Eiji Matsunaga

Differential cadherin expression in the developing postnatal telencephalon of a New World monkey.

Cadherins are cell adhesion molecules widely expressed in the nervous system, where they play various roles in neural patterning, nuclei formation, axon guidance, and synapse formation and function. Although many published articles have reported on cadherin expression in rodents and ferrets, there are limited data on their expression in primate brains. In this study, in situ hybridization analysis was performed for 10 cadherins [nine classic cadherins (Cdh4, ‐6, ‐7, ‐8, ‐9, ‐10, ‐11, ‐12, and ‐20) and T‐cadherin (Cdh13)] in the developing postnatal telencephalon of the common marmoset (Callithrix jacchus). Each cadherin showed broad expression in the cerebral cortex, basal ganglia, amygdala, and hippocampus, as previously shown in the rodent brain. However, detailed expression patterns differed between rodents and marmosets. In contrast to rodents, cadherin expression was reduced overall and localized to restricted areas of the brain during the developmental process, suggesting that cadherins are more crucially involved in developmental or maturation processes rather than in neural functioning. These results also highlight the possibility that restricted/less redundant cadherin expression allows primate brains to generate functional diversity among neurons, allowing morphological and functional differences between rodents and primates. J. Comp. Neurol. 521:4027–4060, 2013.

Dynamic Expression of Cadherins Regulates Vocal Development in a Songbird

Background Since, similarly to humans, songbirds learn their vocalization through imitation during their juvenile stage, they have often been used as model animals to study the mechanisms of human verbal learning. Numerous anatomical and physiological studies have suggested that songbirds have a neural network called ‘song system’ specialized for vocal learning and production in their brain. However, it still remains unknown what molecular mechanisms regulate their vocal development. It has been suggested that type-II cadherins are involved in synapse formation and function. Previously, we found that type-II cadherin expressions are switched in the robust nucleus of arcopallium from cadherin-7-positive to cadherin-6B-positive during the phase from sensory to sensorimotor learning stage in a songbird, the Bengalese finch. Furthermore, in vitro analysis using cultured rat hippocampal neurons revealed that cadherin-6B enhanced and cadherin-7 suppressed the frequency of miniature excitatory postsynaptic currents via regulating dendritic spine morphology. Methodology/Principal Findings To explore the role of cadherins in vocal development, we performed an in vivo behavioral analysis of cadherin function with lentiviral vectors. Overexpression of cadherin-7 in the juvenile and the adult stages resulted in severe defects in vocal production. In both cases, harmonic sounds typically seen in the adult Bengalese finch songs were particularly affected. Conclusions/Significance Our results suggest that cadherins control vocal production, particularly harmonic sounds, probably by modulating neuronal morphology of the RA nucleus. It appears that the switching of cadherin expressions from sensory to sensorimotor learning stage enhances vocal production ability to make various types of vocalization that is essential for sensorimotor learning in a trial and error manner.

Expression patterns of mineralocorticoid and glucocorticoid receptors in Bengalese finch (Lonchura striata var. domestica) brain suggest a relationship between stress hormones and song-system development.

Much evidence suggests that song traits function as an honest signal of male quality during mate choice in songbirds. Because songbirds learn vocalizations during the juvenile stage, development of the song system and song traits is affected by stressful conditions. However, it remains unknown how stressful conditions affect later song traits during development. To explore the relationship between glucocorticoids and song-system development, we performed in situ hybridization analysis of the glucocorticoid and mineralocorticoid receptors in juvenile and adult brains. The glucocorticoid receptor showed weak expression in song nuclei and strong expression in the hypothalamus, whereas the mineralocorticoid receptor showed strong song-nuclei-related expression. Thus, it appears that glucocorticoids are involved in song development directly by binding to receptors in song nuclei or indirectly by regulating sex hormones through hypothalamic hormones.

Evolution and diversity in avian vocal system: An Evo‐Devo model from the morphological and behavioral perspectives

Birds use various vocalizations to mark their territory and attract mates. Three groups of birds (songbirds, parrots, and hummingbirds) learn their vocalizations through imitation. In the brain of such vocal learners, there is a neural network called the song system specialized for vocal learning and production. In contrast, birds such as chickens and pigeons do not have such a neural network and can only produce innate sounds. Since each avian species shows distinct, genetically inherited vocal learning abilities that are related to its morphology, the avian vocal system is a good model for studying the evolution of functional neural circuits. Nevertheless, studies on avian vocalization from an evolutionary developmental‐biological (Evo‐Devo) perspective are scant. In the present review, we summarize the results of songbird studies and our recent work that used the Evo‐Devo approach to understand the evolution of the avian vocal system.

Vocal control area-related expression of neuropilin-1, plexin-A4, and the ligand semaphorin-3A has implications for the evolution of the avian vocal system.

The avian vocal system is a good model for exploring the molecular basis of neural circuit evolution related to behavioral diversity. Previously, we conducted a comparative gene expression analysis among two different families of vocal learner, the Bengalese finch (Lonchura striata var. domestica), a songbird, and the budgerigar (Melopsittacus undulatus), a parrot; and a non‐learner, the quail (Coturnix coturnix), to identify various axon guidance molecules such as cadherin and neuropilin‐1 as vocal control area‐related genes. Here, we continue with this study and examine the expression of neuropilin and related genes in these species in more detail. We found that neuropilin‐1 and its coreceptor, plexin‐A4, were expressed in several vocal control areas in both Bengalese finch and budgerigar brains. In addition, semaphorin‐3A, the ligand of neuropilin‐1, expression was not detected in vocal control areas in both species. Furthermore, there was some similar gene expression in the quail brain. These results suggest the possibility that a change in the expression of a combination of semaphorin/neuropilin/plexin was involved in the acquisition of vocal learning ability during evolution.

Comparative analysis of developmentally regulated expressions of Gadd45a, Gadd45b, and Gadd45g in the mouse and marmoset cerebral cortex

The cerebral cortex is an indispensable region that is involved in higher cognitive function in the mammalian brain, and is particularly evolved in the primate brain. It has been demonstrated that cortical areas are formed by both innate and activity-dependent mechanisms. However, it remains unknown what molecular changes induce cortical expansion and complexity during primate evolution. Active DNA methylation/demethylation is one of the epigenetic mechanisms that can modify gene expression via the methylation/demethylation of promoter regions. Three growth arrest and DNA damage-inducible small nuclear proteins, Gadd45 alpha, beta, and gamma, have been identified as regulators of methylation status. To understand the involvement of epigenetic factors in primate cortical evolution, we started by analyzing expression of these demethylation genes in the developing common marmoset (Callithrix jacchus) and mouse (Mus musculus) brain. In the marmoset brain, we found that cortical expression levels of Gadd45 alpha and gamma were reduced during development, whereas there was high expression of Gadd45 beta in some areas of the adult brain, including the prefrontal, temporal, posterior parietal and insula cortices, which are particularly expanded in greater primates and humans. Compared to the marmoset brain, there were no clear regional differences and constant or reduced Gadd45 expression was seen between juvenile and adult mouse brain. Double staining with a neuronal marker revealed that most Gadd45-expressing cells were NeuN-positive neurons. Thus, these results suggest the possibility that differential Gadd45 expression affects neurons, contributing cortical evolution and diversity.

Defects in ultrasonic vocalization of cadherin-6 knockout mice.

Background Although some molecules have been identified as responsible for human language disorders, there is still little information about what molecular mechanisms establish the faculty of human language. Since mice, like songbirds, produce complex ultrasonic vocalizations for intraspecific communication in several social contexts, they can be good mammalian models for studying the molecular basis of human language. Having found that cadherins are involved in the vocal development of the Bengalese finch, a songbird, we expected cadherins to also be involved in mouse vocalizations. Methodology/Principal Findings To examine whether similar molecular mechanisms underlie the vocalizations of songbirds and mammals, we categorized behavioral deficits including vocalization in cadherin-6 knockout mice. Comparing the ultrasonic vocalizations of cadherin-6 knockout mice with those of wild-type controls, we found that the peak frequency and variations of syllables were differed between the mutant and wild–type mice in both pup-isolation and adult-courtship contexts. Vocalizations during male-male aggression behavior, in contrast, did not differ between mutant and wild–type mice. Open-field tests revealed differences in locomotors activity in both heterozygote and homozygote animals and no difference in anxiety behavior. Conclusions/Significance Our results suggest that cadherin-6 plays essential roles in locomotor activity and ultrasonic vocalization. These findings also support the idea that different species share some of the molecular mechanisms underlying vocal behavior.

Comparative Gene Expression Analysis Among Vocal Learners (Bengalese Finch and Budgerigar) and Non-Learners (Quail and Ring Dove) Reveals Variable Cadherin Expressions in the Vocal System

Birds use various vocalizations to communicate with one another, and some are acquired through learning. So far, three families of birds (songbirds, parrots, and hummingbirds) have been identified as having vocal learning ability. Previously, we found that cadherins, a large family of cell-adhesion molecules, show vocal control-area-related expression in a songbird, the Bengalese finch. To investigate the molecular basis of evolution in avian species, we conducted comparative analysis of cadherin expressions in the vocal and other neural systems among vocal learners (Bengalese finch and budgerigar) and a non-learner (quail and ring dove). The gene expression analysis revealed that cadherin expressions were more variable in vocal and auditory areas compared to vocally unrelated areas such as the visual areas among these species. Thus, it appears that such diverse cadherin expressions might have been related to generating species diversity in vocal behavior during the evolution of avian vocal learning.

Expression pattern of cadherins in the naked mole rat (Heterocephalus glaber) suggests innate cortical diversification of the cerebrum

The cerebral cortex is an indispensable region for higher cognitive function that is remarkably diverse among mammalian species. Although previous research has shown that the cortical area map in the mammalian cerebral cortex is formed by innate and activity‐dependent mechanisms, it remains unknown how these mechanisms contribute to the evolution and diversification of the functional cortical areas in various species. The naked mole rat (Heterocephalus glaber) is a subterranean, eusocial rodent. Physiological and anatomical studies have revealed that the visual system is regressed and the somatosensory system is enlarged. To examine whether species differences in cortical area development are caused by intrinsic factors or environmental factors, we performed comparative gene expression analysis of neonatal naked mole rat and mouse brains. The expression domain of cadherin‐6, a somatosensory marker, was expanded caudally and shifted dorsally in the cortex, whereas the expression domain of cadherin‐8, a visual marker, was reduced caudally in the neonatal naked mole rat cortex. The expression domain of cadherin‐8 was also reduced in other visual areas, such as the lateral geniculate nucleus and superior colliculus. Immunohistochemical analysis of thalamocortical fibers further suggested that somatosensory input did not affect cortical gene expression in the neonatal naked mole rat brain. These results suggest that the development of the somatosensory system and the regression of the visual system in the naked mole rat cortex are due to intrinsic genetic mechanisms as well as sensory input‐dependent mechanisms. Intrinsic genetic mechanisms thus appear to contribute to species diversity in cortical area formation. J. Comp. Neurol. 519:1736‐1747, 2011.

Type-II cadherins modulate neural activity in cultured rat hippocampal neurons.

Cadherins, cell adhesion molecules widely expressed in the nervous system, are thought to be involved in synapse formation and function. To explore the role of cadherins in neuronal activity, we performed electrophysiological and morphological analyses of rat hippocampal cultured neurons overexpressing type-II cadherins, such as cadherin-6B and cadherin-7. We found that cadherin-6B increased but cadherin-7 decreased the number of protrusions of dendritic spines, and affected the frequency of miniature excitatory postsynaptic currents. Our results suggest that type-II cadherins may modulate neural activity by regulating neuronal morphology.