Microglial activation and pyroptosis induced by nano-TiO2 in marine medaka brain.

Recently, nano-titanium dioxide (nano-TiO2) has been widely distributed over surface water. However, there are few reports on its effects on the central nervous system of fish. In this study, we investigated whether nano-TiO2 enters the medaka brain after exposure and its effect on the brain. Marine medaka brains were examined after exposure to 0.01 g/L nano-TiO2 for 3, 10, and 20 d. Nano-TiO2-like particles were found in the telencephalon of treated fish. There was no obvious brain histopathological injury. The number of irregular mitochondria with absent cristae increased. Gene expression of the apoptosis-related genes, casp8, bcl2b, and bax, decreased significantly in the nano-TiO2 group at 3 d. In contrast, the pyroptosis-related genes, gsdmeb and casp1, and inflammation-related factor, il18, increased significantly. As an activated microglia marker, mRNA expression of cd68 increased significantly in the nano-TiO2 treated group. Moreover, CD68 protein expression also increased significantly at 10 d. Altogether, we show that nano-TiO2 can alter mitochondrial morphology in the telencephalon of medaka, leading to microglial activation and pyroptosis.

Evolution of Telencephalon Anterior-Posterior Patterning through Core Endogenous Network Bifurcation.

How did the complex structure of the telencephalon evolve? Existing explanations are based on phenomena and lack a first-principles account. The Darwinian dynamics and endogenous network theory-established decades ago-provides a mathematical and theoretical framework and a general constitutive structure for theory-experiment coupling for answering this question from a first-principles perspective. By revisiting a gene network that explains the anterior-posterior patterning of the vertebrate telencephalon, we found that upon increasing the cooperative effect within this network, fixed points gradually evolve, accompanied by the occurrence of two bifurcations. The dynamic behavior of this network is informed by the knowledge obtained from experiments on telencephalic evolution. Our work provides a quantitative explanation for how telencephalon anterior-posterior patterning evolved from the pre-vertebrate chordate to the vertebrate and provides a series of verifiable predictions from a first-principles perspective.

Adult sex change leads to extensive forebrain reorganization in clownfish.

BACKGROUND: Sexual differentiation of the brain occurs in all major vertebrate lineages but is not well understood at a molecular and cellular level. Unlike most vertebrates, sex-changing fishes have the remarkable ability to change reproductive sex during adulthood in response to social stimuli, offering a unique opportunity to understand mechanisms by which the nervous system can initiate and coordinate sexual differentiation. METHODS: This study explores sexual differentiation of the forebrain using single nucleus RNA-sequencing in the anemonefish Amphiprion ocellaris, producing the first cellular atlas of a sex-changing brain. RESULTS: We uncover extensive sex differences in cell type-specific gene expression, relative proportions of cells, baseline neuronal excitation, and predicted inter-neuronal communication. Additionally, we identify the cholecystokinin, galanin, and estrogen systems as central molecular axes of sexual differentiation. Supported by these findings, we propose a model of sexual differentiation in the conserved vertebrate social decision-making network spanning multiple subtypes of neurons and glia, including neuronal subpopulations within the preoptic area that are positioned to regulate gonadal differentiation. CONCLUSIONS: This work deepens our understanding of sexual differentiation in the vertebrate brain and defines a rich suite of molecular and cellular pathways that differentiate during adult sex change in anemonefish.

This study provides key insights into brain sex differences in sex-changing anemonefish (Amphiprion ocellaris), a species that changes sex in adulthood in response to the social environment. Using single nucleus RNA-sequencing, the study provides the first brain cellular atlas showing sex differences in two crucial reproductive areas: the preoptic area and telencephalon. The research identifies notable sex-differences in cell-type proportions and gene expression, particularly in radial glia and glutamatergic neurons that co-express the neuropeptide cholecystokinin. It also highlights differences in preoptic area neurons likely involved in gonadal regulation. This work deepens our understanding of sexual differentiation of the brain in vertebrates, especially those capable of adult sex change, and illuminates key molecular and cellular beginning and endpoints of the process.

Single-cell sequencing unveils the impact of aging on the progenitor cell diversity in the telencephalon of the female killifish N. furzeri.

The African turquoise killifish (Nothobranchius furzeri) combines a short lifespan with spontaneous age-associated loss of neuro-regenerative capacity, an intriguing trait atypical for a teleost. The impact of aging on the cellular composition of the adult stem cell niches, leading to this dramatic decline in the postnatal neuro- and gliogenesis, remains elusive. Single-cell RNA sequencing of the telencephalon of young adult female killifish of the short-lived GRZ-AD strain unveiled progenitors of glial and non-glial nature, different excitatory and inhibitory neuron subtypes, as well as non-neural cell types. Sub-clustering of the progenitors identified four radial glia (RG) cell types, two non-glial progenitor (NGP) and four intermediate (intercell) cell states. Two astroglia-like, one ependymal, and one neuroepithelial-like (NE) RG subtype were found at different locations in the forebrain in line with their role, while proliferative, active NGPs were spread throughout. Lineage inference pointed to NE-RG and NGPs as start and intercessor populations for glio- and neurogenesis. Upon aging, single-cell RNA sequencing revealed major perturbations in the proportions of the astroglia and intercell states, and in the molecular signatures of specific subtypes, including altered MAPK, mTOR, Notch, and Wnt pathways. This cell catalog of the young regeneration-competent killifish telencephalon, combined with the evidence for aging-related transcriptomic changes, presents a useful resource to understand the molecular basis of age-dependent neuroplasticity. This data is also available through an online database (killifishbrain_scseq).

Soft-tissue fossilization illuminates the stepwise evolution of the ray-finned fish brain.

A complex brain is central to the success of backboned animals. However, direct evidence bearing on vertebrate brain evolution comes almost exclusively from extant species, leaving substantial knowledge gaps. Although rare, soft-tissue preservation in fossils can yield unique insights on patterns of neuroanatomical evolution. Paleontological evidence from an exceptionally preserved Pennsylvanian (∼318 Ma) actinopterygian, Coccocephalus, calls into question prior interpretations of ancestral actinopterygian brain conditions. However, the ordering and timing of major evolutionary innovations, such as an everted telencephalon, modified meningeal tissues, and hypothalamic inferior lobes, remain unclear. Here, we report two distinct actinopterygian morphotypes from the latest Carboniferous-earliest Permian (∼299 Ma) of Brazil that show extensive soft-tissue preservation of brains, cranial nerves, eyes, and potential cardiovascular tissues. These fossils corroborate inferences drawn from ✝Coccocephalus, while adding new information about neuroanatomical evolution. Skeletal features indicate that one of these Brazilian morphotypes is more closely related to living actinopterygians than the other, which is also reflected in soft-tissue features. Significantly, the more crownward morphotype shows a key neuroanatomical feature of extant actinopterygians-an everted telencephalon-that is absent in the other morphotype and ✝Coccocephalus. All preserved Paleozoic actinopterygian brains show broad similarities, including an invaginated cerebellum, hypothalamus inferior lobes, and a small forebrain. In each case, preserved brains are substantially smaller than the enclosing cranial chamber. The neuroanatomical similarities shared by this grade of Permo-Carboniferous actinopterygians reflect probable primitive conditions for actinopterygians, providing a revised model for interpreting brain evolution in a major branch of the vertebrate tree of life.

Atypical telencephalic reflux pattern in a cavernous sinus dural AV fistula related to an anatomical variation of the basal vein of Rosenthal.

Cavernous sinus dural arteriovenous fistula can cause cerebral edema and hemorrhage due to cortical venous reflux and congestion. Understanding complex venous reflux and drainage routes is crucial for treatment planning. Here, we present a case of a cavernous sinus dural arteriovenous fistula with cortical venous reflux via two separate terminations of the telencephalic veins caused by an aplastic basal vein of Rosenthal. The patient presented with diplopia and eye redness and was diagnosed with a Cognard type IIa + b cavernous sinus dural arteriovenous fistula. The shunt was supplied by the dural branches of the internal and external carotid arteries. Multiple shunt points involving the intercavernous sinus and the medial aspect of the left cavernous sinus were identified, with drainage into the supraorbital and intracranial veins, including two separate terminations of the telencephalic veins, one leading to the laterocavernous sinus via the superficial middle cerebral vein and the other to the cavernous sinus via the uncal vein, resulting in basal ganglia venous congestion in the absence of the basal vein of Rosenthal. During transvenous embolization, the intracranial veins, cavernous sinus, and intercavernous sinus were obliterated using a double-catheter technique with a combination of coils and liquid embolics. Telencephalic venous variations can lead to cavernous sinus drainage into the basal ganglia and orbitofrontal brain. This unique drainage pattern underscores the importance of recognizing anatomical variations when managing cavernous sinus dural arteriovenous fistula.

Variations in touch representation in the hummingbird and zebra finch forebrain.

Somatosensation is essential for animals to perceive the external world through touch, allowing them to detect physical contact, temperature, pain, and body position. Studies on rodent vibrissae have highlighted the organization and processing in mammalian somatosensory pathways.1,2 Comparative research across vertebrates is vital for understanding evolutionary influences and ecological specialization on somatosensory systems. Birds, with their diverse morphologies, sensory abilities, and behaviors, serve as ideal models for investigating the evolution of somatosensation. Prior studies have uncovered tactile-responsive areas within the avian telencephalon, particularly in pigeons,3,4,5,6 parrots,7 and finches,8 but variations in somatosensory maps and responses across avian species are not fully understood. This study aims to explore somatotopic organization and neural coding in the telencephalon of Anna's hummingbirds (Calypte anna) and zebra finches (Taeniopygia guttata) by using in vivo extracellular electrophysiology to record activity in response to controlled tactile stimuli on various body regions. These findings reveal unique representations of body regions across distinct forebrain somatosensory nuclei, indicating significant differences in the extent of areas dedicated to certain body surfaces, which may correlate with their behavioral importance.

Relative telencephalon size does not affect collective motion in the guppy (Poecilia reticulata).

Collective motion is common across all animal taxa, from swarming insects to schools of fish. The collective motion requires intricate behavioral integration among individuals, yet little is known about how evolutionary changes in brain morphology influence the ability for individuals to coordinate behavior in groups. In this study, we utilized guppies that were selectively bred for relative telencephalon size, an aspect of brain morphology that is normally associated with advanced cognitive functions, to examine its role in collective motion using an open-field assay. We analyzed high-resolution tracking data of same-sex shoals consisting of 8 individuals to assess different aspects of collective motion, such as alignment, attraction to nearby shoal members, and swimming speed. Our findings indicate that variation in collective motion in guppy shoals might not be strongly affected by variation in relative telencephalon size. Our study suggests that group dynamics in collectively moving animals are likely not driven by advanced cognitive functions but rather by fundamental cognitive processes stemming from relatively simple rules among neighboring individuals.

Cell coupling compensates for changes in single-cell Her6 dynamics and provides phenotypic robustness.

This paper investigates the effect of altering the protein expression dynamics of the bHLH transcription factor Her6 at the single-cell level in the embryonic zebrafish telencephalon. Using a homozygote endogenous Her6:Venus reporter and 4D single-cell tracking, we show that Her6 oscillates in neural telencephalic progenitors and that the fusion of protein destabilisation (PEST) domain alters its expression dynamics, causing most cells to downregulate Her6 prematurely. However, counterintuitively, oscillatory cells increase, with some expressing Her6 at high levels, resulting in increased heterogeneity of Her6 expression in the population. These tissue-level changes appear to be an emergent property of coupling between single-cells, as revealed by experimentally disrupting Notch signalling and by computationally modelling alterations in Her6 protein stability. Despite the profound differences in the single-cell Her6 dynamics, the size of the telencephalon is only transiently altered and differentiation markers do not exhibit significant differences early on; however, a small increase is observed at later developmental stages. Our study suggests that cell coupling provides a compensation strategy, whereby an almost normal phenotype is maintained even though single-cell gene expression dynamics are abnormal, granting phenotypic robustness.

Input and Output Connections of the Crow Nidopallium Caudolaterale.

The avian telencephalic structure nidopallium caudolaterale (NCL) functions as an analog to the mammalian prefrontal cortex. In crows, corvid songbirds, it plays a crucial role in higher cognitive and executive functions. These functions rely on the NCL's extensive telencephalic connections. However, systematic investigations into the brain-wide connectivity of the NCL in crows or other songbirds are lacking. Here, we studied its input and output connections by injecting retrograde and anterograde tracers into the carrion crow NCL. Our results, mapped onto a published carrion crow brain atlas, confirm NCL multisensory connections and extend prior pigeon findings by identifying a novel input from the hippocampal formation. Furthermore, we analyze crow NCL efferent projections to the arcopallium and report newly identified arcopallial neurons projecting bilaterally to the NCL. These findings help to clarify the role of the NCL as central executive hub in the corvid songbird brain.

Transgenic tools targeting the basal ganglia reveal both evolutionary conservation and specialization of neural circuits in zebrafish.

The cortico-basal ganglia circuit mediates decision making. Here, we generated transgenic tools for adult zebrafish targeting specific subpopulations of the components of this circuit and utilized them to identify evolutionary homologs of the mammalian direct- and indirect-pathway striatal neurons, which respectively project to the homologs of the internal and external segment of the globus pallidus (dorsal entopeduncular nucleus [dEN] and lateral nucleus of the ventral telencephalic area [Vl]) as in mammals. Unlike in mammals, the Vl mainly projects to the dEN directly, not by way of the subthalamic nucleus. Further single-cell RNA sequencing analysis reveals two pallidal output pathways: a major shortcut pathway directly connecting the dEN with the pallium and the evolutionarily conserved closed loop by way of the thalamus. Our resources and circuit map provide the common basis for the functional study of the basal ganglia in a small and optically tractable zebrafish brain for the comprehensive mechanistic understanding of the cortico-basal ganglia circuit.

Telencephalic eversion in embryos and early larvae of four teleost species.

The telencephalon of ray-finned fishes undergoes eversion, which is very different to the evagination that occurs in most other vertebrates. Ventricle morphogenesis is key to build an everted telencephalon. Thus, here we use the apical marker zona occludens 1 to understand ventricle morphology, extension of the tela choroidea and the eversion process during early telencephalon development of four teleost species: giant danio (Devario aequipinnatus), blind cavefish (Astyanax mexicanus), medaka (Oryzias latipes), and paradise fish (Macroposus opercularis). In addition, by using immunohistochemistry against tubulin and calcium-binding proteins, we analyze the general morphology of the telencephalon, showing changes in the location and extension of the olfactory bulb and other telencephalic regions from 2 to 5 days of development. We also analyze the impact of abnormal eye and telencephalon morphogenesis on eversion, showing that cyclops mutants do undergo eversion despite very dramatic abnormal eye morphology. We discuss how the formation of the telencephalic ventricle in teleost fish, with its characteristic shape, is a crucial event during eversion.

Adult sex change leads to extensive forebrain reorganization in clownfish.

Sexual differentiation of the brain occurs in all major vertebrate lineages but is not well understood at a molecular and cellular level. Unlike most vertebrates, sex-changing fishes have the remarkable ability to change reproductive sex during adulthood in response to social stimuli, offering a unique opportunity to understand mechanisms by which the nervous system can initiate and coordinate sexual differentiation. This study explores sexual differentiation of the forebrain using single nucleus RNA-sequencing in the anemonefish Amphiprion ocellaris, producing the first cellular atlas of a sex-changing brain. We uncover extensive sex differences in cell type-specific gene expression, relative proportions of cells, baseline neuronal excitation, and predicted inter-neuronal communication. Additionally, we identify the cholecystokinin, galanin, and estrogen systems as central molecular axes of sexual differentiation. Supported by these findings, we propose a model of neurosexual differentiation in the conserved vertebrate social decision-making network spanning multiple subtypes of neurons and glia, including neuronal subpopulations within the preoptic area that are positioned to regulate gonadal differentiation. This work deepens our understanding of sexual differentiation in the vertebrate brain and defines a rich suite of molecular and cellular pathways that differentiate during adult sex change in anemonefish.

MR insights into fetal brain development: what is normal and what is not.

Fetal brain development is a complex, rapid, and multi-dimensional process that can be documented with MRI. In the second and third trimesters, there are predictable developmental changes that must be recognized and differentiated from disease. This review delves into the key biological processes that drive fetal brain development, highlights normal developmental anatomy, and provides a framework to identify pathology. We will summarize the development of the cerebral hemispheres, sulci and gyri, extra-axial and ventricular cerebrospinal fluid, and corpus callosum and illustrate the most common abnormal findings in the clinical setting.

Involvement of nr2f genes in brain regionalization and eye development during early zebrafish development.

Nuclear receptor subfamily 2 group F (Nr2f) proteins are essential for brain development in mice, but little is known about their precise roles and their evolutionary diversification. In the present study, the expression patterns of major nr2f genes (nr2f1a, nr2f1b, and nr2f2) during early brain development were investigated in zebrafish. Comparisons of their expression patterns revealed similar but temporally and spatially distinct patterns after early somite stages in the brain. Frameshift mutations in the three nr2f genes, achieved using the CRISPR/Cas9 method, resulted in a smaller telencephalon and smaller eyes in the nr2f1a mutants; milder forms of those defects were present in the nr2f1b and nr2f2 mutants. Acridine orange staining revealed enhanced cell death in the brain and/or eyes in all nr2f homozygous mutants. The expression of regional markers in the brain did not suggest global defects in brain regionalization; however, shha expression in the preoptic area and hypothalamus, as well as fgf8a expression in the anterior telencephalon, was disturbed in nr2f1a and nr2f1b mutants, potentially leading to a defective telencephalon. Specification of the retina and optic stalk was also significantly affected. The overexpression of nr2f1b by injection of mRNA disrupted the anterior brain at a high dose, and the expression of pax6a in the eyes and fgf8a in the telencephalon at a low dose. The results of these loss- and gain-of-function approaches showed that nr2f genes regulate the development of the telencephalon and eyes in zebrafish embryos.

Molecular organization of neuronal cell types and neuromodulatory systems in the zebrafish telencephalon.

The function of neuronal networks is determined not only by synaptic connectivity but also by neuromodulatory systems that broadcast information via distributed connections and volume transmission. To understand the molecular constraints that organize neuromodulatory signaling in the telencephalon of adult zebrafish, we used transcriptomics and additional approaches to delineate cell types, to determine their phylogenetic conservation, and to map the expression of marker genes at high granularity. The combinatorial expression of GPCRs and cell-type markers indicates that all neuronal cell types are subject to modulation by multiple monoaminergic systems and distinct combinations of neuropeptides. Individual cell types were associated with multiple (typically >30) neuromodulatory signaling networks but expressed only a few diagnostic GPCRs at high levels, suggesting that different neuromodulatory systems act in combination, albeit with unequal weights. These results provide a detailed map of cell types and brain areas in the zebrafish telencephalon, identify core components of neuromodulatory networks, highlight the cell-type specificity of neuropeptides and GPCRs, and begin to decipher the logic of combinatorial neuromodulation.

Brain regions controlling courtship behavior in the bluehead wrasse.

Bluehead wrasses (Thalassoma bifasciatum) follow a socially controlled mechanism of sex determination. A socially dominant initial-phase (IP) female is able to transform into a new terminal-phase (TP) male if the resident TP male is no longer present. TP males display an elaborate array of courtship behaviors, including both color changes and motor behaviors. Little is known concerning the neural circuits that control male-typical courtship behaviors. This study used glutamate iontophoresis to identify regions that may be involved in courtship. Stimulation of the following brain regions elicited diverse types of color change responses, many of which appear similar to courtship color changes: the ventral telencephalon (supracommissural nucleus of the ventral telencephalon [Vs], lateral nucleus of the ventral telencephalon [Vl], ventral nucleus of the ventral telencephalon [Vv], and dorsal nucleus of the ventral telencephalon [Vd]), parts of the preoptic area (NPOmg and NPOpc), entopeduncular nucleus, habenular nucleus, and pretectal nuclei (PSi and PSm). Stimulation of two regions in the posterior thalamus (central posterior thalamic [CP] and dorsal posterior thalamic [DP]) caused movements of the pectoral fins that are similar to courtship fluttering and vibrations. Furthermore, these responses were elicited in female IP fish, indicating that circuits for sexual behaviors typical of TP males exist in females. Immunohistochemistry results revealed regions that are more active in fish that are not courting: interpeduncular nucleus, red nucleus, and ventrolateral thalamus (VL). Taken together, we propose that the telencephalic-habenular-interpeduncular pathway plays an important role in controlling and regulating courtship behaviors in TP males; in this model, in response to telencephalic input, the habenular nucleus inhibits the interpeduncular nucleus, thereby dis-inhibiting forebrain regions and promoting the expression of courtship behaviors.

Spatial transcriptomics map of the embryonic mouse brain - a tool to explore neurogenesis.

The developing brain has a well-organized anatomical structure comprising different types of neural and non-neural cells. Stem cells, progenitors and newborn neurons tightly interact with their neighbouring cells and tissue microenvironment, and this intricate interplay ultimately shapes the output of neurogenesis. Given the relevance of spatial cues during brain development, we acknowledge the necessity for a spatial transcriptomics map accessible to the neurodevelopmental community. To fulfil this need, we generated spatially resolved RNA sequencing (RNAseq) data from embryonic day 13.5 mouse brain sections immunostained for mitotic active neural and vascular cells. Unsupervised clustering defined specific cell type populations of diverse lineages and differentiation states. Differential expression analysis revealed unique transcriptional signatures across specific brain areas, uncovering novel features inherent to particular anatomical domains. Finally, we integrated existing single-cell RNAseq datasets into our spatial transcriptomics map, adding tissue context to single-cell RNAseq data. In summary, we provide a valuable tool that enables the exploration and discovery of unforeseen molecular players involved in neurogenesis, particularly in the crosstalk between different cell types.

Self-formation of concentric zones of telencephalic and ocular tissues and directional retinal ganglion cell axons.

The telencephalon and eye in mammals are originated from adjacent fields at the anterior neural plate. Morphogenesis of these fields generates telencephalon, optic-stalk, optic-disc, and neuroretina along a spatial axis. How these telencephalic and ocular tissues are specified coordinately to ensure directional retinal ganglion cell (RGC) axon growth is unclear. Here, we report self-formation of human telencephalon-eye organoids comprising concentric zones of telencephalic, optic-stalk, optic-disc, and neuroretinal tissues along the center-periphery axis. Initially-differentiated RGCs grew axons towards and then along a path defined by adjacent PAX2+ VSX2+ optic-disc cells. Single-cell RNA sequencing of these organoids not only confirmed telencephalic and ocular identities but also identified expression signatures of early optic-disc, optic-stalk, and RGCs. These signatures were similar to those in human fetal retinas. Optic-disc cells in these organoids differentially expressed FGF8 and FGF9; FGFR inhibitions drastically decreased early RGC differentiation and directional axon growth. Through the RGC-specific cell-surface marker CNTN2 identified here, electrophysiologically excitable RGCs were isolated under a native condition. Our findings provide insight into the coordinated specification of early telencephalic and ocular tissues in humans and establish resources for studying RGC-related diseases such as glaucoma.

A quantitative characterization of early neuron generation in the developing zebrafish telencephalon.

The adult brain is made up of anatomically and functionally distinct regions with specific neuronal compositions. At the root of this neuronal diversity are neural stem and progenitor cells (NPCs) that produce many neurons throughout embryonic development. During development, NPCs switch from initial expanding divisions to neurogenic divisions, which marks the onset of neurogenesis. Here, we aimed to understand when NPCs switch division modes to generate the first neurons in the anterior-most part of the zebrafish brain, the telencephalon. To this end, we used the deep learning-based segmentation method Cellpose and clonal analysis of individual NPCs to assess the production of neurons by NPCs in the first 24 h of zebrafish telencephalon development. Our results provide a quantitative atlas detailing the production of telencephalic neurons and NPC division modes between 14 and 24 h postfertilization. We find that within this timeframe, the switch to neurogenesis is gradual, with considerable heterogeneity in individual NPC neurogenic potential and division rates. This quantitative characterization of initial neurogenesis in the zebrafish telencephalon establishes a basis for future studies aimed at illuminating the molecular mechanisms and regulators of early neurogenesis.

Quantification of neuron production and neural progenitor division modes in zebrafish embryonic telencephalon up to 24 h postfertilization using deep learning-based segmentation and clonal analysis methods.