Melanopsin retinal ganglion cells mediate light-promoted brain development.

During development, melanopsin-expressing intrinsically photosensitive retinal ganglion cells (ipRGCs) become light sensitive much earlier than rods and cones. IpRGCs project to many subcortical areas, whereas physiological functions of these projections are yet to be fully elucidated. Here, we found that ipRGC-mediated light sensation promotes synaptogenesis of pyramidal neurons in various cortices and the hippocampus. This phenomenon depends on activation of ipRGCs and is mediated by the release of oxytocin from the supraoptic nucleus (SON) and the paraventricular nucleus (PVN) into cerebral-spinal fluid. We further characterized a direct connection between ipRGCs and oxytocin neurons in the SON and mutual projections between oxytocin neurons in the SON and PVN. Moreover, we showed that the lack of ipRGC-mediated, light-promoted early cortical synaptogenesis compromised learning ability in adult mice. Our results highlight the importance of light sensation early in life on the development of learning ability and therefore call attention to suitable light environment for infant care.

Heparan Sulfate Organizes Neuronal Synapses through Neurexin Partnerships.

Synapses are fundamental units of communication in the brain. The prototypical synapse-organizing complex neurexin-neuroligin mediates synapse development and function and is central to a shared genetic risk pathway in autism and schizophrenia. Neurexin's role in synapse development is thought to be mediated purely by its protein domains, but we reveal a requirement for a rare glycan modification. Mice lacking heparan sulfate (HS) on neurexin-1 show reduced survival, as well as structural and functional deficits at central synapses. HS directly binds postsynaptic partners neuroligins and LRRTMs, revealing a dual binding mode involving intrinsic glycan and protein domains for canonical synapse-organizing complexes. Neurexin HS chains also bind novel ligands, potentially expanding the neurexin interactome to hundreds of HS-binding proteins. Because HS structure is heterogeneous, our findings indicate an additional dimension to neurexin diversity, provide a molecular basis for fine-tuning synaptic function, and open therapeutic directions targeting glycan-binding motifs critical for brain development.

Astrocyte-immune cell interactions in physiology and pathology.

Astrocytes play both physiological and pathological roles in maintaining central nervous system (CNS) function. Here, we review the varied functions of astrocytes and how these might change in subsets of reactive astrocytes. We review the current understanding of astrocyte interactions with microglia and the vasculature and protective barriers in the central nervous system as well as highlight recent insights into physiologic and reactive astrocyte sub-states identified by transcriptional profiling. Our goal is to stimulate inquiry into how these molecular identifiers link to specific functional changes in astrocytes and to define the implications of these heterogeneous molecular and functional changes in brain function and pathology. Defining these complex interactions has the potential to yield new therapies in CNS injury, infection, and disease.

Control of Synaptic Connectivity by a Network of Drosophila IgSF Cell Surface Proteins.

We have defined a network of interacting Drosophila cell surface proteins in which a 21-member IgSF subfamily, the Dprs, binds to a nine-member subfamily, the DIPs. The structural basis of the Dpr-DIP interaction code appears to be dictated by shape complementarity within the Dpr-DIP binding interface. Each of the six dpr and DIP genes examined here is expressed by a unique subset of larval and pupal neurons. In the neuromuscular system, interactions between Dpr11 and DIP-γ affect presynaptic terminal development, trophic factor responses, and neurotransmission. In the visual system, dpr11 is selectively expressed by R7 photoreceptors that use Rh4 opsin (yR7s). Their primary synaptic targets, Dm8 amacrine neurons, express DIP-γ. In dpr11 or DIP-γ mutants, yR7 terminals extend beyond their normal termination zones in layer M6 of the medulla. DIP-γ is also required for Dm8 survival or differentiation. Our findings suggest that Dpr-DIP interactions are important determinants of synaptic connectivity.

Oxytocin signaling is necessary for synaptic maturation of adult-born neurons.

Neural circuit plasticity and sensory response dynamics depend on forming new synaptic connections. Despite recent advances toward understanding the consequences of circuit plasticity, the mechanisms driving circuit plasticity are unknown. Adult-born neurons within the olfactory bulb have proven to be a powerful model for studying circuit plasticity, providing a broad and accessible avenue into neuron development, migration, and circuit integration. We and others have shown that efficient adult-born neuron circuit integration hinges on presynaptic activity in the form of diverse signaling peptides. Here, we demonstrate a novel oxytocin-dependent mechanism of adult-born neuron synaptic maturation and circuit integration. We reveal spatial and temporal enrichment of oxytocin receptor expression within adult-born neurons in the murine olfactory bulb, with oxytocin receptor expression peaking during activity-dependent integration. Using viral labeling, confocal microscopy, and cell type-specific RNA-seq, we demonstrate that oxytocin receptor signaling promotes synaptic maturation of newly integrating adult-born neurons by regulating their morphological development and expression of mature synaptic AMPARs and other structural proteins.

Pathophysiological roles of thrombospondin-4 in disease development.

Thrombospondin-4 (TSP-4) belongs to the extracellular matrix glycoprotein family of thrombospondins (TSPs). The multidomain, pentameric structure of TSP-4 allows its interactions with numerous extracellular matrix components, proteins and signaling molecules that enable its modulation to various physiological and pathological processes. Characterization of TSP-4 expression under development and pathogenesis of disorders has yielded important insights into mechanisms underlying the unique role of TSP-4 in mediating various processes including cell-cell, cell-extracellular matrix interactions, cell migration, proliferation, tissue remodeling, angiogenesis, and synaptogenesis. Maladaptation of these processes in response to pathological insults and stress can accelerate the development of disorders including skeletal dysplasia, osteoporosis, degenerative joint disease, cardiovascular diseases, tumor progression/metastasis and neurological disorders. Overall, the diverse functions of TSP-4 suggest that it may be a potential marker or therapeutic target for prognosis, diagnosis, and treatment of various pathological conditions upon further investigations. This review article highlights recent findings on the role of TSP-4 in both physiological and pathological conditions with a focus on what sets it apart from other TSPs.

Neuropsin promotes hippocampal synaptogenesis by regulating the expression and cleavage of L1CAM.

During early postnatal brain development, the formation of proper synaptic connections between neurons is crucial for the development of functional neural networks. Recent studies have established the involvement of protease-mediated modulations of extracellular components in both synapse formation and elimination. The secretory serine protease neuropsin (also known as kallikrein-8) cleaves a few transmembrane or extracellular matrix proteins in a neural activity-dependent manner and regulates neural plasticity. However, neuropsin-dependent proteolysis of extracellular components and the involvement of these components in mouse brain development are poorly understood. We have observed that during hippocampus development, expression of neuropsin and levels of full-length or cleaved fragments of the neuropsin substrate protein L1 cell adhesion molecule (L1CAM) positively correlate with synaptogenesis. Our subcellular fractionation studies show that the expression of neuropsin and its proteolytic activity on L1CAM are enriched at developing hippocampal synapses. Activation of neuropsin expression upregulates the transcription and cleavage of L1CAM. Furthermore, blocking of neuropsin activity, as well as knockdown of L1CAM expression, significantly downregulates in vitro hippocampal synaptogenesis. Taken together, these findings provide evidence for the involvement of neuropsin activity-dependent regulation of L1CAM expression and cleavage in hippocampal synaptogenesis.

Immunoglobulin-Like Receptors and Their Impact on Wiring of Brain Synapses.

Synapse formation is mediated by a surprisingly large number and wide variety of genes encoding many different protein classes. One of the families increasingly implicated in synapse wiring is the immunoglobulin superfamily (IgSF). IgSF molecules are by definition any protein containing at least one Ig-like domain, making this family one of the most common protein classes encoded by the genome. Here, we review the emerging roles for IgSF molecules in synapse formation specifically in the vertebrate brain, focusing on examples from three classes of IgSF members: ( a) cell adhesion molecules, ( b) signaling molecules, and ( c) immune molecules expressed in the brain. The critical roles for IgSF members in regulating synapse formation may explain their extensive involvement in neuropsychiatric and neurodevelopmental disorders. Solving the IgSF code for synapse formation may reveal multiple new targets for rescuing IgSF-mediated deficits in synapse formation and, eventually, new treatments for psychiatric disorders caused by altered IgSF-induced synapse wiring.

Engineered adhesion molecules drive synapse organization.

In multicellular organisms, cell-adhesion molecules connect cells into tissues and mediate intercellular signaling between these cells. In vertebrate brains, synaptic cell-adhesion molecules (SAMs) guide the formation, specification, and plasticity of synapses. Some SAMs, when overexpressed in cultured neurons or in heterologous cells co-cultured with neurons, drive formation of synaptic specializations onto the overexpressing cells. However, genetic deletion of the same SAMs from neurons often has no effect on synapse numbers, but frequently severely impairs synaptic transmission, suggesting that most SAMs control the function and plasticity of synapses (i.e., organize synapses) instead of driving their initial establishment (i.e., make synapses). Since few SAMs were identified that mediate initial synapse formation, it is difficult to develop methods that enable experimental control of synaptic connections by targeted expression of these SAMs. To overcome this difficulty, we engineered novel SAMs from bacterial proteins with no eukaryotic homologues that drive synapse formation. We named these engineered adhesion proteins "Barnoligin" and "Starexin" because they were assembled from parts of Barnase and Neuroligin-1 or of Barstar and Neurexin3ß, respectively. Barnoligin and Starexin robustly induce the formation of synaptic specializations in a specific and directional manner in cultured neurons. Synapse formation by Barnoligin and Starexin requires both their extracellular Barnase- and Barstar-derived interaction domains and their Neuroligin- and Neurexin-derived intracellular signaling domains. Our findings support a model of synapse formation whereby trans-synaptic interactions by SAMs drive synapse organization via adhesive interactions that activate signaling cascades.

RNA-binding FMRP and Staufen sequentially regulate the Coracle scaffold to control synaptic glutamate receptor and bouton development.

Both mRNA-binding Fragile X mental retardation protein (FMRP; Fmr1) and mRNA-binding Staufen regulate synaptic bouton formation and glutamate receptor (GluR) levels at the Drosophila neuromuscular junction (NMJ) glutamatergic synapse. Here, we tested whether these RNA-binding proteins act jointly in a common mechanism. We found that both dfmr1 and staufen mutants, and trans-heterozygous double mutants, displayed increased synaptic bouton formation and GluRIIA accumulation. With cell-targeted RNA interference, we showed a downstream Staufen role within postsynaptic muscle. With immunoprecipitation, we showed that FMRP binds staufen mRNA to stabilize postsynaptic transcripts. Staufen is known to target actin-binding, GluRIIA anchor Coracle, and we confirmed that Staufen binds to coracle mRNA. We found that FMRP and Staufen act sequentially to co-regulate postsynaptic Coracle expression, and showed that Coracle, in turn, controls GluRIIA levels and synaptic bouton development. Consistently, we found that dfmr1, staufen and coracle mutants elevate neurotransmission strength. We also identified that FMRP, Staufen and Coracle all suppress pMad activation, providing a trans-synaptic signaling linkage between postsynaptic GluRIIA levels and presynaptic bouton development. This work supports an FMRP-Staufen-Coracle-GluRIIA-pMad pathway regulating structural and functional synapse development.

Imaging three-dimensional brain organoid architecture from meso- to nanoscale across development.

Organoids are stem cell-derived three-dimensional cultures offering a new avenue to model human development and disease. Brain organoids allow the study of various aspects of human brain development in the finest details in vitro in a tissue-like context. However, spatial relationships of subcellular structures, such as synaptic contacts between distant neurons, are hardly accessible by conventional light microscopy. This limitation can be overcome by systems that quickly image the entire organoid in three dimensions and in super-resolution. To that end we have developed a system combining tissue expansion and light-sheet fluorescence microscopy for imaging and quantifying diverse spatial parameters during organoid development. This technique enables zooming from a mesoscopic perspective into super-resolution within a single imaging session, thus revealing cellular and subcellular structural details in three spatial dimensions, including unequivocal delineation of mitotic cleavage planes as well as the alignment of pre- and postsynaptic proteins. We expect light-sheet fluorescence expansion microscopy to facilitate qualitative and quantitative assessment of organoids in developmental and disease-related studies.

Cyclase-associated protein (CAP) inhibits inverted formin 2 (INF2) to induce dendritic spine maturation.

The morphology of dendritic spines, the postsynaptic compartment of most excitatory synapses, decisively modulates the function of neuronal circuits as also evident from human brain disorders associated with altered spine density or morphology. Actin filaments (F-actin) form the backbone of spines, and a number of actin-binding proteins (ABP) have been implicated in shaping the cytoskeleton in mature spines. Instead, only little is known about the mechanisms that control the reorganization from unbranched F-actin of immature spines to the complex, highly branched cytoskeleton of mature spines. Here, we demonstrate impaired spine maturation in hippocampal neurons upon genetic inactivation of cyclase-associated protein 1 (CAP1) and CAP2, but not of CAP1 or CAP2 alone. We found a similar spine maturation defect upon overactivation of inverted formin 2 (INF2), a nucleator of unbranched F-actin with hitherto unknown synaptic function. While INF2 overactivation failed in altering spine density or morphology in CAP-deficient neurons, INF2 inactivation largely rescued their spine defects. From our data we conclude that CAPs inhibit INF2 to induce spine maturation. Since we previously showed that CAPs promote cofilin1-mediated cytoskeletal remodeling in mature spines, we identified them as a molecular switch that control transition from filopodia-like to mature spines.

Microglia regulate chandelier cell axo-axonic synaptogenesis.

SignificanceChandelier cells (ChCs) are a unique type of GABAergic interneuron that form axo-axonic synapses exclusively on the axon initial segment (AIS) of neocortical pyramidal neurons (PyNs), allowing them to exert powerful yet precise control over PyN firing and population output. The importance of proper ChC function is further underscored by the association of ChC connectivity defects with various neurological conditions. Despite this, the cellular mechanisms governing ChC axo-axonic synapse formation remain poorly understood. Here, we identify microglia as key regulators of ChC axonal morphogenesis and AIS synaptogenesis, and show that disease-induced aberrant microglial activation perturbs proper ChC synaptic development/connectivity in the neocortex. In doing so, such findings highlight the therapeutic potential of manipulating microglia to ensure proper brain wiring.

Progesterone and contraceptive progestin actions on the brain: A systematic review of animal studies and comparison to human neuroimaging studies.

In this review we systematically summarize the effects of progesterone and synthetic progestins on neurogenesis, synaptogenesis, myelination and six neurotransmitter systems. Several parallels between progesterone and older generation progestin actions emerged, suggesting actions via progesterone receptors. However, existing results suggest a general lack of knowledge regarding the effects of currently used progestins in hormonal contraception regarding these cellular and molecular brain parameters. Human neuroimaging studies were reviewed with a focus on randomized placebo-controlled trials and cross-sectional studies controlling for progestin type. The prefrontal cortex, amygdala, salience network and hippocampus were identified as regions of interest for future preclinical studies. This review proposes a series of experiments to elucidate the cellular and molecular actions of contraceptive progestins in these areas and link these actions to behavioral markers of emotional and cognitive functioning. Emotional effects of contraceptive progestins appear to be related to 1) alterations in the serotonergic system, 2) direct/indirect modulations of inhibitory GABA-ergic signalling via effects on the allopregnanolone content of the brain, which differ between androgenic and anti-androgenic progestins. Cognitive effects of combined oral contraceptives appear to depend on the ethinylestradiol dose.

Neuroestradiol and neuronal development: Not an exclusive male tale anymore.

The brain synthesizes a variety of neurosteroids, including neuroestradiol. Inhibition of neuroestradiol synthesis results in alterations in basic neurodevelopmental processes, such as neurogenesis, neuroblast migration, neuritogenesis and synaptogenesis. Although the neurodevelopmental actions of neuroestradiol are exerted in both sexes, some of them are sex-specific, such as the well characterized effects of neuroestradiol derived from the metabolism of testicular testosterone during critical periods of male brain development. In addition, recent findings have shown sex-specific actions of neuroestradiol on neuroblast migration, neuritic growth and synaptogenesis in females. Among other factors, the epigenetic regulation exerted by X linked genes, such as Kdm6a/Utx, may determine sex-specific actions of neuroestradiol in the female brain. This review evidences the impact of neuroestradiol on brain formation in both sexes and highlights the interaction of neural steriodogenesis, hormones and sex chromosomes in sex-specific brain development.

Calsyntenin-1 expression and function in brain tissue of lithium-pilocarpine rat seizure models.

To present the expression of calsyntenin-1 (Clstn1) in the brain and investigate the potential mechanism of Clstn1 in lithium-pilocarpine rat seizure models. Thirty-five male SD adult rats were induced to have seizures by intraperitoneal injection of lithium chloride pilocarpine. Rats exhibiting spontaneous seizures were divided into the epilepsy (EP) group (n = 15), whereas those without seizures were divided into the control group (n = 14). Evaluate the expression of Clstn1 in the temporal lobe of two groups using Western blotting, immunohistochemistry, and immunofluorescence. Additionally, 55 male SD rats were subjected to status epilepticus (SE) using the same induction method. Rats experiencing seizures exceeding Racine's level 4 (n = 48) were randomly divided into three groups: SE, SE + control lentivirus (lentiviral vector expressing green fluorescent protein [LV-GFP]), and SE + Clstn1-targeted RNA interference lentivirus (LV-Clstn1-RNAi). The LV-GFP group served as a control for the lentiviral vector, whereas the LV-Clstn1-RNAi group received a lentivirus designed to silence Clstn1 expression. These lentiviral treatments were administered via hippocampal stereotactic injection 2 days after SE induction. Seven days after SE, Western blot analysis was performed to evaluate the expression of Clstn1 in the hippocampus and temporal lobe. Meanwhile, we observed the latency of spontaneous seizures and the frequency of spontaneous seizures within 8 weeks among the three groups. The expression of Clstn1 in the cortex and hippocampus of the EP group was significantly increased compared to the control group (p < .05). Immunohistochemistry and immunofluorescence showed that Clstn1 was widely distributed in the cerebral cortex and hippocampus of rats, and colocalization analysis revealed that it was mainly co expressed with neurons in the cytoplasm. Compared with the SE group (11.80 ± 2.17 days) and the SE + GFP group (12.40 ± 1.67 days), there was a statistically significant difference (p < .05) in the latency period of spontaneous seizures (15.14 ± 2.41 days) in the SE + Clstn1 + RNAi group rats. Compared with the SE group (4.60 ± 1.67 times) and the SE + GFP group (4.80 ± 2.05 times), the SE + Clstn1 + RNAi group (2.0 ± .89 times) showed a significant reduction in the frequency of spontaneous seizures within 2 weeks of chronic phase in rats (p < .05). Elevated Clstn1 expression in EP group suggests its role in EP onset. Targeting Clstn1 may be a potential therapeutic approach for EP management.

Zebrafish and nematodes as whole organism models to measure developmental neurotoxicity.

Despite the growing epidemiological evidence of an association between toxin exposure and developmental neurotoxicity (DNT), systematic testing of DNT is not mandatory in international regulations for admission of pharmaceuticals or industrial chemicals. However, to date around 200 compounds, ranging from pesticides, pharmaceuticals and industrial chemicals, have been tested for DNT in the current OECD test guidelines (TG-443 or TG-426). There are calls for the development of new approach methodologies (NAMs) for DNT, which has resulted in a DNT testing battery using in vitro human cell-based assays. These assays provide a means to elucidate the molecular mechanisms of toxicity in humans which is lacking in animal-based toxicity tests. However, cell-based assays do not represent all steps of the complex process leading to DNT. Validated models with a multi-organ network of pathways that interact at the molecular, cellular and tissue level at very specific timepoints in a life cycle are currently missing. Consequently, whole model organisms are being developed to screen for, and causally link, new molecular targets of DNT compounds and how they affect whole brain development and neurobehavioral endpoints. Given the practical and ethical restraints associated with vertebrate testing, lower animal models that qualify as 3 R (reduce, refine and replace) models, including the nematode (Caenorhabditis elegans) and the zebrafish (Danio rerio) will prove particularly valuable for unravelling toxicity pathways leading to DNT. Although not as complex as the human brain, these 3 R-models develop a complete functioning brain with numerous neurodevelopmental processes overlapping with human brain development. Importantly, the main signalling pathways relating to (neuro)development, metabolism and growth are highly conserved in these models. We propose the use of whole model organisms specifically zebrafish and C. elegans for DNT relevant endpoints.

The impact of purine nucleosides on neuroplasticity in the adult brain.

Neuroplasticity refers to the nervous system's ability to adapt and reorganize its cell structures and neuronal networks in response to internal and external stimuli. In adults, this process involves neurogenesis, synaptogenesis, and synaptic and neurochemical plasticity. Several studies have reported the significant impact of the purinergic system on neuroplasticity modulation. And, there is considerable evidence supporting the role of purine nucleosides, such as adenosine, inosine, and guanosine, in this process. This review presents extensive research on how these nucleosides enhance the neuroplasticity of the adult central nervous system, particularly in response to damage. The mechanisms through which these nucleosides exert their effects involve complex interactions with various receptors and signaling pathways. Adenosine's influence on neurogenesis involves interactions with adenosine receptors, specifically A1R and A2AR. A1R activation appears to inhibit neuronal differentiation and promote astrogliogenesis, while A2AR activation supports neurogenesis, neuritogenesis, and synaptic plasticity. Inosine and guanosine positively impact cell proliferation, neurogenesis, and neuritogenesis. Inosine seems to modulate extracellular adenosine levels, and guanosine might act through interactions between purinergic and glutamatergic systems. Additionally, the review discusses the potential therapeutic implications of purinergic signaling in neurodegenerative and neuropsychiatric diseases, emphasizing the importance of these nucleosides in the neuroplasticity of brain function and recovery.

Biomimetic Fibers Derived from an Equidistant Micropillar Platform Dictate Osteocyte Fate via Mechanoreception.

It is a big challenge to design a biomimetic physical microenvironment with greater similarity to in vivo tissue to observe real cell behaviors. We established a novel cell culture platform based on patterned equidistant micropillars with stiff and soft stiffnesses to mimic the changes that happened in the transition from normal to osteoporotic disease. We first demonstrated that the soft micropillar substrate decreased osteocyte synaptogenesis through synaptogyrin 1 and that this decrease was accompanied by impairment of cell mechanoperception and a decrease in cellular cytoskeletal rearrangement. We then found that the soft equidistant micropillar substrate reduced the osteocyte synaptogenesis mainly via the inactivation of Erk/MAPK signaling. We finally found that soft micropillar substrate-mediated synaptogenesis impacted the cell-to-cell communication and matrix mineralization of osteocytes. Taken together, this study provides evidence of cellular mechanical responses that are much more similar to those of real osteocytes at the bone tissue level.

Structure and evolution of neuronal wiring receptors and ligands.

One of the fundamental properties of a neuronal circuit is the map of its connections. The cellular and developmental processes that allow for the growth of axons and dendrites, selection of synaptic targets, and formation of functional synapses use neuronal surface receptors and their interactions with other surface receptors, secreted ligands, and matrix molecules. Spatiotemporal regulation of the expression of these receptors and cues allows for specificity in the developmental pathways that wire stereotyped circuits. The families of molecules controlling axon guidance and synapse formation are generally conserved across animals, with some important exceptions, which have consequences for neuronal connectivity. Here, we summarize the distribution of such molecules across multiple taxa, with a focus on model organisms, evolutionary processes that led to the multitude of such molecules, and functional consequences for the diversification or loss of these receptors.