Conserved Serotonergic Background of Experience-Dependent Behavioral Responsiveness in Zebrafish (Danio rerio).

Forming effective responses to threatening stimuli requires the adequate and coordinated emergence of stress-related internal states. Such ability depends on early-life experiences and, in connection, the adequate formation of neuromodulatory systems, particularly serotonergic signaling. Here, we assess the serotonergic background of experience-dependent behavioral responsiveness using male and female zebrafish (Danio rerio). For the first time, we have characterized a period during behavioral metamorphosis in which zebrafish are highly reactive to their environment. Absence of social stimuli during this phase established by isolated rearing fundamentally altered the behavioral phenotype of postmetamorphic zebrafish in a challenge-specific manner, partially due to reduced responsiveness and an inability to develop stress-associated arousal state. In line with this, isolation differentially affected whole-brain serotonergic signaling in resting and stress-induced conditions, an effect that was localized in the dorsal pallium and was negatively associated with responsiveness. Administration of the serotonin receptor 1A partial agonist buspirone prevented the isolation-induced serotonin response to novelty in the level of the whole brain and the forebrain as well, without affecting catecholamine levels, and rescued stress-induced arousal along with challenge-induced behaviors, which together indicates functional connection between these changes. In summary, there is a consistent negative association between behavioral responsiveness and serotonergic signaling in zebrafish, which is well recognizable through the modifying effects of developmental perturbation and pharmacological manipulations as well. Our results imply a conserved serotonergic mechanism that context-dependently modulates environmental reactivity and is highly sensitive to experiences acquired during a specific early-life time window, a phenomenon that was previously only suggested in mammals.SIGNIFICANCE STATEMENT The ability to respond to challenges is a fundamental factor in survival. We show that zebrafish that lack appropriate social stimuli in a sensitive developmental period show exacerbated alertness in nonstressful conditions while failing to react adequately to stressors. This shift is reflected inversely by central serotonergic signaling, a system that is implicated in numerous mental disorders in humans. Serotonergic changes in brain regions modulating responsivity and behavioral impairment were both prevented by the pharmacological blockade of serotonergic function. These results imply a serotonergic mechanism in zebrafish that transmits early-life experiences to the later phenotype by shaping stress-dependent behavioral reactivity, a phenomenon that was previously only suggested in mammals. Zebrafish provide new insights into early-life-dependent neuromodulation of behavioral stress-responses.

The swimming plus-maze test: a novel high-throughput model for assessment of anxiety-related behaviour in larval and juvenile zebrafish (Danio rerio).

Larval zebrafish (Danio rerio) has the potential to supplement rodent models due to the availability of resource-efficient, high-throughput screening and high-resolution imaging techniques. Although behavioural models are available in larvae, only a few can be employed to assess anxiety. Here we present the swimming plus-maze (SPM) test paradigm, a tool to assess anxiety-related avoidance of shallow water bodies in early developmental stages. The "+" shaped apparatus consists of arms of different depth, representing different levels of aversiveness similarly to the rodent elevated plus-maze. The paradigm was validated (i) in larval and juvenile zebrafish, (ii) after administration of compounds affecting anxiety and (iii) in differentially aversive experimental conditions. Furthermore, we compared the SPM with conventional "anxiety tests" of zebrafish to identify their shared characteristics. We have clarified that the preference of deeper arms is ontogenetically conserved and can be abolished by anxiolytic or enhanced by anxiogenic agents, respectively. The behavioural readout is insensitive to environmental aversiveness and is unrelated to behaviours assessed by conventional tests involving young zebrafish. Taken together, we have developed a sensitive high-throughput test allowing the assessment of anxiety-related responses of zebrafish regardless of developmental stage, granting the opportunity to combine larva-based state-of-the-art methods with detailed behavioral analysis.