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The apical dendrite of a cortical projection neuron (CPN) is generated from the leading process of the migrating neuron as the neuron completes migration. This transformation occurs in the cortical marginal zone (MZ), a layer that contains the Cajal-Retzius neurons and their axonal projections. Cajal-Retzius neurons (CRNs) are well known for their critical role in secreting Reelin, a glycoprotein that controls dendritogenesis and cell positioning in many regions of the developing brain. In this study, we examine the possibility that CRNs in the MZ may provide additional signals to arriving CPNs, that may promote the maturation of CPNs and thus shape the development of the cortex. We use whole embryonic hemisphere explants and multiphoton microscopy to confirm that CRNs display intracellular calcium transients of <1-min duration and high amplitude during early corticogenesis. In contrast, developing CPNs do not show high-amplitude calcium transients, but instead show a steady increase in intracellular calcium that begins at the time of dendritic initiation, when the leading process of the migrating CPN is encountering the MZ. The possible existence of CRN to CPN communication was revealed by the application of veratridine, a sodium channel activator, which has been shown to preferentially stimulate more mature cells in the MZ at an early developmental time. Surprisingly, veratridine application also triggers large calcium transients in CPNs, which can be partially blocked by a cocktail of antagonists that block glutamate and glycine receptor activation. These findings outline a model in which CRN spontaneous activity triggers the release of glutamate and glycine, neurotransmitters that can trigger intracellular calcium elevations in CPNs. These elevations begin as CPNs initiate dendritogenesis and continue as waves in the post-migratory cells. Moreover, we show that the pharmacological blockade of glutamatergic signaling disrupts migration, while forced expression of a bacterial voltage-gated calcium channel (CavMr) in the migrating neurons promotes dendritic growth and migration arrest. The identification of CRN to CPN signaling during early development provides insight into the observation that many autism-linked genes encode synaptic proteins that, paradoxically, are expressed in the developing cortex well before the appearance of synapses and the establishment of functional circuits. [ABSTRACT FROM AUTHOR]

BackgroundProper patterning of dendritic and axonal arbors is a critical step in the formation of functional neuronal circuits. Developing circuits rely on an array of molecular cues to shape arbor morphology, but the underlying mechanisms guiding the structural formation and interconnectivity of pre- and postsynaptic arbors in real time remain unclear. Here we explore how Down syndrome cell adhesion molecule (DSCAM) differentially shapes the dendritic morphology of central neurons and their presynaptic retinal ganglion cell (RGC) axons in the developing vertebrate visual system.MethodsThe cell-autonomous role of DSCAM, in tectal neurons and in RGCs, was examined using targeted single-cell knockdown and overexpression approaches in developing Xenopus laevis tadpoles. Axonal arbors of RGCs and dendritic arbors of tectal neurons were visualized using real-time in vivo confocal microscopy imaging over the course of 3 days.ResultsIn the Xenopus visual system, DSCAM immunoreactivity is present in RGCs, cells in the optic tectum and the tectal neuropil at the time retinotectal synaptic connections are made. Downregulating DSCAM in tectal neurons significantly increased dendritic growth and branching rates while inducing dendrites to take on tortuous paths. Overexpression of DSCAM, in contrast, reduced dendritic branching and growth rate. Functional deficits mediated by tectal DSCAM knockdown were examined using visually guided behavioral assays in swimming tadpoles, revealing irregular behavioral responses to visual stimulus. Functional deficits in visual behavior also corresponded with changes in VGLUT/VGAT expression, markers of excitatory and inhibitory transmission, in the tectum. Conversely, single-cell DSCAM knockdown in the retina revealed that RGC axon arborization at the target is influenced by DSCAM, where axons grew at a slower rate and remained relatively simple. In the retina, dendritic arbors of RGCs were not affected by the reduction of DSCAM expression.ConclusionsTogether, our observations implicate DSCAM in the control of both pre- and postsynaptic structural and functional connectivity in the developing retinotectal circuit, where it primarily acts as a neuronal brake to limit and guide postsynaptic dendrite growth of tectal neurons while it also facilitates arborization of presynaptic RGC axons cell autonomously.

Microtubules (MTs) are dynamic components of the cell cytoskeleton involved in several cellular functions, such as structural support, migration and intracellular trafficking. Despite their high similarity, MTs have functional heterogeneity that is generated by the incorporation into the MT lattice of different tubulin gene products and by their post-translational modifications (PTMs). Such regulations, besides modulating the tubulin composition of MTs, create on their surface a "biochemical code" that is translated, through the action of protein effectors, into specific MT-based functions. This code, known as "tubulin code", plays an important role in neuronal cells, whose highly specialized morphologies and activities depend on the correct functioning of the MT cytoskeleton and on its interplay with a myriad of MT-interacting proteins. In recent years, a growing number of mutations in genes encoding for tubulins, MT-interacting proteins and enzymes that post-translationally modify MTs, which are the main players of the tubulin code, have been linked to neurodegenerative processes or abnormalities in neural migration, differentiation and connectivity. Nevertheless, the exact molecular mechanisms through which the cell writes and, downstream, MT-interacting proteins decipher the tubulin code are still largely uncharted. The purpose of this review is to describe the molecular determinants and the readout mechanisms of the tubulin code, and briefly elucidate how they coordinate MT behavior during critical neuronal events, such as neuron migration, maturation and axonal transport. [ABSTRACT FROM AUTHOR]

Electrical activity is considered a key driver for the neurochemical and morphological maturation of neurons and the formation of neuronal networks. Designer receptors exclusively activated by designer drugs (DREADDs) are tools for controlling neuronal activity at the single cell level by triggering specific G protein signaling. Our objective was to investigate if prolonged silencing of differentiating cortical neurons can influence dendritic and axonal maturation. The DREADD hM4Di couples to Gi/o signaling and evokes hyperpolarization via GIRK channels. HM4Di was biolistically transfected into neurons in organotypic slice cultures of rat visual cortex, and activated by clozapine-Noxide (CNO) dissolved in H2O; controls expressed hM4Di, but were mock-stimulated with H2O. Neurons were analyzed after treatment for two postnatal time periods, DIV 5-10 and 10-20. We found that CNO treatment delays the maturation of apical dendrites of L2/3 pyramidal cells. Further, the number of collaterals arising from the main axon was significantly lower, as was the number of bouton terminaux along pyramidal cell and basket cell axons. The dendritic maturation of L5/6 pyramidal cells and of multipolar interneurons (basket cells and bitufted cells) was not altered by CNO treatment. Returning CNO-treated cultures to CNO-free medium for 7 days was sufficient to recover dendritic and axonal complexity. Our findings add to the view that activity is a key driver in particular of postnatal L2/3 pyramidal cell maturation. Our results further suggest that inhibitory G protein signaling may represent a factor balancing the strong driving force of neurotrophic factors, electrical activity and calcium signaling. [ABSTRACT FROM AUTHOR]

Many membrane proteins are highly enriched in either dendrites or axons. This non-uniform distribution is a critical feature of neuronal polarity and underlies neuronal function. The molecular mechanisms responsible for polarized distribution of membrane proteins has been studied for some time and many answers have emerged. A less well studied feature of neurons is that organelles are also frequently non-uniformly distributed. For instance, EEA1-positive early endosomes are somatodendritic whereas synaptic vesicles are axonal. In addition, some organelles are present in both axons and dendrites, but not distributed uniformly along the processes. One well known example are lysosomes which are abundant in the soma and proximal dendrite, but sparse in the distal dendrite and the distal axon. The mechanisms that determine the spatial distribution of organelles along dendrites are only starting to be studied. In this review, we will discuss the cell biological mechanisms of how the distribution of diverse sets of endosomes along the proximal-distal axis of dendrites might be regulated. In particular, we will focus on the regulation of bulk homeostatic mechanisms as opposed to local regulation. We posit that immature dendrites regulate organelle motility differently from mature dendrites in order to spatially organize dendrite growth, branching and sculpting. [Display omitted] • Different endosome subtypes display nonuniform distribution along growing dendrites. • Immature and mature dendrites differ in how they regulate organelle positioning. • Local changes in organelle motility might contribute to dendrite differentiation. [ABSTRACT FROM AUTHOR]

The prenatal period of cortical development is important for the establishment of neural circuitry and functional connectivity of the brain; however, the molecular mechanisms underlying this process remain unclear. Here we report that disruption of the actin-cytoskeletal network in the developing mouse prefrontal cortex alters dendritic morphogenesis and synapse formation, leading to enhanced formation of fear-related memory in adulthood. These effects are mediated by a brain-enriched microRNA, miR-9, through its negative regulation of diaphanous homologous protein 1 (Diap1), a key organizer of the actin cytoskeletal assembly. Our findings not only revealed important regulation of dendritogenesis and synaptogenesis during early brain development but also demonstrated a tight link between these early developmental events and cognitive functions later in life.

The prenatal period of cortical development is important for the establishment of neural circuitry and functional connectivity of the brain; however, the molecular mechanisms underlying this process remain unclear. Here we report that disruption of the actin-cytoskeletal network in the developing mouse prefrontal cortex alters dendritic morphogenesis and synapse formation, leading to enhanced formation of fear-related memory in adulthood. These effects are mediated by a brain-enriched microRNA, miR-9, through its negative regulation of diaphanous homologous protein 1 (Diap1), a key organizer of the actin cytoskeletal assembly. Our findings not only revealed important regulation of dendritogenesis and synaptogenesis during early brain development but also demonstrated a tight link between these early developmental events and cognitive functions later in life.

Electrical activity is considered a key driver for the neurochemical and morphological maturation of neurons and the formation of neuronal networks. Designer receptors exclusively activated by designer drugs (DREADDs) are tools for controlling neuronal activity at the single cell level by triggering specific G protein signaling. Our objective was to investigate if prolonged silencing of differentiating cortical neurons can influence dendritic and axonal maturation. The DREADD hM4Di couples to Gi/o signaling and evokes hyperpolarization via GIRK channels. HM4Di was biolistically transfected into neurons in organotypic slice cultures of rat visual cortex, and activated by clozapine-N-oxide (CNO) dissolved in H2O; controls expressed hM4Di, but were mock-stimulated with H2O. Neurons were analyzed after treatment for two postnatal time periods, DIV 5-10 and 10-20. We found that CNO treatment delays the maturation of apical dendrites of L2/3 pyramidal cells. Further, the number of collaterals arising from the main axon was significantly lower, as was the number of bouton terminaux along pyramidal cell and basket cell axons. The dendritic maturation of L5/6 pyramidal cells and of multipolar interneurons (basket cells and bitufted cells) was not altered by CNO treatment. Returning CNO-treated cultures to CNO-free medium for 7 days was sufficient to recover dendritic and axonal complexity. Our findings add to the view that activity is a key driver in particular of postnatal L2/3 pyramidal cell maturation. Our results further suggest that inhibitory G protein signaling may represent a factor balancing the strong driving force of neurotrophic factors, electrical activity and calcium signaling.

BackgroundNetrins are a family of extracellular proteins that function as chemotropic guidance cues for migrating cells and axons during neural development. In the visual system, netrin-1 has been shown to play a key role in retinal ganglion cell (RGC) axon growth and branching at the target, where presynaptic RGC axons form partnerships with the dendrites of tectal neurons. However, the signals that guide the connections between RGC axons and their postsynaptic partners are yet unknown. Here, we explored dynamic cellular mechanisms by which netrin-1 influences visual circuit formation, particularly those that impact postsynaptic neuronal morphology and connectivity during retinotectal wiring.ResultsTime-lapse in vivo imaging of individual Xenopus laevis optic tectal neurons co-expressing tdTomato and PSD95-GFP revealed rapid remodeling and reorganization of dendritic arbors following acute manipulations in netrin-1 levels. Effects of altered netrin signaling on developing dendritic arbors of tectal neurons were distinct from its effects on presynaptic RGC axons. Within 4 h of treatment, tectal injection of recombinant netrin-1 or sequestration of endogenous netrin with an UNC-5 receptor ectodomain induced significant changes in the directionality and orientation of dendrite growth and in the maintenance of already established dendrites, demonstrating that relative levels of netrin are important for these functions. In contrast, altering DCC-mediated netrin signaling with function-blocking antibodies induced postsynaptic specialization remodeling and changed growth directionality of already established dendrites. Reducing netrin signaling also decreased avoidance behavior in a visually guided task, suggesting that netrin is essential for emergent visual system function.ConclusionsThese in vivo findings together with the patterns of expression of netrin and its receptors reveal an important role for netrin in the early growth and guidance of vertebrate central neuron dendritic arbors. Collectively, our studies indicate that netrin shapes both pre- and postsynaptic arbor morphology directly and in multiple ways at stages critical for functional visual system development.

Brain circuit development involves tremendous structural formation and rearrangement of dendrites, axons, and the synaptic connections between them. Direct studies of neuronal morphogenesis are now possible through recent developments in multiple technologies, including single-neuron labeling, time-lapse imaging in intact tissues, and 4D rendering software capable of tracking neural growth over periods spanning minutes to days. These methods allow detailed quantification of structural changes of neurons over time, called dynamic morphometrics, providing new insights into fundamental growth patterns, underlying molecular mechanisms, and the intertwined influences of external factors, including neural activity, and intrinsic genetic programs. Here, we review the methods of dynamic morphometrics sampling and analyses, focusing on their applications to studies of activity-driven dendritogenesis in vertebrate systems. Advances in single-cell labeling, rapid time-lapse imaging in awake developing vertebrate brains, and software for deep feature extraction from 4D data sets now allow rich quantification of neuronal growth. Tracking growth of individual neurons has revealed mechanisms of dendritogenesis not accessible from static images of different neurons at multiple maturational stages. Multiple independent rapid neurite growth behaviors, influenced by intrinsic genetic programs and extrinsic factors, culminate in dendritic arbors with both individual-neuron and neuron-type specific patterning. Dynamic morphometrics reveals that synapse formation and maturation stabilize growing dendrites. Dual in vivo imaging of neurite growth and neural activity reveals an intertwined relationship between synaptogenesis and dendritogenesis, allowing sensory experience to sculpt both dendritic arbor growth and receptive field tuning. [ABSTRACT FROM AUTHOR]

Fetal alcohol syndrome (FAS) is characterized by disrupted fetal brain development and postnatal cognitive impairment. The targets of alcohol are diverse, and it is not clear whether there are common underlying molecular mechanisms producing these disruptions. Prior work established that acute ethanol exposure causes a transient increase in tyrosine phosphorylation of multiple proteins in cultured embryonic cortical cells. In this study, we show that a similar tyrosine phosphorylation transient occurs in the fetal brain after maternal dosing with ethanol. Using phospho-specific antibodies and immunohistochemistry, we mapped regions of highest tyrosine phosphorylation in the fetal cerebral cortex and found that areas of dendritic and axonal growth showed elevated tyrosine phosphorylation 10 min after maternal ethanol exposure. These were also areas of Src expression and Src family kinase (SFK) activation loop phosphorylation (pY416) expression. Importantly, maternal pretreatment with the SFK inhibitor dasatinib completely prevents both the pY416 increase and the tyrosine phosphorylation response. The phosphorylation response was observed in the perisomatic region and neurites of immature migrating and differentiating primary neurons. Importantly, the initial phosphotyrosine transient (~ 30 min) targets both Src and Dab1, two critical elements in Reelin signaling, a pathway required for normal cortical development. This initial phosphorylation response is followed by sustained reduction in Ser3 phosphorylation of n-cofilin, a critical actin severing protein and an identified downstream effector of Reelin signaling. This biochemical disruption is associated with sustained reduction of F-actin content and disrupted Golgi apparatus morphology in developing cortical neurons. The finding outlines a model in which the initial activation of SFKs by ethanol has the potential to disrupt multiple developmentally important signaling systems for several hours after maternal exposure. [ABSTRACT FROM AUTHOR]

Autism spectrum disorders and Williams syndrome exhibit quite opposite features in the social domain, but also share some common underlying behavioral and cognitive deficits. It is not clear, however, which genes account for the attested differences (and similarities) in the socio-cognitive domain. In this article, we adopted a comparative molecular approach and looked for genes that might be differentially (or similarly) regulated in the blood of subjects with these two conditions. We found a significant overlap between differentially expressed genes compared to neurotypical controls, with most of them exhibiting a similar trend in both conditions, but with genes being more dysregulated in Williams syndrome than in autism spectrum disorders. These genes are involved in aspects of brain development and function (particularly dendritogenesis) and are expressed in brain areas (particularly the cerebellum, the thalamus, and the striatum) of relevance for the autism spectrum disorder and the Williams syndrome etiopathogenesis. Autism spectrum disorders and Williams syndrome are complex cognitive conditions exhibiting quite opposite features in the social domain: whereas people with autism spectrum disorders are mostly hyposocial, subjects with Williams syndrome are usually reported as hypersocial. At the same time, autism spectrum disorders and Williams syndrome share some common underlying behavioral and cognitive deficits. It is not clear, however, which genes account for the attested differences (and similarities) in the socio-cognitive domain. In this article, we adopted a comparative molecular approach and looked for genes that might be differentially (or similarly) regulated in the blood of people with these conditions. We found a significant overlap between genes dysregulated in the blood of patients compared to neurotypical controls, with most of them being upregulated or, in some cases, downregulated. Still, genes with similar expression trends can exhibit quantitative differences between conditions, with most of them being more dysregulated in Williams syndrome than in autism spectrum disorders. Differentially expressed genes are involved in aspects of brain development and function (particularly dendritogenesis) and are expressed in brain areas (particularly the cerebellum, the thalamus, and the striatum) of relevance for the autism spectrum disorder and the Williams syndrome etiopathogenesis. Overall, these genes emerge as promising candidates for the similarities and differences between the autism spectrum disorder and the Williams syndrome socio-cognitive profiles. [ABSTRACT FROM AUTHOR]

• In newborn adult hippocampal neurons, FGFR signaling through FRS is required to promote proximal dendrite length. • In contrast, FGFRs are required to inhibit distal dendrite length through PLCγ in newborn adult hippocampal neurons. • FGFRs through as yet unidentified mediators promote or counterbalance inhibition of dendrite length by FGFR-PLCγ. Fibroblast Growth Factor Receptors (FGFRs) play crucial roles in promoting dendrite growth and branching during development. In mice, three FGFR genes, Fgfr1 , Fgfr2 , and Fgfr3 , remain expressed in the adult neurogenic niche of the hippocampal dentate gyrus. However, the function of FGFRs in the dendritic maturation of adult-born neurons remains largely unexplored. Here, using conditional alleles of Fgfr1, Fgfr2, and Fgfr3 as well as Fgfr1 alleles lacking binding sites for Phospholipase-Cγ (PLCγ) and FGF Receptor Substrate (FRS) proteins, we test the requirement for FGFRs in dendritogenesis of adult-born granule cells. We find that deleting all three receptors results in a small decrease in proximal dendrite elaboration. In contrast, specifically mutating Tyr766 in FGFR1 (a PLCγ binding site) in an Fgfr2;Fgfr3 deficient background results in a dramatic increase of overall dendrite elaboration, while blocking FGFR1-FRS signaling causes proximal dendrite deficits and, to a lesser extent than Tyr766 mutants, increases distal dendrite elaboration. These findings reveal unexpectedly complex roles for FGFRs and their intracellular mediators in regulating proximal and distal dendrite elaboration, with the most notable role in suppressing distal elaboration through the PLCγbinding site. [ABSTRACT FROM AUTHOR]

Dysfunctional dendritic arborization is a key feature of many developmental neurological disorders. Across various human brain regions, basal dendritic complexity is known to increase along a caudal-to-rostral gradient. We recently discovered that basal dendritic complexity of layer II/III cortical pyramidal neurons in the mouse increases along a caudomedial-to-rostrolateral gradient spanning multiple regions, but at the time, no molecules were known to regulate that exquisite pattern. Integrin subunits have been implicated in dendritic development, and the subunit with the strongest associations with autism spectrum disorder and intellectual disability is integrin β3 (Itgb3). In mice, global knockout of Itgb3 leads to autistic-like neuroanatomy and behavior. Here, we tested the hypothesis that Itgb3 is required for increasing dendritic complexity along the recently discovered tangential gradient among layer II/III cortical pyramidal neurons. We targeted a subset of layer II/III cortical pyramidal neurons for Itgb3 loss-of-function via Cre-loxP-mediated excision of Itgb3. We tracked the rostrocaudal and mediolateral position of the targeted neurons and reconstructed their dendritic arbors. In contrast to controls, the basal dendritic complexity of Itgb3 mutant neurons was not related to their cortical position. Basal dendritic complexity of mutant and control neurons differed because of overall changes in branch number across multiple branch orders (primary, secondary, etc.), rather than any changes in the average length at those branch orders. Furthermore, dendritic spine density was related to cortical position in control but not mutant neurons. Thus, the autism susceptibility gene Itgb3 is required for establishing a tangential pattern of basal dendritic complexity among layer II/III cortical pyramidal neurons, suggesting an early role for this molecule in the developing brain. [ABSTRACT FROM AUTHOR]

A battery of genetically encoded calcium indicators (GECIs) with different binding kinetics and calcium affinities was developed over the recent years to permit long-term calcium imaging. GECIs are calcium buffers and therefore, expression of GECIs may interfere with calcium homeostasis and signaling pathways important for neuronal differentiation and survival. Our objective was to investigate if the biolistically induced expression of five commonly used GECIs at two postnatal time points (days 14 and 22–25) could affect the morphological maturation of cortical neurons in organotypic slice cultures of rat visual cortex. Expression of GCaMP3 in both time windows, and of GCaMP5G and TN-XXL in the later time window impaired apical and /or basal dendrite growth of pyramidal neurons. With time, the proportion of GECI transfectants with nuclear filling increased, but an only prolonged expression of TN-XXL caused higher levels of neurodegeneration. In multipolar interneurons, only GCaMP3 evoked a transient growth delay during the early time window. GCaMP6m and GCaMP6m-XC were quite “neuron-friendly.” Since growth-impaired neurons might not have the physiological responses typical of age-matched wildtype neurons the results obtained after prolonged developmental expression of certain GECIs might need to be interpreted with caution.

During development the vast majority of cells that will later compose the mature cerebral cortex undergo extensive migration to reach their final position. In addition to intrinsically distinct migratory behaviors, cells encounter and respond to vastly different microenvironments. These range from axonal tracts to cell-dense matrices, electrically active regions and extracellular matrix components, which may all change overtime. Furthermore, migrating neurons themselves not only adapt to their microenvironment but also modify the local niche through cell-cell contacts, secreted factors and ions. In the radial dimension, the developing cortex is roughly divided into dense progenitor and cortical plate territories, and a less crowded intermediate zone. The cortical plate is bordered by the subplate and the marginal zone, which are populated by neurons with high electrical activity and characterized by sophisticated neuritic ramifications. Neuronal migration is influenced by these boundaries resulting in dramatic changes in migratory behaviors as well as morphology and electrical activity. Modifications in the levels of any of these parameters can lead to alterations and even arrest of migration. Recent work indicates that morphology and electrical activity of migrating neuron are interconnected and the aim of this review is to explore the extent of this connection. We will discuss on one hand how the response of migrating neurons is altered upon modification of their intrinsic electrical properties and whether, on the other hand, the electrical properties of the cellular environment can modify the morphology and electrical activity of migrating cortical neurons.

Abstract Background Proper patterning of dendritic and axonal arbors is a critical step in the formation of functional neuronal circuits. Developing circuits rely on an array of molecular cues to shape arbor morphology, but the underlying mechanisms guiding the structural formation and interconnectivity of pre- and postsynaptic arbors in real time remain unclear. Here we explore how Down syndrome cell adhesion molecule (DSCAM) differentially shapes the dendritic morphology of central neurons and their presynaptic retinal ganglion cell (RGC) axons in the developing vertebrate visual system. Methods The cell-autonomous role of DSCAM, in tectal neurons and in RGCs, was examined using targeted single-cell knockdown and overexpression approaches in developing Xenopus laevis tadpoles. Axonal arbors of RGCs and dendritic arbors of tectal neurons were visualized using real-time in vivo confocal microscopy imaging over the course of 3 days. Results In the Xenopus visual system, DSCAM immunoreactivity is present in RGCs, cells in the optic tectum and the tectal neuropil at the time retinotectal synaptic connections are made. Downregulating DSCAM in tectal neurons significantly increased dendritic growth and branching rates while inducing dendrites to take on tortuous paths. Overexpression of DSCAM, in contrast, reduced dendritic branching and growth rate. Functional deficits mediated by tectal DSCAM knockdown were examined using visually guided behavioral assays in swimming tadpoles, revealing irregular behavioral responses to visual stimulus. Functional deficits in visual behavior also corresponded with changes in VGLUT/VGAT expression, markers of excitatory and inhibitory transmission, in the tectum. Conversely, single-cell DSCAM knockdown in the retina revealed that RGC axon arborization at the target is influenced by DSCAM, where axons grew at a slower rate and remained relatively simple. In the retina, dendritic arbors of RGCs were not affected by the reduction of DSCAM expression. Conclusions Together, our observations implicate DSCAM in the control of both pre- and postsynaptic structural and functional connectivity in the developing retinotectal circuit, where it primarily acts as a neuronal brake to limit and guide postsynaptic dendrite growth of tectal neurons while it also facilitates arborization of presynaptic RGC axons cell autonomously.

A battery of genetically encoded calcium indicators (GECIs) with different binding kinetics and calcium affinities was developed over the recent years to permit long-term calcium imaging. GECIs are calcium buffers and therefore, expression of GECIs may interfere with calcium homeostasis and signaling pathways important for neuronal differentiation and survival. Our objective was to investigate if the biolistically induced expression of five commonly used GECIs at two postnatal time points (days 14 and 22–25) could affect the morphological maturation of cortical neurons in organotypic slice cultures of rat visual cortex. Expression of GCaMP3 in both time windows, and of GCaMP5G and TN-XXL in the later time window impaired apical and /or basal dendrite growth of pyramidal neurons. With time, the proportion of GECI transfectants with nuclear filling increased, but an only prolonged expression of TN-XXL caused higher levels of neurodegeneration. In multipolar interneurons, only GCaMP3 evoked a transient growth delay during the early time window. GCaMP6m and GCaMP6m-XC were quite "neuron-friendly." Since growth-impaired neurons might not have the physiological responses typical of age-matched wildtype neurons the results obtained after prolonged developmental expression of certain GECIs might need to be interpreted with caution. [ABSTRACT FROM AUTHOR]

One of the characteristic features of different classes of neurons that is vital for their proper functioning within neuronal networks is the shape of their dendritic arbors. To properly develop dendritic trees, neurons need to accurately control the intracellular transport of various cellular cargo (e.g., mRNA, proteins, and organelles). Microtubules and motor proteins (e.g., dynein and kinesins) that move along microtubule tracks play an essential role in cargo sorting and transport to the most distal ends of neurons. Equally important are motor adaptors, which may affect motor activity and specify cargo that is transported by the motor. Such transport undergoes very dynamic fine‐tuning in response to changes in the extracellular environment and synaptic transmission. Such regulation is achieved by the phosphorylation of motors, motor adaptors, and cargo, among other mechanisms. This review focuses on the contribution of the dynein–dynactin complex, kinesins, their adaptors, and the phosphorylation of these proteins in the formation of dendritic trees by maturing neurons. We primarily review the effects of the motor activity of these proteins in dendrites on dendritogenesis. We also discuss less anticipated mechanisms that contribute to dendrite growth, such as dynein‐driven axonal transport and non‐motor functions of kinesins. [ABSTRACT FROM AUTHOR]

• Phenytoin interrupts neural cell development due to ion homeostasis disruption. • Phenytoin retards arborization of hippocampal neurons in cell culture. • Phenytoin causes cell body scramble that finally lead to cell death. • Short and thick dendrites in neuron create abnormal morphology in PHT treated cells. Ions are key regulators of the morphogenesis, dendritogenesis and development of neurons therefore drugs that perturb ion homeostasis are associated with high risk of mental retardation, intellectual disability and even abortion of fetus. Phenytoin (PHT) is an antiepileptic drug which regulates ion influx especially Ca2+ and Na+ and widely prescribed to pregnant women suffer from epilepsy. This study aimed to investigate neurodevelopmental features of primary culture of hippocampal cells such as morphology, dendritogenesis, cytotoxicity and cell death in the presence and absence of PHT. Primary culture of hippocampal neurons from neonatal rat was treated by 25 and 50 μg/ml of PHT and morphological development was evaluated during the 14 days. Arborization of neurons during the time was monitored by light microscopy. MTT assay and lactate dehydrogenase (LDH) penetrating test also assessed PHT imposed cytotoxicity. Our results confirmed high dose of PHT could cause excessive cell death in neural cells. PHT exposing causes morphological abnormalities in hippocampal neurons such as shrieked cell body or thick and short dendrite. PHT also prevents branching of dendrites and induces LDH leakage that refers to cytotoxicity. By considering the Ca2+ and Na+ important roles in cell development process, PHT affect neural shape and arborization rate. It could retard neural development and lead neurons to the cell death. PHT is an anticonvulsant that prescribed to pregnant women so could disrupt brain development and increase the risk of mental retardation in newborn children. [ABSTRACT FROM AUTHOR]

ObjectivePerinatal exposure to polychlorinated biphenyls (PCBs) is associated with decreased IQ scores, impaired learning and memory, psychomotor difficulties, and attentional deficits in children. It is postulated that these neuropsychological deficits reflect altered patterns of neuronal connectivity. To test this hypothesis, we examined the effects of developmental PCB exposure on dendritic growth.MethodsRat dams were gavaged from gestational day 6 through postnatal day (PND) 21 with vehicle (corn oil) or the commercial PCB mixture Aroclor 1254 (6 mg/kg/day). Dendritic growth and molecular markers were examined in pups during development.ResultsGolgi analyses of CA1 hippocampal pyramidal neurons and cerebellar Purkinje cells indicated that developmental exposure to PCBs caused a pronounced age-related increase in dendritic growth. Thus, even though dendritic lengths were significantly attenuated in PCB-treated animals at PND22, the rate of growth was accelerated at later ages such that by PND60, dendritic growth was comparable to or even exceeded that observed in vehicle controls. Quantitative reverse transcriptase polymerase chain reaction analyses demonstrated that from PND4 through PND21, PCBs generally increased expression of both spinophilin and RC3/neurogranin mRNA in the hippocampus, cerebellum, and cortex with the most significant increases observed in the cortex.ConclusionsThis study demonstrates that developmental PCB exposure alters the ontogenetic profile of dendritogenesis in critical brain regions, supporting the hypothesis that disruption of neuronal connectivity contributes to neuropsychological deficits seen in exposed children.

The objective of this chapter is to provide an overview of the methods used to investigate the connectivity and structure of the nervous system. These methods allow neuronal cells to be categorized according to their location, shape, and connections to other cells. The Golgi-Cox staining gives a thorough picture of all significant neuronal structures found in the brain that may be distinguished from one another. The most significant characteristic is its three-dimensional integrity since all neuronal structures may be followed continuously from one part to the next. Successions of sections of the brain's neurons are seen with the Golgi stain. The Golgi method is used to serially segment chosen brain parts, and the resulting neurons are produced from those sections., (© 2024. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.)

Dendritogenesis can be impaired by exposure to alcohol, and aspects of this impairment share phenotypic similarities to dendritic defects observed after blockade of the Reelin-Dab1 tyrosine kinase signaling pathway. In this study, we find that 10 min of alcohol exposure (400 mg/dL ethanol) by itself causes an unexpected increase in tyrosine phosphorylation of many proteins including Src and Dab1 that are essential downstream effectors of Reelin signaling. This increase in phosphotyrosine is dose-dependent and blockable by selective inhibitors of Src Family Kinases (SFKs). However, the response is transient, and phosphotyrosine levels return to baseline after 30 min of continuous ethanol exposure, both in vitro and in vivo. During this latter period, Src is inactivated and Reelin application cannot stimulate Dab1 phosphorylation. This suggests that ethanol initially activates but then silences the Reelin-Dab1 signaling pathway by brief activation and then sustained inactivation of SFKs. Time-lapse analyses of dendritic growth dynamics show an overall decrease in growth and branching compared to controls after ethanol-exposure that is similar to that observed with Reelin-deficiency. However, unlike Reelin-signaling disruptions, the dendritic filopodial speeds are decreased after ethanol exposure, and this decrease is associated with sustained dephosphorylation and activation of cofilin, an F-actin severing protein. These findings suggest that persistent Src inactivation coupled to cofilin activation may contribute to the dendritic disruptions observed with fetal alcohol exposure. [ABSTRACT FROM AUTHOR]

During neuronal development, AMPA receptors (AMPARs) and NMDA receptors (NMDARs) are important for neuronal differentiation. Kainate receptors (KARs) are closely related to AMPARs and involved in the regulation of cortical network activity. However, their role for neurite growth and differentiation of cortical neurons is unclear. Here, we used KAR agonists and overexpression of selected KAR subunits and their auxiliary neuropilin and tolloid-like proteins, NETOs, to investigate their influence on dendritic growth and network activity in organotypic cultures of rat visual cortex. Kainate at 500 nM enhanced network activity and promoted development of dendrites in layer II/III pyramidal cells, but not interneurons. GluK2 overexpression promoted dendritic growth in pyramidal cells and interneurons. GluK2 transfectants were highly active and acted as drivers for network activity. GluK1 and NETO1 specifically promoted dendritic growth of interneurons. Our study provides new insights for the roles of KARs and NETOs in the morphological and physiological development of the visual cortex. [ABSTRACT FROM AUTHOR]

Parkinson's disease (PD) is a multisystem disorder, involving several monoaminergic neurotransmitter systems resulting in a broad range of motor and non-motor symptoms. Pathological hallmarks of PD are the loss of dopaminergic neurons and the accumulation of alpha-synuclein, however also being present in the serotonergic raphe nuclei early in the disease course. The dysfunction of the serotonergic system projecting to the hippocampus may contribute to early non-motor symptoms such as anxiety and depression. The adult hippocampal dentate gyrus (DG), a unique niche of the forebrain continuously generating new neurons, may particularly present enhanced susceptibility towards accumulating alpha-synuclein levels. The underlying molecular mechanisms in the context of neuronal maturation and survival of new-born neurons are yet not well understood. To characterize the effects of overexpression of human full-length alpha-synuclein on hippocampal cellular and synaptic plasticity, we used a recently generated BAC alpha-synuclein transgenic rat model showing important features of PD such as widespread and progressive alpha-synuclein aggregation pathology, dopamine loss and age-dependent motor decline. At the age of four months, thus prior to the occurrence of the motor phenotype, we observed a profoundly impaired dendritogenesis of neuroblasts in the hippocampal DG resulting in severely reduced survival of adult new-born neurons. Diminished neurogenesis concurred with a serotonergic deficit in the hippocampus as defined by reduced levels of serotonin (5-HT) 1B receptor, decreased 5-HT neurotransmitter levels, and a loss of serotonergic nerve terminals innervating the DG/CA3 subfield, while the number of serotonergic neurons in the raphe nuclei remained unchanged. Moreover, alpha-synuclein overexpression reduced proteins involved in vesicle release, in particular synapsin-1 and Rab3 interacting molecule (RIM3), in conjunction with an altered ultrastructural architecture of hippocampal synapses. Importantly, BAC alpha-synuclein rats showed an early anxiety-like phenotype consisting of reduced exploratory behavior and feeding. Taken together, these findings imply that accumulating alpha-synuclein severely affects hippocampal neurogenesis paralleled by impaired 5-HT neurotransmission prior to the onset of aggregation pathology and overt motor deficits in this transgenic rat model of PD.

The building of the brain is a multistep process that requires the coordinate expression of thousands of genes and an intense nucleocytoplasmic transport of RNA and proteins. This transport is mediated by karyopherins that comprise importins and exportins. Here, we investigated the role of the ß-importin, importin-8 (IPO8) during mouse cerebral corticogenesis as several of its cargoes have been shown to be essential during this process. First, we showed that Ipo8 mRNA is expressed in mouse brain at various embryonic ages with a clear signal in the sub-ventricular/ventricular zone (SVZ/VZ), the cerebral cortical plate (CP) and the ganglionic eminences. We found that acute knockdown of IPO8 in cortical progenitors reduced both their proliferation and cell cycle exit leading to the increase in apical progenitor pool without influencing the number of basal progenitors (BPs). Projection neurons ultimately reached their appropriate cerebral cortical layer, but their dendritogenesis was specifically affected, resulting in neurons with reduced dendrite complexity. IPO8 knockdown also slowed the migration of cortical interneurons. Together, our data demonstrate that IPO8 contribute to the coordination of several critical steps of cerebral cortex development. These results suggest that the impairment of IPO8 function might be associated with some diseases of neuronal migration defects.

microRNAs (miRNAs) have emerged as critical regulators of neuronal dendrite development. Specific precursor (pre-)miRNAs are actively transported to dendrites, but whether this process is regulated by neuronal activity and involved in activity-dependent dendritogenesis is unknown. Here we show that BDNF, a neurotrophin that is released in response to increased neuronal activity, promotes dendritic accumulation of pre-miR-134. Dendritic accumulation, but not transcription of pre-miR-134, is abrogated by treatment of neurons with the NMDA receptor (NMDAR) antagonist APV. Furthermore, APV interferes with BDNF-mediated repression of the known miR-134 target Pumilio 2 (Pum2) in a miR-134 binding site-specific manner. At the functional level, both APV treatment and knockdown of the pre-miR-134 transport protein DHX36 antagonize BDNF-induced dendritogenesis. These effects are likely mediated by reduced dendritic miR-134 activity, since both transfection of a synthetic miR-134 duplex or of a dendritically targeted pre-miR-134-181a chimera rescues BDNF-dependent dendritogenesis in the presence of APV. In conclusion, we have identified a novel NMDAR-dependent mechanism involved in the activity-dependent control of miRNA function during neuronal development.

Dendritic outgrowth in immature neurons is enhanced by neuronal activity and is considered one of the mechanisms of neural circuit optimization. It is known that calcium signals affect transcriptional regulation and cytoskeletal remodeling necessary for dendritic outgrowth. Here, we demonstrate that activity-dependent calcium signaling also controls mitochondrial homeostasis via AMP-activated protein kinase (AMPK) in growing dendrites of differentiating mouse hippocampal neurons. We found that the inhibition of neuronal activity induced dendritic hypotrophy with abnormally elongated mitochondria. In growing dendrites, AMPK is activated by neuronal activity and dynamically oscillates in synchrony with calcium spikes, and this AMPK oscillation was inhibited by CaMKK2 knockdown. AMPK activation led to phosphorylation of MFF and ULK1, which initiate mitochondrial fission and mitophagy, respectively. Dendritic mitochondria in AMPK-depleted neurons exhibited impaired fission and mitophagy and displayed multiple signs of dysfunction. Genetic inhibition of fission led to dendritic hypoplasia that was reminiscent of AMPK-deficient neurons. Thus, AMPK activity is finely tuned by the calcium-CaMKK2 pathway and regulates mitochondrial homeostasis by facilitating removal of damaged components of mitochondria in growing neurons during normal brain development., Competing Interests: Competing interests The authors declare no competing or financial interests., (© 2023. Published by The Company of Biologists Ltd.)

Polarity is a fundamental feature of cells. Protein complexes, including the PAR3-PAR6-aPKC complex, have conserved roles in establishing polarity across a number of eukaryotic cell types. In neurons, polarity is evident as distinct axonal versus dendritic domains. The PAR3, PAR6, and aPKC proteins also play important roles in neuronal polarization. During this process, either aPKC kinase activity, the assembly of the PAR3-PAR6-aPKC complex or the localization of these proteins is regulated downstream of a number of signaling pathways. In turn, the PAR3, PAR6, and aPKC proteins control various effector molecules to establish neuronal polarity. Herein, we discuss the many signaling mechanisms and effector functions that have been linked to PAR3, PAR6, and aPKC during the establishment of neuronal polarity. [ABSTRACT FROM AUTHOR]

The building of the brain is a multistep process that requires the coordinate expression of thousands of genes and an intense nucleocytoplasmic transport of RNA and proteins. This transport is mediated by karyopherins that comprise importins and exportins. Here, we investigated the role of the ß-importin, importin-8 (IPO8) during mouse cerebral corticogenesis as several of its cargoes have been shown to be essential during this process. First, we showed that Ipo8 mRNA is expressed in mouse brain at various embryonic ages with a clear signal in the sub-ventricular/ventricular zone (SVZ/VZ), the cerebral cortical plate (CP) and the ganglionic eminences. We found that acute knockdown of IPO8 in cortical progenitors reduced both their proliferation and cell cycle exit leading to the increase in apical progenitor pool without influencing the number of basal progenitors (BPs). Projection neurons ultimately reached their appropriate cerebral cortical layer, but their dendritogenesis was specifically affected, resulting in neurons with reduced dendrite complexity. IPO8 knockdown also slowed the migration of cortical interneurons. Together, our data demonstrate that IPO8 contribute to the coordination of several critical steps of cerebral cortex development. These results suggest that the impairment of IPO8 function might be associated with some diseases of neuronal migration defects. [ABSTRACT FROM AUTHOR]

microRNAs (miRNAs) have emerged as critical regulators of neuronal dendrite development. Specific precursor (pre-)miRNAs are actively transported to dendrites, but whether this process is regulated by neuronal activity and involved in activitydependent dendritogenesis is unknown. Here we show that BDNF, a neurotrophin that is released in response to increased neuronal activity, promotes dendritic accumulation of pre-miR-134. Dendritic accumulation, but not transcription of pre-miR-134, is abrogated by treatment of neurons with the NMDA receptor (NMDAR) antagonist APV. Furthermore, APV interferes with BDNF-mediated repression of the known miR-134 target Pumilio 2 (Pum2) in a miR-134 binding site-specific manner. At the functional level, both APV treatment and knockdown of the pre-miR-134 transport protein DHX36 antagonize BDNF-induced dendritogenesis. These effects are likely mediated by reduced dendritic miR-134 activity, since both transfection of a synthetic miR-134 duplex or of a dendritically targeted pre-miR-134-181a chimera rescues BDNF-dependent dendritogenesis in the presence of APV. In conclusion, we have identified a novel NMDAR-dependent mechanism involved in the activity-dependent control of miRNA function during neuronal development. [ABSTRACT FROM AUTHOR]

The low-density lipoprotein receptor-related protein 4 (LRP4) is essential in muscle fibers for the establishment of the neuromuscular junction. Here, we show that LRP4 is also expressed by embryonic cortical and hippocampal neurons, and that downregulation of LRP4 in these neurons causes a reduction in density of synapses and number of primary dendrites. Accordingly, overexpression of LRP4 in cultured neurons had the opposite effect inducing more but shorter primary dendrites with an increased number of spines. Transsynaptic tracing mediated by rabies virus revealed a reduced number of neurons presynaptic to the cortical neurons in which LRP4 was knocked down. Moreover, neuron-specific knockdown of LRP4 by in utero electroporation of LRP4 miRNA in vivo also resulted in neurons with fewer primary dendrites and a lower density of spines in the developing cortex and hippocampus. Collectively, our results demonstrate an essential and novel role of neuronal LRP4 in dendritic development and synaptogenesis in the CNS. [ABSTRACT FROM AUTHOR]

Fibroblast growth factors (FGFs) and FGF receptors (FGFRs) are known for their potent effects on cell proliferation/differentiation and cortical patterning in the developing brain. However, little is known regarding FGFs/FGFRs' roles in cortical circuit formation. Here we show that Fgfr1/2/3 and Fgf7/9/10/22 mRNAs are expressed in the developing primary somatosensory (S1) barrel cortex. Barrel cortex layer IV spiny stellate cells (bSCs) are the primary recipients of ascending sensory information via thalamocortical axons (TCAs). Detail quantification revealed distinctive phases for bSC dendritogenesis: orienting dendrites towards TCAs, adding de-novo dendritic segments, and elongating dendritic length, while maintaining dendritic patterns. Deleting Fgfr1/2/3 in bSCs had minimal impact on dendritic polarity but transiently increased the number of dendritic segments. However, six days later, FGFR1/2/3 LOF reduced dendritic branch numbers. These data suggest that FGF-FGFR has a role in stabilizing dendritic patterning. Depolarization of cultured mouse cortical neurons upregulated the levels of several Fgf/Fgfr mRNAs within two hours. In vivo, within 6 hours of systemic kainic acid administration at postnatal day 6, mRNA levels of Fgf9, Fgf10, Fgfr2c, and Fgfr3b in S1 cortices were enhanced and this was accompanied by exuberant dendritogenesis of bSCs by 24 hrs. Deleting Fgfr1/2/3 abolished kainic acid-induced bSC dendritic overgrowth. Finally, FGF9/10 gain-of-function also resulted in extensive dendritogenesis. Taken together, our data suggest that FGF/FGFR can be regulated by glutamate transmission to modulate/stabilize bSC dendritic complexity. Both male and female mice were used for our study. [ABSTRACT FROM AUTHOR]

Local translation of synaptic mRNAs is an important process related to key aspects of central nervous system development and physiology, including dendritogenesis, axonal growth cone morphology and guidance and synaptic plasticity. Accordingly, local translation is compromised in several intellectual disabilities, including Fragile X syndrome, tuberous sclerosis and Down syndrome. Down Syndrome Cell Adhesion Molecule (DSCAM) is a gene with ascribed functions in neuronal wiring that belongs to the Down Syndrome Critical Region (DSCR) of chromosome 21. In this review, we discuss the evidence for local translation of theDSCAMmRNA in dendrites and axonal growth cones of mouse hippocampal neurons, as well as the possible functions of the locally translated DSCAM protein. [ABSTRACT FROM PUBLISHER]

Thyroid hormone 3,3′,5-Triiodo-L-thyronine (T3) is essential for proper brain development. Perinatal loss of T3 causes severe growth defects in neurons and glia, including strong inhibition of dendrite formation in Purkinje cells in the cerebellar cortex. Here we show that T3 promotes dendritic outgrowth of Purkinje cells through induction of peroxisome proliferator-activated receptor gamma (PPARγ) co-activator 1α (PGC-1α), a master regulator of mitochondrial biogenesis. PGC-1α expression in Purkinje cells is upregulated during dendritic outgrowth in normal mice, while it is significantly retarded in hypothyroid mice or in cultures depleted of T3. In cultured Purkinje cells, PGC-1α knockdown or molecular perturbation of PGC-1α signaling inhibits enhanced dendritic outgrowth and mitochondrial generation and activation caused by T3 treatment. In contrast, PGC-1α overexpression promotes dendrite extension even in the absence of T3. PGC-1α knockdown also downregulates dendrite formation in Purkinje cells in vivo. Our findings suggest that the growth-promoting activity of T3 is partly mediated by PGC-1α signaling in developing Purkinje cells.

The prenatal period of cortical development is important for the establishment of neural circuitry and functional connectivity of the brain; however, the molecular mechanisms underlying this process remain unclear. Here we report that disruption of the actin–cytoskeletal network in the developing mouse prefrontal cortex alters dendritic morphogenesis and synapse formation, leading to enhanced formation of fear-related memory in adulthood. These effects are mediated by a brain-enriched microRNA, miR-9, through its negative regulation of diaphanous homologous protein 1 (Diap1), a key organizer of the actin cytoskeletal assembly. Our findings not only revealed important regulation of dendritogenesis and synaptogenesis during early brain development but also demonstrated a tight link between these early developmental events and cognitive functions later in life. [ABSTRACT FROM AUTHOR]

Vascular endothelial growth factor (VEGF) is well known as the growth factor with wide-ranging functions even in the central nervous system (CNS). Presently, most attention is given to the investigation of its role in neuronal protection, growth and maturation processes, whereby most effects are mediated through VEGF receptor 2 (VEGFR-2). The purpose of our current study is to provide new insights into the impact of VEGF on immature and mature Purkinje cells (PCs) in accordance with maturity and related receptor expression. Therefore, to expand our knowledge of VEGF effects in PCs development and associated VEGFR-2 expression, we used cultivated organotypic cerebellar slice cultures in immunohistochemical or microinjection studies, followed by confocal laser scanning microscopy (CLSM) and morphometric analysis. Additionally, we incorporated in our study the method of laser microdissection, followed by quantitative polymerase chain reaction (qPCR). For the first time we could show the age-dependent VEGF sensitivity of PCs with the largest promoting effects being on dendritic length and cell soma size in neonatal and juvenile stages. Once mature, PCs were no longer susceptible to VEGF stimulation. Analysis of VEGFR-2 expression revealed its presence in PCs throughout development, which underlined its mediating functions in neuronal cells. [ABSTRACT FROM AUTHOR]

Several neurodevelopmental disorders are associated with increased mTOR activity that results in pathogenic neuronal dysmorphogenesis (i.e., soma and dendrite overgrowth), leading to circuit alterations associated with epilepsy and neurologic disabilities. Although an mTOR analog is approved for the treatment of epilepsy in one of these disorders, it has limited efficacy and is associated with a wide range of side effects. There is a need to develop novel agents for the treatment of mTOR-pathway related disorders. Here, we developed a medium-throughput phenotypic assay to test drug efficacy on neurite morphogenesis of mouse neurons in a hyperactive mTOR condition. Our assay involved in utero electroporation (IUE) of a selective population of cortical pyramidal neurons with a plasmid encoding the constitutively active mTOR activator, Rheb, and tdTomato. Labeled neurons from the somatosensory cortex (SSC) were cultured onto 96-well plates and fixed at various days in vitro or following Torin 1 treatment. Automated systems were used for image acquisition and neuron morphologic measurements. We validated our automated approach using traditional manual methods of neuron morphologic assessment. Both automated and manual analyses showed increased neurite length and complexity over time, and decreased neurite overgrowth and soma size with Torin 1. These data validate the accuracy of our automated approach that takes hours compared with weeks when using traditional manual methods. Taken together, this assay can be scaled to screen 32 compounds simultaneously in two weeks, highlighting its robustness and efficiency for medium-throughput screening of candidate therapeutics on a defined population of wild-type or diseased neurons., Competing Interests: The authors declare no competing financial interests., (Copyright © 2023 Sokolov et al.)

Isoprenylation is a posttranslational modification that transfers farnesyl pyrophosphate (FPP) or geranylgeranyl pyrophosphate (GGPP) to cysteine residues of a particular set of proteins, causing their localization to the plasma membrane and other cellular compartments and so rendering them biologically active. Such a modification process, catalyzed by protein prenyltransferase including farnesyltransferase (FT), geranylgeranyltransferase I (GGTI), and geranylgeranyltransferase II (GGTII), is required for the transforming activity of many oncogenic proteins, including some RAS family members. In the past three decades, prenyltransferase has been extensively studied as a promising cancer therapeutic target in vitro, in animal models, and in the clinic. Recently, a growing number of studies suggest that prenyltransferases and the substrates FPP and GGPP also play fundamental roles in nervous system development and brain disorders. However, a systemic review about the advances of prenyltransferases in the field of neuroscience is lacking so far. Herein, we give a brief introduction for the structure and distribution of GGTI and comprehensively updated the recent advances of GGTI in neuronal dendritogenesis/synaptogenesis and in learning/memory-related behavioral performance. More importantly, we discussed the involvement of GGTI and its substrate GGPP in neurodegenerative disorders, such as aging, Alzheimer's disease, multiple sclerosis, and Niemann-Pick disease type C. The role of FT-FPP and GGTII is mentioned as well to compare with GGTI in these physiological and pathological processes. We hope that this systematical review about what we know about GGTI research in the brain can stimulate further studies on the underlying mechanism of GGTI-mediated isoprenylation in the pathogenesis of neurodegenerative and neurodevelopmental disorders. [ABSTRACT FROM AUTHOR]

Inhibitory interneurons in the olfactory bulb are generated continuously throughout life in the subventricular zone and differentiate into periglomerular and granule cells. Neural circuits that undergo reorganization by newborn olfactory bulb interneurons are necessary for odor detection, odor discrimination, olfactory memory, and innate olfactory responses. Although sensory experience has been shown to regulate development in a variety of species and in various structures, including the retina, cortex, and hippocampus, little is known about how sensory experience regulates the dendritic development of newborn olfactory bulb interneurons. Recent studies revealed that the 5T4 oncofetal trophoblast glycoprotein and the neuronal Per/Arnt/Sim domain protein 4 (Npas4) transcription factor regulate dendritic branching and dendritic spine formation, respectively, in olfactory bulb interneurons. Here, we summarize the molecular mechanisms that underlie the sensory input-dependent development of newborn interneurons and the formation of functional neural circuitry in the olfactory bulb.

Iron deficiency (ID), with and without anemia, affects an estimated 2 billion people worldwide. ID is particularly deleterious during early-life brain development, leading to longterm neurological impairments including deficits in hippocampus-mediated learning and memory. Neonatal rats with fetal/neonatal ID anemia (IDA) have shorter hippocampal CA1 apical dendrites with disorganized branching. ID-induced dendritic structural abnormalities persist into adulthood despite normalization of the iron status. However, the specific developmental effects of neuronal iron loss on hippocampal neuron dendrite growth and branching are unknown. Embryonic hippocampal neuron cultures were chronically treated with deferoxamine (DFO, an iron chelator) beginning at 3 days in vitro (DIV). Levels of mRNA for Tfr1 and Slc11a2, iron-responsive genes involved in iron uptake, were significantly elevated in DFO-treated cultures at 11DIV and 18DIV, indicating a degree of neuronal ID similar to that seen in rodent ID models. DFO treatment decreased mRNA levels for genes indexing dendritic and synaptic development (i.e. BdnfVI, Camk2a, Vamp1, Psd95, Cfl1, Pfn1, Pfn2, and Gda) and mitochondrial function (i.e. Ucp2, Pink1, and Cox6a1). At 18DIV, DFO reduced key aspects of energy metabolism including basal respiration, maximal respiration, spare respiratory capacity, ATP production, and glycolytic rate, capacity, and reserve. Sholl analysis revealed a significant decrease in distal dendritic complexity in DFO-treated neurons at both 11DIV and 18DIV. At 11DIV, the length of primary dendrites and the number and length of branches in DFO-treated neurons were reduced. By 18DIV, partial recovery of the dendritic branch number in DFO-treated neurons was counteracted by a significant reduction in the number and length of primary dendrites and the length of branches. Our findings suggest that early neuronal iron loss, at least partially driven through altered mitochondrial function and neuronal energy metabolism, is responsible for the effects of fetal/neonatal ID and IDA on hippocampal neuron dendritic and synaptic maturation. Impairments in these neurodevelopmental processes likely underlie the negative impact of early life ID and IDA on hippocampus-mediated learning and memory. [ABSTRACT FROM AUTHOR]

Glutamate neurotransmission refines synaptic connections to establish the precise neural circuits underlying sensory processing. Deleting metabotropic glutamate receptor 5 (mGluR5) in mice perturbs cortical somatosensory map formation in the primary somatosensory (SI) cortex at both functional and anatomical levels. To examine the cell-autonomous influences of mGluR5 signaling in the morphological and functional development of layer IV spiny stellate glutamatergic neurons receiving sensory input, mGluR5 genetic mosaic mice were generated through in utero electroporation. In the SI cortex of these mosaic brains, we found that most wild-type neurons were located in barrel rings encircling thalamocortical axon (TCA) clusters while mGluR5 knock-out (KO) neurons were placed in the septal area, the cell-sparse region separating barrels. These KO neurons often displayed a symmetrical dendritic morphology with increased dendritic complexity, in contrast to the polarized pattern of wild-type neurons. The dendritic spine density of mGluR5 KO spiny stellate neurons was significantly higher than in wild-type neurons. Whole-cell electrophysiological recordings detected a significant increase in the frequencies of spontaneous and miniature excitatory postsynaptic events in mGluR5 KO neurons compared with neighboring wildtype neurons. Our mosaic analysis provides strong evidence supporting the cell-autonomous influence of mGluR5 signaling on the functional and anatomical development of cortical glutamatergic neurons. Specifically, mGluR5 is required in cortical glutamatergic neurons for the following processes: (1) the placement of cortical glutamatergic neurons close to TCA clusters; (2) the regulation of dendritic complexity and outgrowth toward TCA clusters; (3) spinogenesis; and (4) tuning of excitatory inputs. [ABSTRACT FROM AUTHOR]

Objective Febrile seizures ( FS) are fever-associated convulsions, being the most common seizure disorder in early childhood. A subgroup of these children later develops epilepsy characterized by a hyperexcitable neuronal network in the hippocampus. Hippocampal excitability is regulated by the hippocampal dentate gyrus ( DG) where postnatal neurogenesis occurs. Experimental FS increase the survival of newborn hippocampal dentate granule cells ( DGCs), yet the significance of this neuronal subpopulation to the hippocampal network remains unclear. In the current study, we characterized the temporal maturation and structural integration of these post- FS born DGCs in the DG. Methods Experimental FS were induced in 10-day-old rat pups. The next day, retroviral particles coding for enhanced green fluorescent protein (eGFP) were stereotactically injected in the DG to label newborn cells. Histochemical analyses of eGFP expressing DGCs were performed one, 4, and 8 weeks later and consisted of the following: (1) colocalization with neurodevelopmental markers doublecortin, calretinin, and the mature neuronal marker NeuN; (2) quantification of dendritic complexity; and (3) quantification of spine density and morphology. Results At neither time point were neurodevelopmental markers differently expressed between FS animals and normothermia ( NT) controls. One week after treatment, DGCs from FS animals showed dendrites that were 66% longer than those from NT controls. At 4 and 8 weeks, Sholl analysis of the outer 83% of the molecular layer showed 20-25% more intersections in FS animals than in NT controls (p < 0.01). Although overall spine density was not affected, an increase in mushroom-type spines was observed after 8 weeks. Significance Experimental FS increase dendritic complexity and the number of mushroom-type spines in post- FS born DGCs, demonstrating a more mature phenotype and suggesting increased incoming excitatory information. The consequences of this hyperconnectivity to signal processing in the DG and the output of the hippocampus remain to be studied. [ABSTRACT FROM AUTHOR]