Papel do sistema nitrérgico em respostas tipo ansiedade decorrentes de estresse agudo por substância de alarme coespecífica em zebrafish (Danio rerio, Hamilton, 1822)

Abstract

The nitrergic system is one of the key modulators of anxiety-like responses, playing a fundamental role in enabling effective signaling by various neurotransmitters to facilitate the behavioral and physiological adjustments observed in relevant models of biological psychiatry. Driven by the fundamental need to understand the proximal mechanisms underlying anxiety-related responses, this study aimed to evaluate the involvement of the nitrergic system in anxiety-like behaviors resulting from acute exposure to conspecific alarm substance (CAS) in adult zebrafish (Danio rerio). A total of 135 adult fish were used. Of these, 39 served as CAS donors, 72 were subdivided into groups of 12 animals under different experimental conditions for behavioral and biochemical assessments, and 24 were used for histochemical analysis. Animals were treated with either 7-NI (10 or 30 mg/kg), an inhibitor of nitric oxide synthase 1 (NOS-1), or vehicle. Following drug administration, fish were individually exposed to CAS or distilled water. Subsequently, they were subjected to the novel tank test (NTT) and then to the light-dark preference test (LDT). After behavioral testing, brains were dissected for spectrophotometric quantification of nitrite levels and thiobarbituric acid reactive species (TBARS). For the histochemistry experiment, groups of 6 animals were exposed to CAS or distilled water, then dissected for histological sectioning and histochemical staining. The results indicated that NOS-1 blockade with both doses of 7-NI appeared to potentiate anxiety-like responses in animals exposed to both distilled water and CAS, with the 10 mg/kg dose producing an intermediate state between the control and the effect observed with the 30 mg/kg dose. Biochemically, NOS-1 inhibition exerted opposing effects depending on the 7-NI dose on nitrite and TBARS concentrations; the decrease in nitrite levels with the lower 10 mg/kg dose was associated with increased TBARS concentrations, whereas the increase in nitrite with the 30 mg/kg dose was linked to decreased TBARS concentrations. Histochemical analysis revealed that CAS exposure leads to increased nitrergic activity in brain regions including the dorsomedial telencephalon, supracommisural telencephalon, preoptic area, postcommisural telencephalon, dorsal and ventral habenulae, griseum centrale, and superior raphe, while decreased activity was observed in the dorsolateral telencephalon and optic tectum. Overall, the NOS-1 isoform appears to enable the production of NO that modulates anxiety-like responses following acute CAS stress in zebrafish, encompassing distinct behavioral and biochemical adjustments directed toward both anxiogenic and anxiolytic outcomes. Furthermore, CAS exposure alters activity patterns in brain regions associated with defensive circuits, functionally marked by an increased centrality of the locus coeruleus coupled with decreased centrality of the raphe nuclei. Future studies should investigate the impact of potential tonic and phasic actions of nitrergic signaling on defensive responses, as well as the influence of sex, threat relevance, stress controllability, stress duration, and social context on these responses in zebrafish.