The domestic chick as an animal model of autism spectrum disorder: building adaptive social perceptions through prenatally formed predispositions.

Equipped with an early social predisposition immediately post-birth, humans typically form associations with mothers and other family members through exposure learning, canalized by a prenatally formed predisposition of visual preference to biological motion, face configuration, and other cues of animacy. If impaired, reduced preferences can lead to social interaction impairments such as autism spectrum disorder (ASD) via misguided canalization. Despite being taxonomically distant, domestic chicks could also follow a homologous developmental trajectory toward adaptive socialization through imprinting, which is guided via predisposed preferences similar to those of humans, thereby suggesting that chicks are a valid animal model of ASD. In addition to the phenotypic similarities in predisposition with human newborns, accumulating evidence on the responsible molecular mechanisms suggests the construct validity of the chick model. Considering the recent progress in the evo-devo studies in vertebrates, we reviewed the advantages and limitations of the chick model of developmental mental diseases in humans.

Temporal hampering of thyroid hormone synthesis just before hatching impeded the filial imprinting in domestic chicks.

Thyroid hormones play a critical role in the initiation of the sensitive period of filial imprinting. The amount of thyroid hormones in the brains of chicks increases intrinsically during the late embryonic stages and peaks immediately before hatching. After hatching, a rapid imprinting-dependent inflow of circulating thyroid hormones into the brain occurs via vascular endothelial cells during imprinting training. In our previous study, inhibition of hormonal inflow impeded imprinting, indicating that the learning-dependent inflow of thyroid hormones after hatching is critical for the acquisition of imprinting. However, it remained unclear whether the intrinsic thyroid hormone level just before hatching affects imprinting. Here, we examined the effect of temporal thyroid hormone decrease on embryonic day 20 on approach behavior during imprinting training and preference for the imprinting object. To this end, methimazole (MMI; a thyroid hormone biosynthesis inhibitor) was administered to the embryos once a day on days 18-20. Serum thyroxine (T4) was measured to evaluate the effect of MMI. In the MMI-administered embryos, the T4 concentration was transiently reduced on embryonic day 20 but recovered to the control level on post-hatch day 0. At the beginning of imprinting training on post-hatch day 1, control chicks approached the imprinting object only when the object was moving. In the late phase of training, control chicks subsequently approached towards the static imprinting object. On the other hand, in the MMI-administered chicks, the approach behavior decreased during the repeated trials in the training, and the behavioral responses to the imprinting object were significantly lower than those of control chicks. This indicates that their persistent responses to the imprinting object were impeded by a temporal thyroid hormone decrease just before hatching. Consequently, the preference scores of MMI-administered chicks were significantly lower than those of control chicks. Furthermore, the preference score on the test was significantly correlated with the behavioral responses to the static imprinting object in the training. These results indicate that the intrinsic thyroid hormone level immediately before hatching is crucial for the learning process of imprinting.

Naïve chicks do not prefer objects with stable body orientation, though they may prefer behavioural variability.

Domestic chicks (Gallus gallus domesticus) have been widely used as a model to study the motion cues that allow visually naïve organisms to detect animate agents shortly after hatching/birth. Our previous work has shown that chicks prefer to approach agents whose main body axis and motion direction are aligned (a feature typical of creatures whose motion is constrained by a bilaterally symmetric body plan). However, it has never been investigated whether chicks are also sensitive to the fact that an agent maintains a stable front-back body orientation in motion (i.e. consistency in which end is leading and which trailing). This is another feature typical of bilateria, which is also associated with the detection of animate agents in humans. The aim of the present study was to fill this gap. Contrary to our initial expectations, after testing 300 chicks across 3 experimental conditions, we found a recurrent preference for the agent which did not maintain a stable front-back body orientation. Since this preference was limited to female chicks, the results are discussed also in relation to sex differences in the social behaviour of this model. Overall, we show for the first time that chicks can discriminate agents based on the stability of their front-back orientation. The unexpected direction of the effect could reflect a preference for agents' whose behaviour is less predictable. Chicks may prefer agents with greater behavioural variability, a trait which has been associated with animate agents, or have a tendency to explore agents performing "odd behaviours".

Life is in motion (through a chick's eye).

Cognitive scientists, social psychologists, computer scientists, neuroscientists, ethologists and many others have all wondered how brains detect and interpret the motion of living organisms. It appears that specific cues, incorporated into our brains by natural selection, serve to signal the presence of living organisms. A simple geometric figure such as a triangle put in motion with specific kinematic rules can look alive, and it can even seem to have intentions and goals. In this article, we survey decades of parallel investigations on the motion cues that drive animacy perception-the sensation that something is alive-in non-human animals, especially in precocial species, such as the domestic chick, to identify inborn biological predispositions. At the same time, we highlight the relevance of these studies for an understanding of human typical and atypical cognitive development.

Spontaneous preference for unpredictability in the temporal contingencies between agents' motion in naive domestic chicks.

The ability to recognize animate agents based on their motion has been investigated in humans and animals alike. When the movements of multiple objects are interdependent, humans perceive the presence of social interactions and goal-directed behaviours. Here, we investigated how visually naive domestic chicks respond to agents whose motion was reciprocally contingent in space and time (i.e. the time and direction of motion of one object can be predicted from the time and direction of motion of another object). We presented a 'social aggregation' stimulus, in which three smaller discs repeatedly converged towards a bigger disc, moving in a manner resembling a mother hen and chicks (versus a control stimulus lacking such interactions). Remarkably, chicks preferred stimuli in which the timing of the motion of one object could not be predicted by that of other objects. This is the first demonstration of a sensitivity to the temporal relationships between the motion of different objects in naive animals, a trait that could be at the basis of the development of the perception of social interaction and goal-directed behaviours.

Pitch-Luminance Crossmodal Correspondence in the Baby Chick: An Investigation on Predisposed and Learned Processes.

Our senses are constantly reached by a multitude of stimuli from all different sensory modalities. To create a coherent representation of the environment, we must integrate the various unimodal inputs that refer to the same object into a single multimodal representation. In some cases, however, we tend to bind certain properties of the stimuli without any apparent reason, which is a phenomenon named crossmodal correspondence. For instance, we match a spiky or a rounded shape with the sound "Kiki" or "Bouba", respectively. Similarly, we associate the left hemispace with low luminance and the right one with high luminance. Instances of crossmodal correspondences were described also in other mammals, and recently, a case of space-luminance crossmodal correspondence was reported in birds (i.e., domestic chicks). Here, we investigate the presence of pitch-luminance crossmodal correspondence in three-day-old chicks, employing experimental methods that exploit either predisposed or learned processes. While failing to report evidence for this phenomenon, we discuss the difference between statistical and structural crossmodal correspondences and the possible role of environmental factors in determining their emergence. Moreover, we discuss the importance of the different experimental methodologies to investigate distinct aspects of this perceptual phenomenon to reach a deeper understanding and unveil the role of innate vs. learned mechanisms.

Embryonic Valproate Exposure Alters Mesencephalic Dopaminergic Neurons Distribution and Septal Dopaminergic Gene Expression in Domestic Chicks.

In recent years, the role of the dopaminergic system in the regulation of social behavior is being progressively outlined, and dysfunctions of the dopaminergic system are increasingly associated with neurodevelopmental disorders, including autism spectrum disorder (ASD). To study the role of the dopaminergic (DA) system in an animal model of ASD, we investigated the effects of embryonic exposure to valproic acid (VPA) on the postnatal development of the mesencephalic DA system in the domestic chick. We found that VPA affected the rostro-caudal distribution of DA neurons, without changing the expression levels of several dopaminergic markers in the mesencephalon. We also investigated a potential consequence of this altered DA neuronal distribution in the septum, a social brain area previously associated to social behavior in several vertebrate species, describing alterations in the expression of genes linked to DA neurotransmission. These findings support the emerging hypothesis of a role of DA dysfunction in ASD pathogenesis. Together with previous studies showing impairments of early social orienting behavior, these data also support the use of the domestic chick model to investigate the neurobiological mechanisms potentially involved in early ASD symptoms.

Imprintability of Newly Hatched Domestic Chicks on an Artificial Object: A Novel High Time-Resolution Apparatus Based on a Running Disc.

In filial imprinting, newly hatched chicks repeatedly approach a conspicuous object nearby and memorize it, even though it is an artificial object instead of their mother hen. Imprinting on an artificial object in a laboratory setting has a clear sensitive period from post hatch days 1-3 in the case of domestic chicks. However, the establishment of imprintability are difficult to investigate because of the limitations of the behavioral apparatus. In this study, we developed a novel behavioral apparatus, based on a running disc, to investigate the learning processes of imprinting in newly hatched domestic chicks. In the apparatus, the chick repeatedly approaches the imprinting object on the disc. The apparatus sends a transistor-transistor-logic signal every 1/10 turn of the disc to a personal computer through a data acquisition system following the chick's approach to the imprinting object on the monitor. The imprinting training and tests were designed to define the three learning processes in imprinting. The first process is the one in which chicks spontaneously approach the moving object. The second is an acquired process in which chicks approach an object even when it is static. In the third process, chicks discriminate between the differently colored imprinting object and the control object in the preference test. Using the apparatus, the difference in the chicks' behavior during or after the sensitive period was examined. During the sensitive period, the chicks at post hatch hour 12 and 18 developed the first imprinting training process. The chicks at post hatch hour 24 maintained learning until the second process. The chicks at post hatch hour 30 reached the discrimination process in the test. After the sensitive period, the chicks reared in darkness until post hatch day 4 exhibited poor first learning process in the training. Thus, this apparatus will be useful for the detection of behavioral changes during neuronal development and learning processes.

Young domestic chicks spontaneously represent the absence of objects.

Absence is a notion that is usually captured by language-related concepts like zero or negation. Whether nonlinguistic creatures encode similar thoughts is an open question, as everyday behavior marked by absence (of food, of social partners) can be explained solely by expecting presence somewhere else. We investigated 8-day-old chicks' looking behavior in response to events violating expectations about the presence or absence of an object. We found different behavioral responses to violations of presence and absence, suggesting distinct underlying mechanisms. Importantly, chicks displayed an avian signature of novelty detection to violations of absence, namely a sex-dependent left-eye-bias. Follow-up experiments excluded accounts that would explain this bias by perceptual mismatch or by representing the object at different locations. These results suggest that the ability to spontaneously form representations about the absence of objects likely belongs to the initial cognitive repertoire of vertebrate species.

Subtype-selective contribution of muscarinic acetylcholine receptors for filial imprinting in newly-hatched domestic chicks.

Muscarinic acetylcholine receptors (mAChRs) play an important role in many brain functions. Our previous study revealed that the injection of mAChRs antagonist scopolamine into the intermediate medial mesopallium (IMM) region, which is critical for filial imprinting, impairs memory formation. In avian brains, four mAChR subtypes have been identified (M2, M3, M4 and M5). M3 and M5 receptors increase the excitability of neurons, whereas M2 and M4 receptors reduce the excitability. Because the scopolamine blocks all subtypes, the previous study did not identify which subtype contributes to the memory formation. By injecting several types of mAChR antagonists into the IMM, in this study we determined which mAChR subtype plays a critical role in imprinting. First, the effects of antagonists on the excitatory receptor subtypes M3 and M5 were examined. Injection of the M3 antagonist (DAU5884) at 20 mM or the M5 antagonist (ML381) at 2 mM impaired imprinting. Considering the pKi value of DAU5884, the impairment seems to be caused by DAU5884 binding to M3 and/or M4 receptors. Second, the effect of antagonists on the inhibitory receptor subtype M2 was examined. The results showed that the M2 antagonist (AQ-RA741) impaired imprinting at a concentration of 20 mM. Considering the pKi value of AQ-RA741, the impairment seems to be caused by AQ-RA741 binding to M2 and/or M4. The findings of this study suggests that the excitatory receptor subtypes M3 and M5 and the inhibitory receptor subtype M2 and/or M4 cooperate to achieve the appropriate balance of acetylcholine signaling to execute imprinting.

Gene expression profiles of the muscarinic acetylcholine receptors in brain regions relating to filial imprinting of newly-hatched domestic chicks.

Muscarinic acetylcholine receptors (mAChRs) in the central nervous system play an important role in regulating complex functions such as learning, memory, and selective attention. Five subtypes of the mAChRs (M1-M5) have been identified in mammals, and are classified into two subfamilies: excitatory (M1, M3, and M5) and inhibitory (M2 and M4) subfamilies. Filial imprinting of domestic chicks is a useful model in the laboratory to investigate the mechanisms of memory formation in early learning. We recently found that mAChRs in the intermediate medial mesopallium (IMM) are involved in the memory formation of imprinting. However, expression profiles of each mAChR subtype in the brain regions including the IMM remain unexplored. Here we show the unique gene expression of each mAChR subtype in the pallial regions involved in imprinting. In terms of the excitatory mAChRs, M5 was expressed in the IMM region and other parts of the pallium, whereas M3 was less expressed in the IMM but highly expressed in the hyperpallium and nidopallium. Regarding the inhibitory mAChRs, M2 was sparsely distributed but clearly in some cells throughout the pallial regions. M4 was highly expressed in the IMM region and other parts of the pallium. These expression profiles can be used as a basis for understanding cholinergic modulation in the memory formation of imprinting and other learning processes in birds, and compared to those of mammals.

Light-incubation effects on lateralisation of single unit responses in the visual Wulst of domestic chicks.

Since the ground-breaking discovery that in-egg light exposure triggers the emergence of visual lateralisation, domestic chicks became a crucial model for research on the interaction of environmental and genetic influences for brain development. In domestic chick embryos, light exposure induces neuroanatomical asymmetries in the strength of visual projections from the thalamus to the visual Wulst. Consequently, the right visual Wulst receives more bilateral information from the two eyes than the left one. How this impacts visual Wulst's physiology is still unknown. This paper investigates the visual response properties of neurons in the left and right Wulst of dark- and light-incubated chicks, studying the effect of light incubation on bilaterally responsive cells that integrate information from both eyes. We recorded from a large number of visually responsive units, providing the first direct evidence of lateralisation in the neural response properties of units of the visual Wulst. While we confirm that some forms of lateralisation are induced by embryonic light exposure, we found also many cases of light-independent asymmetries. Moreover, we found a strong effect of in-egg light exposure on the general development of the functional properties of units in the two hemispheres. This indicates that the effect of embryonic stimulation goes beyond its contribution to the emergence of some forms of lateralisation, with influences on the maturation of visual units in both hemispheres.

Space-luminance crossmodal correspondences in domestic chicks.

Crossmodal correspondences are spontaneous associations of non-redundant information across different modalities. Infants and some non-human animals (i.e., chimpanzees, rhesus monkeys, and dogs) showed crossmodal correspondences like adult humans, suggesting a shared origin (at least among mammals) of such a phenomenon. Here we investigate visual-spatial crossmodal correspondences in a precocial avian species, i.e., the domestic chicken. Three-day-old chicks (n = 40) were presented with two (one in the left and one in the right hemispace of an arena) identical panels, either dyed black (low luminance) or white (high luminance). Chicks could circumnavigate either panel to obtain a food reward. Akin to humans, they preferentially chose the left side when presented with black panels and the right side when presented with white panels. The control group (n = 39), tested with grey panels, showed no spatial preference. In light of our results, we discuss crossmodal correspondences in terms of an early available mechanism widespread across different species.

Anatomical asymmetries in the tectofugal pathway of dark-incubated domestic chicks: Rightwards lateralization of parvalbumin neurons in the entopallium.

The discovery of the role of light exposure for the development of lateralization in domestic chick embryos revolutionized this research field. However, two main issues remain unresolved: (i) while in chicks anatomical light-dependent lateralization is mostly confined to the thalamofugal visual pathway, in pigeons only the tectofugal pathway is lateralized after light exposure. However, no study in either species ever investigated anatomical lateralization in the entopallium, the forebrain station of the tectofugal pathway. (ii) It is now known that lateralization can be observed also in dark-incubated chicks, both at the behavioural and at the Immediate Early Gene-expression level. We hypothesized that lateralization of the tectofugal system may underlie these light-independent effects. To investigate structural lateralization in the tectofugal pathway of dark-incubated chicks, we used parvalbumin (PV) as a marker of a sub population of entopallial neurons, quantifying PV-ir cell densities in the left and right entopallium. We found higher density in the right hemisphere, revealing for the first time anatomical lateralization in entopallium and confirming its potential role in supporting lateralized brain processing in dark-incubated birds. Results are discussed in relation to the possible functional role of PV-ir cells in inhibitory neural functions.

Sensitive periods for social development: Interactions between predisposed and learned mechanisms.

We analysed research that makes use of precocial species as animal models to describe the interaction of predisposed mechanisms and environmental factors in early learning, in particular for the development of social cognition. We also highlight the role of sensitive periods in this interaction, focusing on domestic chicks as one of the main animal models for this field. In the first section of the review, we focus on the emergence of early predispositions to attend to social partners. These attentional biases appear before any learning experience about social stimuli. However, non-specific experiences occurring during sensitive periods of the early post-natal life determine the emergence of these predisposed mechanisms for the detection of social partners. Social predispositions have an important role for the development learning-based social cognitive functions, showing the interdependence of predisposed and learned mechanisms in shaping social development. In the second part of the review we concentrate on the reciprocal interactions between filial imprinting and spontaneous (not learned) social predispositions. Reciprocal influences between these two sets of mechanisms ensure that, in the natural environment, filial imprinting will target appropriate social objects. Neural and physiological mechanisms regulating the sensitive periods for the emergence of social predispositions and for filial imprinting learning are also described.

Response of male and female domestic chicks to change in the number (quantity) of imprinting objects.

When facing two sets of imprinting objects of different numerousness, domestic chicks prefer to approach the larger one. Given that choice for familiar and novel stimuli in imprinting situations is known to be affected by the sex of the animals, we investigated how male and female domestic chicks divide the time spent in the proximity of a familiar versus an unfamiliar number of objects, and how animals interact (by pecking) with these objects. We confirmed that chicks discriminate among the different numerousnesses, but we also showed that females and males behave differently, depending on the degree of familiarity of the objects. When objects in the testing sets were all familiar, females equally explored both sets and pecked at all objects individually. Males instead selectively approached the familiar numerousness and pecked more at it. When both testing sets comprised familiar as well as novel objects, both males and females approached the larger numerousness of familiar objects. However, chicks directed all their pecks toward the novel object within the set. Differences in the behavior of males and females can be accounted for in terms of sex difference in the motivation to reinstate social contact with the familiar objects and to explore novel ones, likely associated with the ecology and the social structure of the species before domestication.

A leftward bias negatively correlated with performance is selectively displayed by domestic chicks during rule reversal (not acquisition).

In order to face a constantly changing environment, animals need to be able to update their knowledge of the world on the basis of new information. Often, this means to inhibit a previously acquired response and flexibly change their behaviour to produce a new response. Here, we measured such abilities in young domestic chicks, employing a Colour Reversal Learning Task. During the acquisition phase, 17 one-week-old male chicks had to learn to peck on one of two coloured boxes to obtain a food reward. After reaching criterion, chicks underwent a reversal phase in which the previously learned colour-reward contingency was swapped. As expected from the literature, chicks performed better in the acquisition phase with respect to the reversal phase. Results moreover highlighted the presence of a lateralized bias selectively during reversal: chicks performed better if the stimulus rewarded was located in the left hemispace (processed by the right hemisphere). Interestingly, the bias correlated with the individual difficulty, i.e., it was stronger in those chicks which needed more trials to complete the reversal session. The present study contributes evidence in support of behavioural flexibility in young chicks, along with a novel perspective on lateralized mechanisms that might underlie such ability.

Selective activation of the right hippocampus during navigation by spatial cues in domestic chicks (Gallus gallus).

In different vertebrate species, hippocampus plays a crucial role for spatial orientation. However, even though cognitive lateralization is widespread in the animal kingdom, the lateralization of this hippocampal function has been poorly studied. The aim of the present study was to investigate the lateralization of hippocampal activation in domestic chicks, during spatial navigation in relation to free-standing objects. Two groups of chicks were trained to find food in one of the feeders located in a large circular arena. Chicks of one group solved the task using the relational spatial information provided by free-standing objects present in the arena, while the other group used the local appearance of the baited feeder as a beacon. The immediate early gene product c-Fos was employed to map neural activation of hippocampus and medial striatum of both hemispheres. Chicks that used spatial cues for navigation showed higher activation of the right hippocampus compared to chicks that oriented by local features and compared to the left hippocampus. Such differences between the two groups were not present in the left hippocampus or in the medial striatum. Relational spatial information seems thus to be selectively processed by the right hippocampus in domestic chicks. The results are discussed in light of existing evidence of hippocampal lateralization of spatial processing in chicks, with particular attention to the contrasting evidence found in pigeons.

Numerical magnitude, rather than individual bias, explains spatial numerical association in newborn chicks.

We associate small numbers with the left and large numbers with the right side of space. Recent evidence from human newborns and non-human animals has challenged the primary role assigned to culture, in determining this spatial numerical association (SNA). Nevertheless, the effect of individual spatial biases has not been considered in previous research. Here, we tested the effect of numerical magnitude in SNA and we controlled for itablendividual biases. We trained 3-day-old chicks (Gallus gallus) on a given numerical magnitude (5). Then chicks could choose between two identical, left or right, stimuli both representing either 2, 8, or 5 elements. We computed the percentage of Left-sided Choice (LC). Numerical magnitude, but not individual lateral bias, explained LC: LC2 vs. 2>LC5 vs. 5>LC8 vs. 8. These findings suggest that SNA originates from pre-linguistic precursors, and pave the way to the investigation of the neural correlates of the number space association.

Most of the world modern-day languages are written from left to right ­ but what about numbers? As it turns out, the majority of people also represent numbers using a 'mental line', with smaller numbers on the left and larger numbers on the right. Some researchers argue that this phenomenon results from the way humans learn to read and write: in other words, that it is a by-product of culture, rather than an innate property of the brain. Recent evidence suggests that newborn infants, as well as certain species of monkeys and birds, also associate smaller numbers with the left and larger numbers with the right side of space. This raises the possibility that human mental number line may stem from an ability that evolved before language, in a common ancestor of humans and other animals. Yet, critics claim that findings in infants and non-human species result from a failure to account for individual biases in responding. To resolve this controversy, Rugani et al. trained three-day-old domestic chicks to approach a target board sporting five red squares. Chicks were then given the choice to approach two identical boards, which would both show two, five or eight red squares. Rugani et al. showed that when both boards had two red squares, the chicks tended to approach the left-hand board more often than the right. By contrast, when both boards had eight red squares, the birds approached the right-hand board more often than the left. Importantly, no left-right bias was observed when the number of red squared remained unchanged (five). These results also could not be explained by individual chicks favoring the left or right side. Instead, the findings suggest that even newborn animals tend to associate numbers with positions on a mental number line. Additional research is needed to determine the role of experience ­ or culture ­ in shaping this tendency, and future studies should also examine which brain regions support the association between number and space.

The use of spatial and local cues for orientation in domestic chicks (Gallus gallus).

Birds have been widely used to study spatial orientation. However, since different birds rely on different types of visual information to find goal locations (such as spatial information from free-standing objects or local cues, i.e. characteristics of a goal location like color and shape), it is important to investigate this aspect in each model species. The aim of the present study was to clarify whether domestic chicks, a ground-living bird and a widely used model for the comparative study of spatial orientation, are able to reorient in relation to free-standing objects and if they preferentially follow local or spatial cues. Furthermore, we also investigated whether monocular eye occlusion influences the ability of chicks to use spatial or local cues. Chicks were trained and tested in a large circular arena with free-standing objects providing relational spatial information, to find food in one of the feeders. We found that dark-incubated male chicks were able to reorient in relation to distinct, free-standing landmarks (Experiment 1), but when local and spatial cues were put in conflict, chicks significantly preferred local cues over spatial cues (Experiment 3). Moreover, while the use of one eye system only was not sufficient to orient by spatial cues (Experiment 2), the preference for local over spatial cues was independent of monocular occlusion (Experiment 4). The results are discussed in relation to our general knowledge of spatial information processing in domestic chicks.