Mutations in MECP2 give rise to Rett syndrome (RTT), an X-linked neurodevelopmental disorder that results in broad cognitive impairments in females. While the exact etiology of RTT symptoms remains unknown, one possible explanation for its clinical presentation is that loss of MECP2 causes miswiring of neural circuits due to defects in the brain’s capacity to respond to changes in neuronal activity and sensory experience. Here, we show that MeCP2 is phosphorylated at four residues in the mouse brain (S86, S274, T308, and S421) in response to neuronal activity, and we generate a quadruple knock-in (QKI) mouse line in which all four activity-dependent sites are mutated to alanines to prevent phosphorylation. QKI mice do not display overt RTT phenotypes or detectable gene expression changes in two brain regions. However, electrophysiological recordings from the retinogeniculate synapse of QKI mice reveal that while synapse elimination is initially normal at P14, it is significantly compromised at P20. Notably, this phenotype is distinct from the synapse refinement defect previously reported for Mecp2 null mice, where synapses initially refine but then regress after the third postnatal week. We thus propose a model in which activity-induced phosphorylation of MeCP2 is critical for the proper timing of retinogeniculate synapse maturation specifically during the early postnatal period.

The complete and non-redundant coverage of sensory tissues by neighboring neurons enables effective detection of stimuli in the environment. How the neurites of adjacent neurons establish their boundaries to achieve this completeness in coverage remains incompletely understood. Here, we use distinct fluorescent reporters to study two neighboring sensory neurons with complex dendritic arbors, FLP and PVD, in C. elegans . We quantify the sizes of their dendritic fields, and identify CWN-2/Wnt and LIN-17/Frizzled as a ligand and receptor that regulate the relative dendritic field sizes of these two neurons. Loss of either cwn-2 or lin-17 results in complementary changes in the size of the dendritic fields of both neurons; the FLP arbor expands, while that of PVD shrinks. Using an endogenous knock-in mNeonGreen-CWN-2/Wnt, we find that CWN-2/Wnt is localized along the path of growing FLP dendrites. Dynamic imaging shows a significant braking of FLP dendrite growth upon CWN-2/Wnt contact. We find that LIN-17/Frizzled functions cell-autonomously in FLP to limit dendritic field size and propose that PVD fills the space left by FLP through contact-induced retraction. Our results reveal that interactions of dendrites with adjacent dendrites and with environmental cues both shape the boundaries of neighboring dendritic fields.Highlights:▫ Secreted Wnt CWN-2 and cell-autonomous activity of neuronal LIN-17/Frizzled receptors restrict FLP dendritic field sizes▫ Endogenously tagged CWN-2/Wnt is punctate and visible in the same plane of growing FLP dendrites▫ Growth of developing FLP dendrites is inhibited upon contact with extracellular CWN-2/Wnt and with PVD dendrites.

Nervous systems exhibit dramatic diversity in cell morphology and size. How neurons regulate their biosynthetic and secretory machinery to support such diversity is not well understood. Endoplasmic reticulum exit sites (ERESs) are essential for maintaining secretory flux, and are required for normal dendrite development, but how neurons of different size regulate secretory capacity remains unknown. In Caenorhabditis elegans, we find that the ERES number is strongly correlated with the size of a neuron’s dendritic arbor. The elaborately branched sensory neuron, PVD, has especially high ERES numbers. Asymmetric cell division provides PVD with a large initial cell size critical for rapid establishment of PVD’s high ERES number before neurite outgrowth, and these ERESs are maintained throughout development. Maintenance of ERES number requires the cell fate transcription factor MEC-3, C. elegans TOR ( ceTOR/let-363), and nutrient availability, with mec-3 and ceTOR/let-363 mutant PVDs both displaying reductions in ERES number, soma size, and dendrite size. Notably, mec-3 mutant animals exhibit reduced expression of a ceTOR/let-363 reporter in PVD, and starvation reduces ERES number and somato-dendritic size in a manner genetically redundant with ceTOR/let-363 perturbation. Our data suggest that both asymmetric cell division and nutrient sensing pathways regulate secretory capacities to support elaborate dendritic arbors.

Nervous systems exhibit dramatic diversity in cell morphology and size. How neurons regulate their biosynthetic and secretory machinery to support such diversity is not well understood. Endoplasmic reticulum exit sites (ERESs) are essential for maintaining secretory flux, and are required for normal dendrite development, but how neurons of different size regulate secretory capacity remains unknown. In C. elegans, we find that the ERES number is strongly correlated with the size of a neuron’s dendritic arbor. The elaborately branched sensory neuron, PVD, has especially high ERES numbers. Asymmetric cell division provides PVD with a large initial cell size critical for rapid establishment of PVD’s high ERES number before neurite outgrowth, and these ERESs are maintained throughout development. Maintenance of ERES number requires the cell fate transcription factor MEC-3, C. elegans TOR ( ceTOR/let-363 ), and nutrient availability, with mec-3 and ceTOR/let-363 mutant PVDs both displaying reductions in ERES number, soma size, and dendrite size. Notably, mec-3 mutant animals exhibit reduced expression of a ceTOR/let-363 reporter in PVD, and starvation reduces ERES number and somato-dendritic size in a manner genetically redundant with ceTOR/let-363 perturbation. Our data suggest that both asymmetric cell division and nutrient sensing pathways regulate secretory capacities to support elaborate dendritic arbors.

Dendritic arbor architecture profoundly impacts neuronal connectivity and function, and aberrant dendritic morphology characterizes neuropsychiatric disorders. Here, we identify the adhesion-GPCR BAI1 as an important regulator of dendritic arborization. BAI1 loss from mouse or rat hippocampal neurons causes dendritic hypertrophy, whereas BAI1 overexpression precipitates dendrite retraction. These defects specifically manifest as dendrites transition from growth to stability. BAI1-mediated growth arrest is independent of its Rac1-dependent synaptogenic function. Instead, BAI1 couples to the small GTPase RhoA, driving late RhoA activation in dendrites coincident with growth arrest. BAI1 loss lowers RhoA activation and uncouples it from dendrite dynamics, causing overgrowth. None of BAI1’s known downstream effectors mediates BAI1-dependent growth arrest. Rather, BAI1 associates with the Rho-GTPase regulatory protein Bcr late in development and stimulates its cryptic RhoA-GEF activity, which functions together with its Rac1-GAP activity to terminate arborization. Our results reveal a late-acting signaling pathway mediating a key transition in dendrite development.

Enhancers are the primary DNA regulatory elements that confer cell type specificity of gene expression. Recent studies characterizing individual enhancers have revealed their potential to direct heterologous gene expression in a highly cell-type-specific manner. However, it has not yet been possible to systematically identify and test the function of enhancers for each of the many cell types in an organism. We have developed PESCA, a scalable and generalizable method that leverages ATAC- and single-cell RNA-sequencing protocols, to characterize cell-type-specific enhancers that should enable genetic access and perturbation of gene function across mammalian cell types. Focusing on the highly heterogeneous mammalian cerebral cortex, we apply PESCA to find enhancers and generate viral reagents capable of accessing and manipulating a subset of somatostatin-expressing cortical interneurons with high specificity. This study demonstrates the utility of this platform for developing new cell-type-specific viral reagents, with significant implications for both basic and translational research.

Sensory experience influences the establishment of neural connectivity through molecular mechanisms that remain unclear. Here, we employ single-nucleus RNA sequencing to investigate the contribution of sensory-driven gene expression to synaptic refinement in the dorsal lateral geniculate nucleus of the thalamus, a region of the brain that processes visual information. We find that visual experience induces the expression of the cytokine receptor Fn14 in excitatory thalamocortical neurons. By combining electrophysiological and structural techniques, we show that Fn14 is dispensable for early phases of refinement mediated by spontaneous activity but that Fn14 is essential for refinement during a later, experience-dependent period of development. Refinement deficits in mice lacking Fn14 are associated with functionally weaker and structurally smaller retinogeniculate inputs, indicating that Fn14 mediates both functional and anatomical rearrangements in response to sensory experience. These findings identify Fn14 as a molecular link between sensory-driven gene expression and vision-sensitive refinement in the brain.

Igf1 is identified in mice as an experience-induced gene that functions cell-autonomously to increase inhibitory input onto a disinhibitory subtype of GABAergic neurons in the cortex, affecting the downstream excitation–inhibition balance within circuits that regulate visual acuity, and providing a novel example of experience modulating neural plasticity. The mechanisms by which experience influences the different types of inhibitory neurons controlling distinct computations in cortical circuits are not well known. Here Michael Greenberg and colleagues identify an experience-dependent gene expression program in vasoactive intestinal polypeptide (VIP)-expressing interneurons of the visual cortex that is distinct from those expression patterns observed in other neuronal subtypes. IGF1 (insulin-like growth factor 1) was identified as an activity-dependent gene that functions cell autonomously to increase inhibitory input onto VIP neurons, affecting downstream excitation–inhibition balance within circuits that regulate visual acuity. Inhibitory neurons regulate the adaptation of neural circuits to sensory experience1, but the molecular mechanisms by which experience controls the connectivity between different types of inhibitory neuron2,3 to regulate cortical plasticity are largely unknown. Here we show that exposure of dark-housed mice to light induces a gene program in cortical vasoactive intestinal peptide (VIP)-expressing neurons that is markedly distinct from that induced in excitatory neurons and other subtypes of inhibitory neuron. We identify Igf1 as one of several activity-regulated genes that are specific to VIP neurons, and demonstrate that IGF1 functions cell-autonomously in VIP neurons to increase inhibitory synaptic input onto these neurons. Our findings further suggest that in cortical VIP neurons, experience-dependent gene transcription regulates visual acuity by activating the expression of IGF1, thus promoting the inhibition of disinhibitory neurons3,4,5 and affecting inhibition onto cortical pyramidal neurons.

The nervous system adapts to experience by inducing a transcriptional program that controls important aspects of synaptic plasticity. Although the molecular mechanisms of experience-dependent plasticity are well characterized in excitatory neurons, the mechanisms that regulate this process in inhibitory neurons are only poorly understood. Here, we describe a transcriptional program that is induced by neuronal activity in inhibitory neurons. We find that, while neuronal activity induces expression of early-response transcription factors such as Npas4 in both excitatory and inhibitory neurons, Npas4 activates distinct programs of late-response genes in inhibitory and excitatory neurons. These late-response genes differentially regulate synaptic input to these two types of neurons, promoting inhibition onto excitatory neurons while inducing excitation onto inhibitory neurons. These findings suggest that the functional outcomes of activity-induced transcriptional responses are adapted in a cell-type-specific manner to achieve a circuit-wide homeostatic response.

Excitatory synapses are polarized structures that primarily reside on dendritic spines in the brain. The small GTPase Rac1 regulates the development and plasticity of synapses and spines by modulating actin dynamics. By restricting the Rac1-guanine nucleotide exchange factor Tiam1 to spines, the polarity protein Par3 promotes synapse development by spatially controlling Rac1 activation. However, the mechanism for recruiting Par3 to spines is unknown. Here, we identify brain-specific angiogenesis inhibitor 1 (BAI1) as a synaptic adhesion GPCR that is required for spinogenesis and synaptogenesis in mice and rats. We show that BAI1 interacts with Par3/Tiam1 and recruits these proteins to synaptic sites. BAI1 knockdown results in Par3/Tiam1 mislocalization and loss of activated Rac1 and filamentous actin from spines. Interestingly, BAI1 also mediates Rac-dependent engulfment in professional phagocytes through its interaction with a different Rac1-guanine nucleotide exchange factor module, ELMO/DOCK180. However, this interaction is dispensable for BAI1's role in synapse development because a BAI1 mutant that cannot interact with ELMO/DOCK180 rescues spine defects in BAI1-knockdown neurons, whereas a mutant that cannot interact with Par3/Tiam1 rescues neither spine defects nor Par3 localization. Further, overexpression of Tiam1 rescues BAI1 knockdown spine phenotypes. These results indicate that BAI1 plays an important role in synaptogenesis that is mechanistically distinct from its role in phagocytosis. Furthermore, our results provide the first example of a cell surface receptor that targets members of the PAR polarity complex to synapses.