Frédéric Laberge

Evolution of the Amygdala: New Insights from Studies in Amphibians

The histology of amphibian brains gives an impression of relative simplicity when compared with that of reptiles or mammals. The amphibian telencephalon is small and contains comparatively few and large neurons, which in most parts constitute a dense periventricular cellular layer. However, the view emerging from the last decade is that the brains of all tetrapods, including amphibians, share a general bauplan resulting from common ancestry and the need to perform similar vital functions. To what extent this common organization also applies to higher brain functions is unknown due to a limited knowledge of the neurobiology of early vertebrates. The amygdala is widely recognized as a brain center critical for basic forms of emotional learning (e.g., fear conditioning) and its structure in amphibians could suggest how this capacity evolved. A functional systems approach is used here to synthesize the results of our anatomical investigations of the amphibian amygdala. It is proposed that the connectivity of the amphibian telencephalon portends a capacity for multi-modal association in a limbic system largely similar to that of amniote vertebrates. One remarkable exception is the presence of new sensory-associative regions of the amygdala in amniotes: the posterior dorsal ventricular ridge plus lateral nuclei in reptiles and the basolateral complex in mammals. These presumably homologous regions apparently are capable of modulating the phylogenetically older central amygdala and allow more complex forms of emotional learning.

Morphology and axonal projection pattern of neurons in the telencephalon of the fire-bellied toad Bombina orientalis: An anterograde, retrograde, and intracellular biocytin labeling study

The connectivity and cytoarchitecture of telencephalic centers except dorsal and medial pallium were studied in the fire‐bellied toad Bombina orientalis by anterograde and retrograde biocytin labeling and intracellular biocytin injection (total of 148 intracellularly labeled neurons or neuron clusters). Our findings suggest the following telencephalic divisions: (1) a central amygdala–bed nucleus of the stria terminalis in the caudal midventral telencephalon, connected to visceral–autonomic centers; (2) a vomeronasal amygdala in the caudolateral ventral telencephalon receiving input from the accessory olfactory bulb and projecting mainly to the preoptic region/hypothalamus; (3) an olfactory amygdala in the caudal pole of the telencephalon lateral to the vomeronasal amygdala receiving input from the main olfactory bulb and projecting to the hypothalamus; (4) a medial amygdala receiving input from the anterior dorsal thalamus and projecting to the medial pallium, septum, and hypothalamus; (5) a ventromedial column formed by a nucleus accumbens and a ventral pallidum projecting to the central amygdala, hypothalamus, and posterior tubercle; (6) a lateral column constituting the dorsal striatum proper rostrally and the dorsal pallidum caudally, and a ventrolateral column constituting the ventral striatum. We conclude that the caudal mediolateral complex consisting of the extended central, vomeronasal, and olfactory amygdala of anurans represents the ancestral condition of the amygdaloid complex. During the evolution of the mammalian telencephalon this complex was shifted medially and involuted. The mammalian basolateral amygdala apparently is an evolutionary new structure, but the medial portion of the amygdalar complex of anurans reveals similarities in input and output with this structure and may serve similar functions. J. Comp. Neurol. 478:35–61, 2004.

Connectivity and cytoarchitecture of the ventral telencephalon in the salamander Plethodon shermani

The cytoarchitecture and axonal connection pattern of centers in the ventral telencephalon of the salamander Plethodon shermani were studied using biocytin for anterograde and retrograde labeling of cell groups, as well as by intracellular injections. Application of biocytin to the main and accessory olfactory bulbs identified the olfactory pallial regions and the vomeronasal portion of the amygdala, respectively. According to our results, the amygdala of Plethodon is divided into (1) a rostral part projecting to visceral and limbic centers and receiving afferents from the dorsal thalamus, and (2) a caudal part receiving accessory olfactory input. The striatopallial transition area (SPTA) lies rostrodorsally to the caudal (vomeronasal) amygdala and is similar in connections and possibly in function. The rostral striatum has few descending projections to the medulla, whereas the intermediate striatum sends strong projections to the tegmentum and medulla. The caudal striatum has strong ascending projections to the striatum and descending projections to the ventral hypothalamus. The dendritic trees of neurons labeled below the striatum and in the SPTA spread laterally from the soma, whereas dendrites of striatal neurons converge into the laterally situated striatal neuropil. In the caudal amygdala, three distinct types of neurons are found differing in dendritic arborization. It is concluded that, hodologically, the rostral part of the urodele amygdala corresponds to the central and basolateral amygdala and the caudal part to the cortical/medial amygdala of mammals. The urodele striatum is divided into a rostral striatum proper, an intermediate dorsal pallidum, and a caudal part, with distinct connections described here for the first time in a vertebrate. J. Comp. Neurol. 482:176–200, 2005.

Organization of the sensory input to the telencephalon in the fire-bellied toad, Bombina orientalis

The functional organization of sensory activity in the amphibian telencephalon is poorly understood. We used an in vitro brain preparation to compare the anatomy of afferent pathways with the localization of electrically evoked sensory potentials and single neuron intracellular responses in the telencephalon of the toad Bombina orientalis. Anatomical tracing showed that the anterior thalamic nucleus innervates the anterior parts of the medial, dorsal, and lateral pallia and the rostralmost part of the pallium in addition to the subpallial amygdala/ventral pallidum region. Additional afferents to the medial telencephalon originate from the thalamic eminence. Electrical stimulation of diverse sensory nerves and brain regions generated evoked potentials with distinct characteristics in the pallium, subpallial amygdala/ventral pallidum, and dorsal striatopallidum. In the pallium, this sensory activity is generated in the anterior medial region. In the case of olfaction, evoked potentials were recorded at all sites, but displayed different characteristics across telencephalic regions. Stimulation of the anterior dorsal thalamus generated a pattern of activity comparable to olfactory evoked potentials, but it became similar to stimulation of the optic nerve or brainstem after bilateral lesion of the lateral olfactory tract, which interrupted the antidromic activation of the olfactohabenular tract. Intracellular bimodal sensory responses were obtained in the anterior pallium, medial amygdala, ventral pallidum, and dorsal striatopallidum. Our results demonstrate that the amphibian anterior pallium, medial amygdala/ventral pallidum, and dorsal striatopallidum are multimodal sensory centers. The organization of the amphibian telencephalon displays striking similarities with the brain pathways recently implicated in mammalian goal‐directed behavior. J. Comp. Neurol. 502:55–74, 2007.

Organization of the pallium in the fire‐bellied toad Bombina orientalis. I: Morphology and axonal projection pattern of neurons revealed by intracellular biocytin labeling

The cytoarchitecture and axonal projection pattern of pallial areas was studied in the fire‐bellied toad Bombina orientalis by intracellular injection of biocytin into a total of 326 neurons forming 204 clusters. Five pallial regions were identified, differing in morphology and projection pattern of neurons. The rostral pallium receiving the bulk of dorsal thalamic afferents has reciprocal connections with all other pallial areas and projects to the septum, nucleus accumbens, and anterior dorsal striatum. The medial pallium projects bilaterally to the medial pallium, septum, nucleus accumbens, mediocentral amygdala, and hypothalamus and ipsilaterally to the rostral, dorsal, and lateral pallium. The ventral part of the medial pallium is distinguished by efferents to the eminentia thalami and the absence of contralateral projections. The dorsal pallium has only ipsilateral projections running to the rostral, medial, and lateral pallium; septum; nucleus accumbens; and eminentia thalami. The lateral pallium has ipsilateral projections to the olfactory bulbs and to the rostral, medial, dorsal, and ventral pallium. The ventral pallium including the striatopallial transition area (SPTA) has ipsilateral projections to the olfactory bulbs, rostral and lateral pallium, dorsal striatopallidum, vomeronasal amygdala, and hypothalamus. The medial pallium can be tentatively homologized with the mammalian hippocampal formation, the dorsal pallium with allocortical areas, the lateral pallium rostrally with the piriform and caudally with the entorhinal cortex, the ventral pallium with the accessory olfactory amygdala. The rostral pallium, with its projections to the dorsal and ventral striatopallidum, resembles the mammalian frontal cortex. J. Comp. Neurol. 501:443–464, 2007.

Thalamo-telencephalic pathways in the fire-bellied toad Bombina orientalis.

It was suggested that among extant vertebrates, anuran amphibians display a brain organization closest to the ancestral tetrapod condition, and recent research suggests that anuran brains share important similarities with the brains of amniotes. The thalamus is the major source of sensory input to the telencephalon in both amphibians and amniote vertebrates, and this sensory input is critical for higher brain functions. The present study investigated the thalamo‐telencephalic pathways in the fire‐bellied toad Bombina orientalis, a basal anuran, by using a combination of retrograde tract tracing and intracellular injections with the tracer biocytin. Intracellular labeling revealed that the majority of neurons in the anterior and central thalamic nuclei project to multiple brain targets involved in behavioral modulation either through axon collaterals or en passant varicosities. Single anterior thalamic neurons target multiple regions in the forebrain and midbrain. Of note, these neurons display abundant projections to the medial amygdala and a variety of pallial areas, predominantly the anterior medial pallium. In Bombina, telencephalic projections of central thalamic neurons are restricted to the dorsal striato‐pallidum. The bed nucleus of the pallial commissure/thalamic eminence similarly targets multiple brain regions including the ventral medial pallium, but this is accomplished through a higher variety of distinct neuron types. We propose that the amphibian diencephalon exerts widespread influence in brain regions involved in behavioral modulation and that a single dorsal thalamic neuron is in a position to integrate different sensory channels and distribute the resulting information to multiple brain regions. J. Comp. Neurol. 508:806–823, 2008.

Food Web Structure Shapes the Morphology of Teleost Fish Brains

Previous work showed that teleost fish brain size correlates with the flexible exploitation of habitats and predation abilities in an aquatic food web. Since it is unclear how regional brain changes contribute to these relationships, we quantitatively examined the effects of common food web attributes on the size of five brain regions in teleost fish at both within-species (plasticity or natural variation) and between-species (evolution) scales. Our results indicate that brain morphology is influenced by habitat use and trophic position, but not by the degree of littoral-pelagic habitat coupling, despite the fact that the total brain size was previously shown to increase with habitat coupling in Lake Huron. Intriguingly, the results revealed two potential evolutionary trade-offs: (i) relative olfactory bulb size increased, while relative optic tectum size decreased, across a trophic position gradient, and (ii) the telencephalon was relatively larger in fish using more littoral-based carbon, while the cerebellum was relatively larger in fish using more pelagic-based carbon. Additionally, evidence for a within-species effect on the telencephalon was found, where it increased in size with trophic position. Collectively, these results suggest that food web structure has fundamentally contributed to the shaping of teleost brain morphology.

Cytoarchitecture of the accessory olfactory bulb in the salamander Plethodon shermani

Plethodontid terrestrial salamanders are emerging models in the study of the evolution of chemical communication in vertebrates. Their vomeronasal system is well defined. It comprises sensory neurons in the epithelium of the vomeronasal organ, whose axons form the vomeronasal nerve projecting to the accessory olfactory bulb (AOB), which in turn projects to the vomeronasal amygdala through the accessory olfactory tract. A detailed description of the cellular elements of the urodele AOB is lacking. Neuronal morphology in the AOB was studied by means of biocytin intracellular injections and retrograde tract tracing in the salamander Plethodon shermani. The AOB exhibits the characteristic lamination of olfactory bulbs, except that it displays a mixed periglomerular and mitral somata layer superficially. Mitral cells are the only AOB neurons projecting to the vomeronasal amygdala. Each mitral cell sends multiple axonal branches, generally through both dorsal and ventral portions of the accessory olfactory tract. Some mitral cells additionally send axon collaterals in the white matter immediately ventral to the AOB. AOB interneurons are divided into superficial periglomerular and deep granule cells, each category exhibiting morphological variety. Some neurons in the granule cell layer of the AOB or the region ventral to the AOB have dendritic trees that cover both regions. The present study is the first to highlight the full anatomical extent of single AOB neurons and surprisingly suggests that the ventrolateral telencephalon found below the AOB is part of the salamander vomeronasal system.

Efferent Axonal Projections of the Habenular Complex in the Fire-Bellied Toad Bombina orientalis

The habenular complex and its associated axonal pathways are often thought of as phylogenetically conserved features of the brain among vertebrates despite the fact that detailed studies of this brain region are limited to a few species. Here, the gross morphology and axonal projection pattern of the habenular complex of an anuran amphibian, the fire-bellied toad Bombina orientalis, was studied to allow comparison with the situation in other vertebrates. Axonal pathways were traced using biocytin applications in dissected brain preparations. The results show that the rostral part of the left dorsal nucleus is enlarged in this species, while the rostral ventral nucleus and caudal parts do not show left-right size differences. Biocytin applications revealed widespread axonal projections of the habenular complex to the posterior tuberculum/dorsal hypothalamic region, ventral tegmentum, interpeduncular nucleus (IPN), and raphe median. Additionally, axons targeting the lateral hypothalamus originated from the ventral habenular nuclei. The results also suggest an asymmetrical pattern of projection to the IPN in the rostral part of the habenular complex, where the left habenula preferentially targeted the dorsal IPN while the right habenula preferentially targeted the ventral IPN. The caudal habenular nuclei showed no asymmetry of projections as both sides targeted the ventral IPN. Comparison of the habenular complex axonal connectivity across vertebrates argues against strong phylogenetic conservation of the axonal projection patterns of different habenular nuclei.

Is there a structure equivalent to the mammalian basolateral amygdaloid complex in amphibians

To the Editor: RE: ‘Evolution of the amygdaloid complex in vertebrates, with special reference to the anamnio-amniotic transition’ by Moreno and Gonzalez. Journal of Anatomy, Vol. 211, pp. 151–163. Moreno and Gonzalez, in their review on the evolution of the amygdala, omitted to mention that their consideration of the amphibian ‘lateral’ amygdala as a multisensorial structure equivalent to the basolateral amygdaloid complex of mammals is controversial. We have proposed elsewhere that the absence of dorsal thalamic input to the lateral portions of the amygdala in amphibians precludes its role as a multisensory associative region, as is expected from parts of the basolateral complex (Laberge et al., 2006). In a study of sensory responses in the toad telencephalon, responses to sensory modalities other than olfaction were only found in the medial parts of the amygdala (Laberge & Roth, 2007). Our recent results further show that intracellularly labeled thalamic neurons do not send axon branches to the lateral parts of the amygdala in the toad Bombina orientalis (F. Laberge, S. Muhlenbrock-Lenter, U. Dicke and G. Roth unpublished data). Species differences could exist, but we feel that the assertion by Moreno and Gonzalez that the lateral amygdala represents a structure equivalent to the basolateral amygdaloid complex of mammals is not yet supported by experimental observations of thalamic input and multimodal sensory responses in this region.