Valproic acid exposure affects social visual lateralization and asymmetric gene expression in zebrafish larvae.

Cerebral asymmetry is critical for typical brain function and development; at the same time, altered brain lateralization seems to be associated with neuropsychiatric disorders. Zebrafish are increasingly emerging as model species to study brain lateralization, using asymmetric development of the habenula, a phylogenetically old brain structure associated with social and emotional processing, to investigate the relationship between brain asymmetry and social behavior. We exposed 5-h post-fertilization zebrafish embryos to valproic acid (VPA), a compound used to model the core signs of ASD in many vertebrate species, and assessed social interaction, visual lateralization and gene expression in the thalamus and the telencephalon. VPA-exposed zebrafish exhibit social deficits and a deconstruction of social visual laterality to the mirror. We also observe changes in the asymmetric expression of the epithalamic marker leftover and in the size of the dorsolateral part of the habenula in adult zebrafish. Our data indicate that VPA exposure neutralizes the animals' visual field bias, with a complete loss of the left-eye use bias in front of their own mirror image, and alters brain asymmetric gene expression and morphology, opening new perspectives to investigate brain lateralization and its link to atypical social cognitive development.

Spatial Learning by Using Non-Visual Geometry and a Visual 3D Landmark in Zebrafish (Danio rerio).

Fish conjoin environmental geometry with conspicuous landmarks to reorient towards foraging sites and social stimuli. Zebrafish (Danio rerio) can merge a rectangular opaque arena with a 2D landmark (a blue-colored wall) but cannot merge a rectangular transparent arena with a 3D landmark (a blue cylinder) without training to "feel" the environment thanks to other-than-sight pathways. Thus, their success is linked to tasks differences (spontaneous vs. rewarded). This study explored the reorientation behavior of zebrafish within a rectangular transparent arena, with a blue cylinder outside, proximal to/distal from a target corner position, on the short/long side of the arena. Adult males were extensively trained to distinguish the correct corner from the rotational one, sharing an equivalent metric-sense relationship (short surface left, long surface right), to access food and companions. Results showed that zebrafish's reorientation behavior was driven by both the non-visual geometry and the visual landmark, partially depending on the landmark's proximity and surface length. Better accuracy was attained when the landmark was proximal to the target corner. When long-term experience was allowed, zebrafish handled non-visual and visual sensory stimulations over time for reorienting. We advance the possibility that multisensory processes affect fish's reorientation behavior and spatial learning, providing a link through which to investigate animals' exploratory strategies to face situations of visual deprivation or impairments.

Two Are Better Than One: Integrating Spatial Geometry with a Conspicuous Landmark in Zebrafish Reorientation Behavior.

Within bounded environments of a distinctive shape, zebrafish locate two geometrically equivalent corner positions, based on surface metrics and left-right directions. For instance, the corners with a short surface right/long surface left cannot be distinguished as unique spatial locations unless other cues break the symmetry. By conjoining geometry with a conspicuous landmark, such as a different-color surface, one of the two geometric twins will have a short different-colored surface right, becoming identifiable. Zebrafish spontaneously combine a rectangular white arena's shape with a blue wall landmark, but only when this landmark is near the target corner; when far, that cue triggers a steady attractiveness bias. In this study, we trained zebrafish to use a blue wall landmark in conjunction with a rectangular-shaped arena, providing them rewards over time. We found that trained zebrafish learned to locate the target corner, regardless of the landmark's length and distance, overcoming the attractiveness bias. Zebrafish preferred geometry after removing the landmark (geometric test), but not if put into conflict geometry and landmark (affine transformation). Analysis on movement patterns revealed wall-following exploration as a consistent strategy for approaching the target corner, with individual left-right direction. The capacity of zebrafish to handle different sources of information may be grounds for investigating how environmental changes affect fish spatial behavior in threatened ecosystems.

"Classifying-together" phenomenon in fish (Xenotoca eiseni): Simultaneous exposure to visual stimuli impairs subsequent discrimination learning.

When animals are previously exposed to two different visual stimuli simultaneously, their learning performance at discriminating those stimuli delays: such a phenomenon is known as "classifying-together" or "Bateson effect". However, the consistency of this phenomenon has not been wholly endorsed, especially considering the evidence collected in several vertebrates. The current study addressed whether a teleost fish, Xenotoca eiseni, was liable to the Bateson effect. Three experiments were designed, by handling the visual stimuli (i.e., a full red disk, an amputated red disk, a red cross) and the presence of an exposure phase, before performing a discriminative learning task (Exp. 1: full red disk vs. amputated red disk; Exp. 2: full red disk vs. red cross). In the exposure phase, three conditions per pairs of training stimuli were arranged: "congruence", where fish were exposed and trained to choose the same stimulus; "wide-incongruence", where fish were exposed to one stimulus and trained to choose the other one; "narrow-incongruence", where fish were exposed to both the stimuli and trained to choose one of them. In the absence of exposure (Exp. 3), the discrimination learning task was carried out to establish a baseline performance as regards the full red disk vs. amputated red disk, and the full red disk vs. red cross. Results showed that fish ran into retardation effects at learning when trained to choose a novel stimulus with respect to the one experienced during the exposure-phase (wide-incongruence condition), as well as after being simultaneously exposed to both stimuli (narrow-incongruence condition). Furthermore, there were no facilitation effects due to the congruence compared with the baseline: in such a case, familiar stimuli did not ease the performance at learning. The study provides the first evidence about the consistency of the classifying-together effect in a fish species, further highlighting the impact of visual similarities on discrimination processes.

The Geometric World of Fishes: A Synthesis on Spatial Reorientation in Teleosts.

Fishes navigate through underwater environments with remarkable spatial precision and memory. Freshwater and seawater species make use of several orientation strategies for adaptative behavior that is on par with terrestrial organisms, and research on cognitive mapping and landmark use in fish have shown that relational and associative spatial learning guide goal-directed navigation not only in terrestrial but also in aquatic habitats. In the past thirty years, researchers explored spatial cognition in fishes in relation to the use of environmental geometry, perhaps because of the scientific value to compare them with land-dwelling animals. Geometric navigation involves the encoding of macrostructural characteristics of space, which are based on the Euclidean concepts of "points", "surfaces", and "boundaries". The current review aims to inspect the extant literature on navigation by geometry in fishes, emphasizing both the recruitment of visual/extra-visual strategies and the nature of the behavioral task on orientation performance.

Visual discrimination and amodal completion in zebrafish.

While zebrafish represent an important model for the study of the visual system, visual perception in this species is still less investigated than in other teleost fish. In this work, we validated for zebrafish two versions of a visual discrimination learning task, which is based on the motivation to reach food and companions. Using this task, we investigated zebrafish ability to discriminate between two different shape pairs (i.e., disk vs. cross and full vs. amputated disk). Once zebrafish were successfully trained to discriminate a full from an amputated disk, we also tested their ability to visually complete partially occluded objects (amodal completion). After training, animals were presented with two amputated disks. In these test stimuli, another shape was either exactly juxtaposed or only placed close to the missing sectors of the disk. Only the former stimulus should elicit amodal completion. In human observers, this stimulus causes the impression that the other shape is occluding the missing sector of the disk, which is thus perceived as a complete, although partially hidden, disk. In line with our predictions, fish reinforced on the full disk chose the stimulus eliciting amodal completion, while fish reinforced on the amputated disk chose the other stimulus. This represents the first demonstration of amodal completion perception in zebrafish. Moreover, our results also indicated that a specific shape pair (disk vs. cross) might be particularly difficult to discriminate for this species, confirming previous reports obtained with different procedures.

Learning by Doing: The Use of Distance, Corners and Length in Rewarded Geometric Tasks by Zebrafish (Danio rerio).

Zebrafish spontaneously use distance and directional relationships among three-dimensional extended surfaces to reorient within a rectangular arena. However, they fail to take advantage of either an array of freestanding corners or an array of unequal-length surfaces to search for a no-longer-present goal under a spontaneous cued memory procedure, being unable to use the information supplied by corners and length without some kind of rewarded training. The present study aimed to tease apart the geometric components characterizing a rectangular enclosure under a procedure recruiting the reference memory, thus training zebrafish in fragmented layouts that provided differences in surface distance, corners, and length. Results showed that fish, besides the distance, easily learned to use both corners and length if subjected to a rewarded exit task over time, suggesting that they can represent all the geometrically informative parts of a rectangular arena when consistently exposed to them. Altogether, these findings highlight crucially important issues apropos the employment of different behavioral protocols (spontaneous choice versus training over time) to assess spatial abilities of zebrafish, further paving the way to deepen the role of visual and nonvisual encodings of isolated geometric components in relation to macrostructural boundaries.

The geometry as an eyed fish feels it in spontaneous and rewarded spatial reorientation tasks.

Disoriented human beings and animals, the latter both sighted and blind, are able to use spatial geometric information (metric and sense properties) to guide their reorientation behaviour in a rectangular environment. Here we aimed to investigate reorientation spatial skills in three fish species (Danio rerio, Xenotoca eiseni, Carassius auratus) in an attempt to discover the possible involvement of extra-visual senses during geometric navigation. We observed the fish's behaviour under different experimental procedures (spontaneous social cued task and rewarded exit task), providing them different temporal opportunities to experience the environmental shape (no experience, short and prolonged experience). Results showed that by using spontaneous social cued memory tasks, fishes were not able to take advantage of extra-visual senses to encode the spatial geometry, neither allowing them short time-periods of environmental exploration. Contrariwise, by using a reference memory procedure, during the rewarded exit tasks, thus providing a prolonged extra-visual experience, fishes solved the geometric task, showing also differences in terms of learning times among species.

The role of learning and environmental geometry in landmark-based spatial reorientation of fish (Xenotoca eiseni).

Disoriented animals and humans use both the environmental geometry and visual landmarks to guide their spatial behavior. Although there is a broad consensus on the use of environmental geometry across various species of vertebrates, the nature of disoriented landmark-use has been greatly debated in the field. In particular, the discrepancy in performance under spontaneous choice conditions (sometimes called "working memory" task) and training over time ("reference memory" task) has raised questions about the task-dependent dissociability of mechanisms underlying the use of landmarks. Until now, this issue has not been directly addressed, due to the inclusion of environmental geometry in most disoriented navigation paradigms. In the present study, therefore, we placed our focus on landmark-based navigation in fish (Xenotoca eiseni), an animal model that has provided fruitful research in spatial reorientation. We began with a test of spontaneous navigation by geometry and landmarks (Experiment 1), showing a preference for the correct corner, even in the absence of reinforced training. We then proceeded to test landmarks without the influence of informative geometry through the use of square environments (Experiment 2-4), varying the numerosity of present landmarks, the distance of landmarks from the target corner, and the type of task (i.e., spontaneous cued memory or reference memory). We found marked differences in landmark-use in the absence of environmental geometry. In the spontaneous memory task, visual landmarks acquired perceptive salience (and attracted the fish) but without serving as a spatial cue to location when they were distal from the target. Across learning in the reference memory task, the fish overcame these effects and gradually improved in their performance, although they were still biased to learn visual landmarks near the target (i.e., as beacons). We discuss these results in relation to the existing literature on dissociable mechanisms of spatial learning.

The Environmental Geometry in Spatial Learning by Zebrafish (Danio rerio).

During navigation, disoriented animals learn to use the spatial geometry of rectangular environments to gain rewards. The length of macroscopic surfaces (metric: short/long) and their spatial arrangement (sense: left/right) are powerful cues that animals prove to encode for reorientation. The aim of this study was to investigate if zebrafish (Danio rerio) could take advantage of such geometric properties in a rewarded exit task, by applying a reference memory procedure. The experiment was performed in a rectangular arena having four white walls, where fish were required to choose the two geometrically equivalent exit corners lying on the reinforced diagonal. Results showed that zebrafish encoded the geometry of the arena during reorientation, solving the spatial task within the first 5 days of training. With the aim to avoid the possible influence of extravisual cues on the zebrafish success, we performed a geometric test in extinction of response after the learning day. At test, fish persisted in choosing the two correct corners, thus confirming that the navigation strategy used at training was based on geometric cues. This study adds evidence about the role of geometric frameworks in fish species, and it further validates an effective spatial learning paradigm for zebrafish.

A Detour Task in Four Species of Fishes.

Four species of fish (Danio rerio, Xenotoca eiseni, Carassius auratus, and Pterophyllum scalare) were tested in a detour task requiring them to temporarily abandon the view of the goal-object (a group of conspecifics) to circumvent an obstacle. Fishes were placed in the middle of a corridor, at the end of which there was an opaque wall with a small window through which the goal was visible. Midline along the corridor two symmetrical apertures allowed animals to access two compartments for each aperture. After passing the aperture, fishes showed searching behavior in the two correct compartments close to the goal, appearing able to localize it, although they had to temporarily move away from the object's view. Here we provide the first evidence that fishes can solve such a detour task and therefore seem able to represent the "permanence in existence" of objects, which continue to exist even if they are not momentarily visible.