Mammalian Amygdala Research Articles

Molecular Diversity of Neuron Types in the Salamander Amygdala and Implications for Amygdalar Evolution

The amygdala is a complex brain structure in the vertebrate telencephalon, essential for regulating social behaviors, emotions, and (social) cognition. In contrast to the vast majority of neuron types described in the many nuclei of the mammalian amygdala, little is known about the neuronal diversity in non-mammals, making reconstruction of its evolution particularly difficult. Here, we characterize glutamatergic neuron types in the amygdala of the urodele amphibian Pleurodeles waltl. Our single-cell RNA sequencing data indicate the existence of at least ten distinct types and subtypes of glutamatergic neurons in the salamander amygdala. These neuron types are molecularly distinct from neurons in the ventral pallium (VP), suggesting that the pallial amygdala and the VP are two separate areas in the telencephalon. In situ hybridization for marker genes indicates that amygdalar glutamatergic neuron types are located in three major subdivisions: the lateral amygdala, the medial amygdala, and a newly defined area demarcated by high expression of the transcription factor Sim1. The gene expression profiles of these neuron types suggest similarities with specific neurons in the sauropsid and mammalian amygdala. In particular, we identify Sim1+ and Sim1+ Otp+ expressing neuron types, potentially homologous to the mammalian nucleus of the lateral olfactory tract (NLOT) and to hypothalamic-derived neurons of the medial amygdala, respectively. Taken together, our results reveal a surprising diversity of glutamatergic neuron types in the amygdala of salamanders, despite the anatomical simplicity of their brain. These results offer new insights on the cellular and anatomical complexity of the amygdala in tetrapod ancestors.

Integrated Behavioral, Genetic and Brain Circuit Visualization Methods to Unravel Functional Anatomy of Zebrafish Amygdala.

The mammalian amygdala is a complex forebrain structure consisting of a heterogeneous group of nuclei derived from the pallial and subpallial telencephalon. It plays a critical role in a broad range of behaviors such as emotion, cognition, and social behavior; within the amygdala each nucleus has a distinct role in these behavioral processes. Topological, hodological, molecular, and functional studies suggest the presence of an amygdala-like structure in the zebrafish brain. It has been suggested that the pallial amygdala homolog corresponds to the medial zone of the dorsal telencephalon (Dm) and the subpallial amygdala homolog corresponds to the nuclei in the ventral telencephalon located close to and topographically basal to Dm. However, these brain regions are broad and understanding the functional anatomy of the zebrafish amygdala requires investigating the role of specific populations of neurons in brain function and behavior. In zebrafish, the highly efficient Tol2 transposon-mediated transgenesis method together with the targeted gene expression by the Gal4-UAS system has been a powerful tool in labeling, visualizing, and manipulating the function of specific cell types in the brain. The transgenic resource combined with neuronal activity imaging, optogenetics, pharmacology, and quantitative behavioral analyses enables functional analyses of neuronal circuits. Here, we review earlier studies focused on teleost amygdala anatomy and function and discuss how the transgenic resource and tools can help unravel the functional anatomy of the zebrafish amygdala.

Regional synapse gain and loss accompany memory formation in larval zebrafish

Defining the structural and functional changes in the nervous system underlying learning and memory represents a major challenge for modern neuroscience. Although changes in neuronal activity following memory formation have been studied [B. F. Grewe etal., Nature 543, 670-675 (2017); M. T. Rogan, U. V. Stäubli, J. E. LeDoux, Nature 390, 604-607 (1997)], the underlying structural changes at the synapse level remain poorly understood. Here, we capture synaptic changes in the midlarval zebrafish brain that occur during associative memory formation by imaging excitatory synapses labeled with recombinant probes using selective plane illumination microscopy. Imaging the same subjects before and after classical conditioning at single-synapse resolution provides an unbiased mapping of synaptic changes accompanying memory formation. In control animals and animals that failed to learn the task, there were no significant changes in the spatial patterns of synapses in the pallium, which contains the equivalent of the mammalian amygdala and is essential for associative learning in teleost fish [M. Portavella, J. P. Vargas, B. Torres, C. Salas, Brain Res. Bull 57, 397-399 (2002)]. In zebrafish that formed memories, we saw a dramatic increase in the number of synapses in the ventrolateral pallium, which contains neurons active during memory formation and retrieval. Concurrently, synapse loss predominated in the dorsomedial pallium. Surprisingly, we did not observe significant changes in the intensity of synaptic labeling, a proxy for synaptic strength, with memory formation in any region of the pallium. Our results suggest that memory formation due to classical conditioning is associated with reciprocal changes in synapse numbers in the pallium.

Development of the mouse anterior amygdalar radial unit marked by Lhx9-expression

The amygdala in mammals plays a key role in emotional processing and learning, being subdivided in pallial and subpallial derivatives. Recently, the cortical ring model and the pallial amygdalar radial model (Puelles et al. 2019; Garcia-Calero et al. 2020) described the pallial amygdala as an histogenetic field external to the allocortical ring, and subdivided it in five major radial domains called lateral, basal, anterior, posterior and retroendopiriform units. The anterior radial unit, whose cells typically express the Lhx9 gene (see molecular profile in Garcia-Calero et al. 2020), is located next to the pallial/subpallial boundary. This radial domain shows massive radial translocation and accumulation of its derivatives into its intermediate and superficial strata, with only a glial palisade representing its final periventricular domain. To better understand the development of this singular radial domain, not described previously, we followed the expression of Lhx9 during mouse amygdalar development in the context of the postulated radial subdivisions of the pallial amygdala and other telencephalic developmental features.

The dorsal arcopallium of chicks displays the expression of orthologs of mammalian fear related serotonin receptor subfamily genes

Fear is an adaptive emotion that elicits defensive behavioural responses against aversive threats in animals. In mammals, serotonin receptors (5-HTRs) have been shown to modulate fear-related neural circuits in the basolateral amygdala complex (BLA). To understand the phylogenetic continuity of the neural basis for fear, it is important to identify the neural circuit that processes fear in other animals. In birds, fear-related behaviours were suggested to be processed in the arcopallium/amygdala complex and modulated by the serotonin (5-HT) system. However, details about the distribution of 5-HTRs in the avian brain are very sparsely reported, and the 5-HTR that is potentially involved in fear-related behaviour has not been elucidated. In this study, we showed that orthologs of mammalian 5-HTR genes that are expressed in the BLA, namely 5-HTR1A, 5-HTR1B, 5-HTR2A, 5-HTR2C, 5-HTR3A, and 5-HTR4, are expressed in a part of the chick arcopallium/amygdala complex called the dorsal arcopallium. This suggests that serotonergic regulation in the dorsal arcopallium may play an important role in regulating fear-related behaviour in birds. Our findings can be used as a basis for comparing the processing of fear and its serotonergic modulation in the mammalian amygdala complex and avian arcopallium/amygdala complex.

The amygdala of the common shrew, guinea pig, rabbit, fox and pig: five flavours of the mammalian amygdala as a consequence of clade-specific mosaic-like evolution.

The amygdala is a part of neural networks that contribute to the regulation of emotional behaviours and emotional learning, stress response, and olfactory, pheromonal and reproductive functions. All these various functions are processed by the three main functional systems, frontotemporal, autonomic and olfactory, which are derived from different telencephalic sources (claustrum, striatum and olfactory cortex) and are represented, respectively, by the basolateral complex (BLC), the central complex (CC) and corticomedial complex (CMC) of the amygdala. The question arises of how these three functional systems evolved during mammalian phylogeny to fit the amygdala to specific needs of various animals. In the present study, we provide quantitative information regarding the individual volumes and neuron numbers in the BLC, CC and CMC of the common shrew, guinea pig, rabbit, fox and pig, a series of animals arranged according to increasing size and complexity of the brain. The results show that, in this series of animals, the BLC underwent a gradual size increase in volume and number of neurons, whereas the CMC was gradually reduced with regard to both these measures. The CC was more or less conserved across studied species. For example, the volume of the amygdala in pigs is ~250 times larger than that in shrews and it also has almost 26 times as many neurons as the amygdala of shrews. However, the volumes of the BLC, CC and CMC were ~380, 208 and 148 times larger, respectively, in pigs than in shrews. The number of neurons in these three regions was ~38, 23 and 20 times greater, respectively, in pigs than in shrews. The results also show striking morphometric similarities of the amygdala in the guinea pig and rabbit as well as fox and pig. For example, the percentages of neurons in the fox and pig are 42.23% and 42.78%, respectively, for the BLC, 16.64% and 16.58%, respectively, for the CC, and 41.12% and 40.64%, respectively, for the CMC. In conclusion, our results indicate that the amygdala does not evolve as a single unit but, instead, the three main functional systems evolved independently, which suggests that brain structures with major functional links evolve together independently of evolutionary changes in other unrelated structures. The size progression of the BLC parallels the size progression of the neocortex with which it is strongly functionally linked, whereas the CMC is strongly connected to olfactory regions, and all these structures follow the same regression course. Remarkable morphometric similarity of the amygdala in the guinea pig and rabbit as well as in the fox and pig, however, suggest that there must also be another mechanism shaping the morphology of the amygdala and the brain during evolution. The gradual nature of size changes in the BLC and CMC support this hypothesis as well.

Neurobiology of Wild and Hatchery-Reared Atlantic Salmon: How Nurture Drives Neuroplasticity.

Life experiences in the rearing environment shape the neural and behavioral plasticity of animals. In fish stocking practices, the hatchery environment is relatively stimulus-deprived and does not optimally prepare fish for release into the wild. While the behavioral differences between wild and hatchery-reared fish have been examined to some extent, few studies have compared neurobiological characteristics between wild and hatchery-reared individuals. Here, we compare the expression of immediate early gene cfos and neuroplasticity marker brain-derived neurotrophic factor (bdnf) in telencephalic subregions associated with processing of stimuli in wild and hatchery-reared Atlantic salmon at basal and 30 min post (acute) stress conditions. Using in situ hybridization, we found that the expression level of these markers is highly specific per neuronal region and affected by both the origin of the fish, and exposure to acute stress. Expression of cfos was increased by stress in all brain regions and cfos was more highly expressed in the Dlv (functional equivalent to the mammalian hippocampus) of hatchery-reared compared to wild fish. Expression of bdnf was higher overall in hatchery fish, while acute stress upregulated bdnf in the Dm (functional equivalent to the mammalian amygdala) of wild, but not hatchery individuals. Our findings demonstrate that the hatchery environment affects neuroplasticity and neural activation in brain regions that are important for learning processes and stress reactivity, providing a neuronal foundation for the behavioral differences observed between wild and hatchery-reared fish.

Neural and behavioural correlates of repeated social defeat

Dominance hierarchies are common across the animal kingdom and have important consequences for reproduction and survival. Animals of lower social status cope with repeated social defeat using proactive and reactive behaviours. However, there remains a paucity of information on how an individual’s coping behaviours changes over time or what neural mechanisms are involved. We used a resident-intruder paradigm in the African cichlid fish Astatotilapia burtoni to investigate the neural correlates of these two opposing behaviour groups. Fish initially used both proactive and reactive behaviours, but had a dramatic increase in use of proactive behaviours during the third interaction, and this was followed by cessation of proactive behaviours and exclusive use of reactive coping. By quantifying neural activation in socially-relevant brain regions, we identify a subset of brain nuclei, including those homologous to the mammalian amygdala, showing higher activation in fish displaying proactive but not reactive behaviours. Fish displaying reactive behaviours had greater neural activation in the superior raphe, suggesting a possible conserved function during social defeat across vertebrates. These data provide the first evidence on the involvement of specific brain regions underlying proactive and reactive coping in fishes, indicating that these nuclei have conserved functions during social defeat across taxa.

Identification of a neuronal population in the telencephalon essential for fear conditioning in zebrafish

BackgroundFear conditioning is a form of learning essential for animal survival and used as a behavioral paradigm to study the mechanisms of learning and memory. In mammals, the amygdala plays a crucial role in fear conditioning. In teleost, the medial zone of the dorsal telencephalon (Dm) has been postulated to be a homolog of the mammalian amygdala by anatomical and ablation studies, showing a role in conditioned avoidance response. However, the neuronal populations required for a conditioned avoidance response via the Dm have not been functionally or genetically defined.ResultsWe aimed to identify the neuronal population essential for fear conditioning through a genetic approach in zebrafish. First, we performed large-scale gene trap and enhancer trap screens, and created transgenic fish lines that expressed Gal4FF, an engineered version of the Gal4 transcription activator, in specific regions in the brain. We then crossed these Gal4FF-expressing fish with the effector line carrying the botulinum neurotoxin gene downstream of the Gal4 binding sequence UAS, and analyzed the double transgenic fish for active avoidance fear conditioning. We identified 16 transgenic lines with Gal4FF expression in various brain areas showing reduced performance in avoidance responses. Two of them had Gal4 expression in populations of neurons located in subregions of the Dm, which we named 120A-Dm neurons. Inhibition of the 120A-Dm neurons also caused reduced performance in Pavlovian fear conditioning. The 120A-Dm neurons were mostly glutamatergic and had projections to other brain regions, including the hypothalamus and ventral telencephalon.ConclusionsHerein, we identified a subpopulation of neurons in the zebrafish Dm essential for fear conditioning. We propose that these are functional equivalents of neurons in the mammalian pallial amygdala, mediating the conditioned stimulus–unconditioned stimulus association. Thus, the study establishes a basis for understanding the evolutionary conservation and diversification of functional neural circuits mediating fear conditioning in vertebrates.

Transmitter receptors reveal segregation of the arcopallium/amygdala complex in pigeons (Columba livia).

At the beginning of the 20th century it was suggested that a complex group of nuclei in the avian posterior ventral telencephalon is comparable to the mammalian amygdala. Subsequent findings, however, revealed that most of these structures share premotor characteristics, while some indeed constitute the avian amygdala. These developments resulted in 2004 in a change of nomenclature of these nuclei, which from then on were named arcopallial or amygdala nuclei and referred to as the arcopallium/amygdala complex. The structural basis for the similarities between avian and mammalian arcopallial and amygdala subregions is poorly understood. Therefore, we analyzed binding site densities for glutamatergic AMPA, NMDA and kainate, GABAergic GABAA , muscarinic M1 , M2 and nicotinic acetylcholine (nACh; α4 β2 subtype), noradrenergic α1 and α2 , serotonergic 5-HT1A and dopaminergic D1/5 receptors using quantitative in vitro receptor autoradiography combined with a detailed analysis of the cyto- and myelo-architecture. Our approach supports a segregation of the pigeon's arcopallium/amygdala complex into the following subregions: the arcopallium anterius (AA), the arcopallium ventrale (AV), the arcopallium dorsale (AD), the arcopallium intermedium (AI), the arcopallium mediale (AM), the arcopallium posterius (AP), the nucleus posterioris amygdalopallii pars basalis (PoAb) and pars compacta (PoAc), the nucleus taeniae amgygdalae (TnA) and the area subpallialis amygdalae (SpA). Some of these subregions showed further subnuclei and each region of the arcopallium/amygdala complex are characterized by a distinct multi-receptor density expression. Here we provide a new detailed map of the pigeon's arcopallium/amygdala complex and compare the receptor architecture of the subregions to their possible mammalian counterparts.

The Cytoarchitecture of the Telencephalon of Betta Splendens Regan 1910 (Perciformes: Anabantoidei) with a Stereological Approach to the Supracommissural and Postcommissural Nuclei.

Teleostean fish brains are useful models to study cellular and functional specializations along the phylogenesis. The Betta splendens Regan 1910 (Siamese fighting fish; Perciformes:Anabantoidei) is known for its aggressive display, courtship behavior, nest building, and offspring care. Here, we present novel and detailed data about the cytoarchitecture of the olfactory bulb and the telencephalic hemispheres of this fish. The hematoxylin-eosin and Nissl techniques served to identify brain nuclei (n = 19 males and n = 21 females) and for the stereological evaluation of the numerical density of cells and the proportion of neurons and glial cells in the ventral telencephalon supracommissural (Vs) and postcommissural (Vp) nuclei of adult males and females. These nuclei are putative homologs of the sexually dimorphic medial amygdala in mammals. The olfactory bulb of Betta splendens consists of 5 concentrically arranged layers plus ganglion cells of the terminal nerves. The dorsal telencephalon consists of 16 different cell groups. The ventral telencephalon has 8 nuclei, plus the lateral septal organ and the nuclei of the preoptic area forming an anatomical continuum. The rostrocaudal extent of the Vs and Vp is not different between sexes. In both nuclei, the proportion of neurons to glial cells is approximately 2:1 and the density of neurons and glial cells is not different between sexes. These morphological findings can subserve future research on the brain function of the Betta splendens and the search for neural sex differences in other central areas of this same species, in other teleost species, or yet in other related vertebrate group. Anat Rec, 00:000-000, 2017. © 2017 Wiley Periodicals, Inc. Anat Rec, 301:88-110, 2018. © 2017 Wiley Periodicals, Inc.

Distribution of Calretinin Immunoreactivity in the Lateral Nucleus of the Bottlenose Dolphin (Tursiops truncatus) Amygdala.

The amgdaloid complex consists of different nuclei, each with unique cytoarchitectonic, chemoarchitectonic and connectional characteristics. Most of the inputs coming from cortical and subcortical areas enter the amygdala via the lateral nucleus, which makes it the main receiving structure of the complex. The activity of its neurons is coordinated and modulated by different inhibitory, GABAergic-interneurons, which can be classified for their expression of various calcium-binding proteins, as well as by morphological characteristics. This research based on the analysis of the amygdala of three bottlenose dolphins, provides the first description of the topography, cytoarchitecture and distribution of calretinin immunoreactivity of the lateral nucleus. Our observations on the bottlenose dolphin confirmed the general topography of the mammalian amygdala and of the lateral nucleus. Notably, we identified six subdivision of the nucleus, more than those reported until now in the rat, monkey and human lateral nucleus. This could reveal an outstanding capability of integration and elaboration of external stimuli. In addition, we observed a strong presence of CR-immunoreactive (-ir) neurons and fibres. CR-ir neurons were mainly non-pyramidal inhibitory neurons; in particular, 80% of IR-cells were represented by large and small polygonal neurons. In the lateral nucleus of the human amygdala, CR-ir neurons form inhibitory synapses on calbindin-D28k-IR inhibitory interneurons. Since calbindin-D28k-ir interneurons make inhibitory synapses on the pyramidal cells, the final goal of the CR-ir interneurons could be the synchronization of cells activity, thus playing an important role in the control of information flow in the lateral amygdalar nucleus. Anat Rec, 2017. © 2017 Wiley Periodicals, Inc. Anat Rec, 300:2008-2016, 2017. © 2017 Wiley Periodicals, Inc.

Selective contribution of the telencephalic arcopallium to the social facilitation of foraging efforts in the domestic chick.

To investigate the neural basis of socio-economic behaviors in birds, we examined the effects of bilateral electrolytic lesions of arcopallium (Arco, the major descending pallial area of the avian telencephalon) and the surrounding nuclei in domestic chicks. We tested foraging effort (running distance) in an I-shaped maze with two food patches that delivered food in a biased manner according to a variable interval schedule. Normally, chicks run back and forth between the patches, and the patch use time matches the respective food delivery rate. In the paired phase, even without actual interference of food, chicks showed social facilitation of running effort compared with the single phase. Chicks with lesions in the Arco and lateral Arco showed significant reductions in social facilitation. The lesion effects of the lateral Arco were particularly selective, as it was not accompanied by changes in running distance in the single phase. Lesions of the nidopallium and nucleus taeniae of the amygdala produced no changes in foraging behavior. On the other hand, the Arco lesion did not impair social facilitation of operant peck latency. In accordance with this, anterograde tracing revealed characteristic projections from the lateral Arco to the extended amygdala, hippocampus, and septum, as well as wide areas of limbic nuclei in the hypothalamus and medial areas of the striatum including the nucleus accumbens. Pathways from the lateral Arco could enable chicks to overcome the extra effort investment of social foraging, suggesting functional and anatomical analogies to the anterior cingulate cortex and basolateral amygdala in mammals.

Potential of zebrafish as a model for exploring the role of the amygdala in emotional memory and motivational behavior.

Emotion is a key aspect of behavior, enabling humans and animals to assign either positive or negative values to sensory inputs and thereby to make appropriate decisions. Classical experiments in mammalian models, mainly in primates and rodents, have shown that the amygdala is essential for appetitive and aversive associative processing and that dysfunction of this brain region leads to various psychiatric conditions, including depression, generalized anxiety disorder, panic disorder, phobias, autism, and posttraumatic stress disorder. In the past 2 decades, the zebrafish (Danio rerio; Cyprinidae) has emerged as a versatile, reliable vertebrate model organism for the in vivo study of development, gene function, and numerous aspects of human pathologies. Small size, high fecundity, rapid external development, transparency, genetic tractability, and high genetic and physiologic homology with humans are among the factors that have contributed to the success with this small fish in different biomedical research areas. Recent findings indicate that, despite the anatomical differences in the brain structure of teleosts and tetrapods, fish possess a structure homologous to the mammalian amygdala, a hypothesis that is supported by the expression of molecular markers, analyses of neuronal projections in different brain areas, and behavioral studies. This Review summarizes this evidence and highlights a number of relevant bioassays in zebrafish to study emotional memory and motivational behavior.

Leaving the door open for fish pain: Evolutionary convergence and the utility of ‘just-so stories’

Leaving the door open for fish pain: Evolutionary convergence and the utility of ‘just-so stories’

Neural responses to songbird vocalizations in the nucleus taeniae of the amygdala in Bengalese finches

Emotions are important psychological processes that induce adaptive behavior in response to communication signals. Many species of songbirds form complex social relationships and communicate with others through a large variety of vocal sounds. Thus, songbirds could be a good model to search for the neural basis of emotion especially in a context of communication. Anatomical studies have shown that the nucleus taeniae of the amygdala (TnA) in birds corresponds to the medial amygdala in mammals. While the amygdala is suggested to be involved in recognition of conspecific vocalizations in rats and bats, the function of the TnA still remains unclear. The present study aimed to explore auditory response properties of the songbird TnA by electrophysiology. We examined activity of the Bengalese finch TnA neurons during the presentations of conspecific and heterospecific vocalizations, as well as synthesized sound. We demonstrated for the first time that a population of TnA neurons exhibited selective auditory responses to songbird vocalizations. Our findings suggested involvement of the songbirds TnA for the recognition of communicative sounds. Further investigation into the TnA response properties should be fruitful in understanding the relationship between emotion and vocal signals in animals. [Work supported by JSPS KAKENHI Grant # 26240019.]

Microbes & neurodevelopment--Absence of microbiota during early life increases activity-related transcriptional pathways in the amygdala.

The mammalian amygdala is a key emotional brain region for eliciting social behaviour, critically involved in anxiety and fear-related behaviours, and hence a focus of research on neurodevelopmental and stress-related disorders such as autism and anxiety. Recently, increasing evidence implicates host-microbe interactions in the aetiology of these conditions. Germ-free (GF) mice, devoid of any microbiota throughout organismal maturation, are a well-established tool to study the effects of absence of the microbiota on host physiology. A growing body of independently replicated findings confirm that GF animals demonstrate altered anxiety-related behaviour and impaired social behaviour. However, the underlying mechanisms of this interaction and the nature of the pathways involved are only insufficiently understood. To further elucidate the molecular underpinnings of microbe-brain interaction, we therefore exploited unbiased genome-wide transcriptional profiling to determine gene expression in the amygdala of GF and GF mice that have been colonised after weaning. Using RNA-sequencing and a comprehensive downstream analysis pipeline we studied the amygdala transcriptome and found significant differences at the levels of differential gene expression, exon usage and RNA-editing. Most surprisingly, we noticed upregulation of several immediate early response genes such as Fos, Fosb, Egr2 or Nr4a1 in association with increased CREB signalling in GF mice. In addition, we found differential expression and recoding of several genes implicated in brain physiology processes such as neurotransmission, neuronal plasticity, metabolism and morphology. In conclusion, our data suggest altered baseline neuronal activity in the amygdala of germ-free animals, which is established during early life and may have implications for understanding development and treatment of neurodevelopmental disorders.

The endocannabinoid system and associative learning and memory in zebrafish

In zebrafish the medial pallium of the dorsal telencephalon represents an amygdala homolog structure, which is crucially involved in emotional associative learning and memory. Similar to the mammalian amygdala, the medial pallium contains a high density of endocannabinoid receptor CB1. To elucidate the role of the zebrafish endocannabinoid system in associative learning, we tested the influence of acute and chronic administration of receptor agonists (THC, WIN55,212-2) and antagonists (Rimonabant, AM-281) on two different learning paradigms. In an appetitively motivated two-alternative choice paradigm, animals learned to associate a certain color with a food reward. In a second set-up, a fish shuttle-box, animals associated the onset of a light stimulus with the occurrence of a subsequent electric shock (avoidance conditioning). Once fish successfully had learned to solve these behavioral tasks, acute receptor activation or inactivation had no effect on memory retrieval, suggesting that established associative memories were stable and not alterable by the endocannabinoid system. In both learning tasks, chronic treatment with receptor antagonists improved acquisition learning, and additionally facilitated reversal learning during color discrimination. In contrast, chronic CB1 activation prevented aversively motivated acquisition learning, while different effects were found on appetitively motivated acquisition learning. While THC significantly improved behavioral performance, WIN55,212-2 significantly impaired color association. Our findings suggest that the zebrafish endocannabinoid system can modulate associative learning and memory. Stimulation of the CB1 receptor might play a more specific role in acquisition and storage of aversive learning and memory, while CB1 blocking induces general enhancement of cognitive functions.

The α2-subunit of the nicotinic cholinergic receptor is specifically expressed in medial subpallium-derived cells of mammalian amygdala.

Nicotinic acetylcholine receptor (nAChR) subtypes are expressed in specific neuronal populations, which are involved in numerous neural functions such as sleep, fatigue, anxiety, and cognition, as well as the central processing of pain and food intake. Moreover, mutations in nAChRs subunits have been related to frontal lobe epilepsy, neurodegenerative diseases, and other neurological disorders, including schizophrenia and attention deficit and hyperactivity disorder (ADHD). Previous studies have shown that the α2-subunit of the AChR (Chrna2) is expressed in the basal forebrain, in the septum, and in some amygdalar nuclei in the adult rodent brain. However, although the importance of this amygdalar expression in emotion-related behavior and the physiopathology of neuropsychiatric disorders has been accepted, a detailed study of the Chrna2 expression pattern during development has been lacking. In this study we found that Chrna2 is specifically expressed in medial subpallium-derived amygdalar nuclei from early developmental stages to adult. This finding could help us to better understand the role of Chrna2 in the differentiation and functional maturation of amygdalar neurons involved in cholinergic-regulated emotional behavior.

Emotions and motivated behavior converge on an amygdala-like structure in the zebrafish.

The brain reward circuitry plays a key role in emotional and motivational behaviors, and its dysfunction underlies neuropsychiatric disorders such as schizophrenia, depression and drug addiction. Here, we characterized the neuronal activity pattern induced by acute amphetamine administration and during drug-seeking behavior in the zebrafish, and demonstrate the existence of conserved underlying brain circuitry. Combining quantitative analyses of cfos expression with neuronal subtype-specific markers at single-cell resolution, we show that acute d-amphetamine administration leads to both increased neuronal activation and the recruitment of neurons in the medial (Dm) and the lateral (Dl) domains of the adult zebrafish pallium, which contain homologous structures to the mammalian amygdala and hippocampus, respectively. Calbindin-positive and glutamatergic neurons are recruited in Dm, and glutamatergic and γ-aminobutyric acid (GABAergic) neurons in Dl. The drug-activated neurons in Dm and Dl are born at juvenile stage rather than in the embryo or during adulthood. Furthermore, the same territory in Dm is activated during both drug-seeking approach and light avoidance behavior, while these behaviors do not elicit activation in Dl. These data identify the pallial territories involved in acute psychostimulant response and reward formation in the adult zebrafish. They further suggest an evolutionarily conserved function of amygdala-like structures in positive emotions and motivated behavior in zebrafish and mammals.