Antoine de Chevigny

The cerebral cortex comprises diverse excitatory and inhibitory neuron subtypes, each with distinct laminar positions and connectivity patterns. Yet, the molecular logic underlying their precise wiring remains poorly understood. To identify ligand–receptor (LR) interactions involved in cortical circuit assembly, we tracked gene expression dynamics in mice across major neuronal populations at 17 developmental stages using single-cell transcriptomics. This generated a comprehensive atlas of LR-mediated communication between excitatory and inhibitory neuron subtypes, capturing known and novel interactions. Notably, we identified NEOGENIN-1 as the principal receptor for CBLN4 during the perinatal period, mediating synapse formation between somatostatin-expressing interneurons and glutamatergic neurons. We also identified members of the cadherin superfamily as candidate regulators of perisomatic inhibition from parvalbumin-expressing basket cells onto deep and superficial excitatory neurons, exerting opposing effects on synapse formation. These findings suggest a context-dependent role for cadherins in synaptic specificity and underscore the power of single-cell transcriptomics for decoding the molecular mechanisms of cortical wiring.

The developmental shift from depolarizing to hyperpolarizing GABA responses is a pivotal step in the maturation of GABAergic transmission and cortical circuits; classically documented in principal neurons during the first postnatal week in the mouse cortex. Surprisingly, whether maturation of GABA-mediated responses follows the same temporal pattern in cortical interneurons (INs) remains unresolved. Leveraging an array of methods, a high-resolution cortical development mouse atlas and single-cell RNA sequencing, we identify and comprehensively characterize a population of early-maturing cortical INs in mice, distinguished by KCC2 expression at embryonic stages and concomitant hyperpolarizing GABA_A responses at birth. These early KCC2-expressing INs exhibit precocious intrinsic excitability, synaptic integration, and dendritic complexity at birth, contrasting delayed maturation in principal neurons and other INs. Spatial transcriptomics and differential gene expression (DGE) analyses reveal early KCC2-expressing INs localize predominantly to layer 5, express somatostatin, and show upregulation of synaptogenic genes, consistent with the recorded elevated synaptic activity. Crucially, evolutionary conservation of early KCC2-expressing INs in humans was demonstrated with analogous genetic profiles enriched for signaling and synaptic maturation pathways. This work resolves a critical gap in developmental neurobiology, demonstrating heterogenous GABAergic functional maturation within IN subpopulations and establishing KCC2 as a marker of early-maturing INs.

The cerebral cortex hosts a diverse array of excitatory glutamatergic and inhibitory GABAergic neuron types, each characterized by distinct positional and synaptic connectivity patterns. However, the molecular mechanisms orchestrating this precise organization remain largely unknown. To identify ligand-receptor (LR) pairs regulating interactions and connectivity among cortical neurons during embryonic and postnatal development, we analyzed the transcriptional dynamics of all genes across major cortical neuron subtypes at 17 developmental time points using single-cell transcriptomics. From these data, we constructed a comprehensive bioinformatic atlas that inferred significant LR-mediated interactions between glutamatergic and GABAergic neurons throughout cortical maturation. This atlas not only corroborated known interactions but also enabled the discovery of novel regulators, identifying two cadherin superfamily members as key mediators of perisomatic inhibition in deep and superficial layer excitatory neurons by parvalbumin-expressing basket cells. These findings underscore the power of large-scale transcriptional profiling to unravel fundamental molecular mechanisms driving cortical circuit assembly.

While the transcription factor NEUROD2 has recently been associated with epilepsy, its precise role during nervous system development remains unclear. Using a multi-scale approach, we set out to understand howNeurod2deletion affects the development of the cerebral cortex in mice. InNeurod2KO embryos, cortical projection neurons over-migrated, thereby altering the final size and position of layers. In juvenile and adults, spine density and turnover were dysregulated in apical but not basal compartments in layer 5 neurons. Patch-clamp recordings in layer 5 neurons of juvenile mice revealed increased intrinsic excitability. Bulk RNA sequencing showed dysregulated expression of many genes associated with neuronal excitability and synaptic function, whose human orthologs were strongly associated with autism spectrum disorders (ASD). At the behavior level,Neurod2KO mice displayed social interaction deficits, stereotypies, hyperactivity, and occasionally spontaneous seizures. Mice heterozygous forNeurod2had similar defects, indicating thatNeurod2is haploinsufficient. Finally, specific deletion ofNeurod2in forebrain excitatory neurons recapitulated cellular and behavioral phenotypes found in constitutive KO mice, revealing the region-specific contribution of dysfunctionalNeurod2in symptoms. Informed by these neurobehavioral features in mouse mutants, we identified eleven patients from eight families with a neurodevelopmental disorder including intellectual disability and ASD associated withNEUROD2pathogenic mutations. Our findings demonstrate crucial roles forNeurod2in neocortical development, whose alterations can cause neurodevelopmental disorders including intellectual disability and ASD.

Dendritic spines are small protrusions studding neuronal dendrites, first described in 1888 by Ramón y Cajal using his famous Golgi stainings. Around 50 years later the advance of electron microscopy (EM) confirmed Cajal’s intuition that spines constitute the postsynaptic site of most excitatory synapses in the mammalian brain. The finding that spine density decreases between young and adult ages in fixed tissues suggested that spines are dynamic. It is only a decade ago that two-photon microscopy (TPM) has unambiguously proven the dynamic nature of spines, through the repeated imaging of single spines in live animals. Spine dynamics comprise formation, disappearance, and stabilization of spines and are modulated by neuronal activity and developmental age. Here, we review several emerging concepts in the field that start to answer the following key questions: What are the external signals triggering spine dynamics and the molecular mechanisms involved? What is, in return, the role of spine dynamics in circuit-rewiring, learning, and neuropsychiatric disorders?

Somatic sensation is defined by the existence of a diversity of primary sensory neurons with unique biological features and response profiles to external and internal stimuli. However, there is no coherent picture about how this diversity of cell states is transcriptionally generated. Here, we use deep single cell analysis to resolve fate splits and molecular biasing processes during sensory neurogenesis in mice. Our results identify a complex series of successive and specific transcriptional changes in post-mitotic neurons that delineate hierarchical regulatory states leading to the generation of the main sensory neuron classes. In addition, our analysis identifies previously undetected early gene modules expressed long before fate determination although being clearly associated with defined sensory subtypes. Overall, the early diversity of sensory neurons is generated through successive bi-potential intermediates in which synchronization of relevant gene modules and concurrent repression of competing fate programs precede cell fate stabilization and final commitment.

The development of the cerebral cortex requires complex sequential processes that have to be precisely orchestrated. The localization and timing of neuronal progenitor proliferation and of neuronal migration define the identity, laminar positioning, and specific connectivity of each single cortical neuron. Alterations at any step of this organized series of events-due to genetic mutations or environmental factors-lead to defined brain pathologies collectively known as malformations of cortical development (MCDs), which are now recognized as a leading cause of drug-resistant epilepsy and intellectual disability. In this heterogeneous group of disorders, macroscopic alterations of brain structure (eg, heterotopic nodules, small or absent gyri, double cortex) can be recognized and probably subtend a general reorganization of neuronal circuits. In this review, we provide an overview of the molecular mechanisms that are implicated in the generation of genetic MCDs associated with aberrations at various steps of neurogenesis and cortical development.

We identified seven families associating NEUROD2 pathogenic mutations with ASD and intellectual disability. To get insight into the pathophysiological mechanisms, we analyzed cortical development in Neurod2 KO mice. Cortical projection neurons (CPNs) over-migrated during embryogenesis, inducing abnormal thickness and laminar positioning of cortical layers. At juvenile ages, dendritic spine turnover and intrinsic excitability were increased in L5 CPNs. Differentially expressed genes in Neurod2 KO mice were enriched for voltage-gated ion channels, and the human orthologs of these genes were strongly associated with ASD. Furthermore, adult Neurod2 KO mice exhibited core ASD-like behavioral abnormalities. Finally, by generating Neurod2 conditional mutant mice we demonstrate that forebrain excitatory neuron-specific Neurod2 deletion recapitulates cellular and behavioral ASD phenotypes found in full KO mice. Our findings demonstrate crucial roles for Neurod2 in cortical development and function, whose alterations likely account for ASD and related symptoms in the newly defined NEUROD2 mutation syndrome.

In the postnatal forebrain regionalized neural stem cells along the ventricular walls produce olfactory bulb (OB) interneurons with varying neurotransmitter phenotypes and positions. To understand the molecular basis of this region-specific variability we analyzed gene expression in the postnatal dorsal and lateral lineages in mice of both sexes from stem cells to neurons. We show that both lineages maintain transcription factor signatures of their embryonic site of origin, the pallium and subpallium. However, additional factors, including Zic1 and Zic2, are postnatally expressed in the dorsal stem cell compartment and maintained in the lineage that generates calretinin-positive GABAergic neurons for the OB. Functionally, we show that Zic1 and Zic2 induce the generation of calretinin-positive neurons while suppressing dopaminergic fate in the postnatal dorsal lineage. We investigated the evolutionary conservation of the dopaminergic repressor function of Zic proteins and show that it is already present inC. elegans.SIGNIFICANCE STATEMENTThe vertebrate brain generates thousands of different neuron types. In this work we investigate the molecular mechanisms underlying this variability. Using a genomics approach we identify the transcription factor signatures of defined neural stem cells and neuron populations. Based thereon we show that two related transcription factors, Zic1 and Zic2, are essential to control the balance between two defined neuron types in the postnatal brain. We show that this mechanism is conserved in evolutionary very distant species.