 before the application of restraint stress presented an increase in locomotor activity after the subcutaneous injection of formaldehyde, similar to fish not exposed to restraint, suggesting that the stress-induced antinociception promoted by restraint in fish is probably mediated by cannabinoid CB1 receptors. The results presented in this paper suggest the participation of the endocannabinoid system in nociception modulation in fish, supporting the hypothesis that an endogenous antinociceptive system activated by restraint stress is present in fish and that the modulation of antinociception by the CB1 receptor is evolutionary well-conserved across vertebrates.

Novelty of the arena impairs the cortisol-related increase in the aggression of matrinxã (Brycon amazonicus).

The dichotomic effect of a cortisol level rise in vertebrate behavior has been widely observed. Generally, a chronic increase of the hormone level inhibits aggression, while an acute rise increases aggression. However, in this study, we show that this increase in aggression through an acute rise of cortisol also depends on the context in which the agonistic interaction occurs in the tropical fish matrinxã, Brycon amazonicus. We combined two factors: the type of housing (resident or non-resident in the trial arena) and the level of cortisol at the beginning of the fight (normal level - control, or high level - hydrocortisone-treated fish). The cortisol treatment increased the aggressiveness in the resident fish, but this effect was not observed in the non-resident fish, which fought in an unknown arena. The novelty of the arena may have elicited an "alerted state" in the non-resident fish; in this situation the fight was not the priority, and the cortisol effect in aggression could be impaired by a conflict between motivational systems (fear and aggression). In our knowledge, in fish, the increase of aggression promoted by an acute rise in cortisol levels was always tested and observed in a resident context, and the inhibition of cortisol effect in the agonist behavior is demonstrated for the first time. As the cortisol effect in aggression is observed in several taxa, the inhibition of aggressiveness increased by the novelty of the arena should be investigated in other groups to clarify the dynamics of this effect of cortisol in animal behavior.

Stress-induced antinociception in fish reversed by naloxone.

Pain perception in non-mammalian vertebrates such as fish is a controversial issue. We demonstrate that, in the fish Leporinus macrocephalus, an imposed restraint can modulate the behavioral response to a noxious stimulus, specifically the subcutaneous injection of 3% formaldehyde. In the first experiment, formaldehyde was applied immediately after 3 or 5 min of the restraint. Inhibition of the increase in locomotor activity in response to formaldehyde was observed, which suggests a possible restraint-induced antinociception. In the second experiment, the noxious stimulus was applied 0, 5, 10 and 15 min after the restraint, and both 3 and 5 min of restraint promoted short-term antinociception of approximately 5 min. In experiments 3 and 4, an intraperitoneal injection of naloxone (30 mg.kg(-1)) was administered 30 min prior to the restraint. The 3- minute restraint-induced antinociception was blocked by pretreatment with naloxone, but the corresponding 5-minute response was not. One possible explanation for this result is that an opioid and a non-preferential µ-opioid and/or non-opioid mechanism participate in this response modulation. Furthermore, we observed that both the 3- and 5- minutes restraint were severely stressful events for the organism, promoting marked increases in serum cortisol levels. These data indicate that the response to a noxious stimulus can be modulated by an environmental stressor in fish, as is the case in mammals. To our knowledge, this study is the first evidence for the existence of an endogenous antinociceptive system that is activated by an acute standardized stress in fish. Additionally, it characterizes the antinociceptive response induced by stress in terms of its time course and the opioid mediation, providing information for understanding the evolution of nociception modulation.

Dietary L-tryptophan alters aggression in juvenile matrinxã Brycon amazonicus.

This study evaluated the effect of dietary supplementation with L: -tryptophan (L-TRP), a serotonin precursor, on the aggressiveness of juvenile matrinxã Brycon amazonicus. Fish were kept in individual aquaria for 7 days receiving the diets: D1 (control: 0.47% of TRP), D2 (0.94% of TRP), D3 (1.88% of TRP), and D4 (3.76% of TRP). After this, they were grouped with an intruder fish to establish a resident-intruder relationship during periods of 20 min. Blood cortisol, glucose, chloride, sodium and calcium; hemoglobin, hematocrit, red blood cell count and volume; liver glycogen and lipids were measured. Territoriality had significant effect on the aggressiveness of matrinxã (the residents were more aggressive than intruders, P < 0.001) and tryptophan significantly affected their behavior. Fish fed with the D2 diet presented a longer latency until the first attack (P = 0.0069) and bit the intruder fewer times (P = 0.0136) during the period of observation, compared to the control group. The frequency of bites and chases after the first attack was not affected by the dietary supplementation of TRP. Physiological variables were not significantly affected by the diet, except for a moderate increase in cortisol level in fish fed with D2 diet after the fight, indicating slight activation of the hypothalamus-pituitary-interrenal axis. The results show that juvenile matrinxã have aggressive and territorial behavior and that a diet containing 9.4 g TRP kg(-1) alter their aggressiveness, without affecting the stress-related physiological parameters.