Differentiation Of Sensory Neurons Research Articles

Shallow neural networks trained to detect collisions recover features of visual loom-selective neurons.

Animals have evolved sophisticated visual circuits to solve a vital inference problem: detecting whether or not a visual signal corresponds to an object on a collision course. Such events are detected by specific circuits sensitive to visual looming, or objects increasing in size. Various computational models have been developed for these circuits, but how the collision-detection inference problem itself shapes the computational structures of these circuits remains unknown. Here, inspired by the distinctive structures of LPLC2 neurons in the visual system of Drosophila, we build anatomically-constrained shallow neural network models and train them to identify visual signals that correspond to impending collisions. Surprisingly, the optimization arrives at two distinct, opposing solutions, only one of which matches the actual dendritic weighting of LPLC2 neurons. Both solutions can solve the inference problem with high accuracy when the population size is large enough. The LPLC2-like solutions reproduces experimentally observed LPLC2 neuron responses for many stimuli, and reproduces canonical tuning of loom sensitive neurons, even though the models are never trained on neural data. Thus, LPLC2 neuron properties and tuning are predicted by optimizing an anatomically-constrained neural network to detect impending collisions. More generally, these results illustrate how optimizing inference tasks that are important for an animal's perceptual goals can reveal and explain computational properties of specific sensory neurons.

Differentiation of iPS-Cells into Peripheral Sensory Neurons.

Induced pluripotent stem cells (iPS-cells) have significantly expanded our experimental possibilities, by creating new strategies for the molecular study of human disease and drug development. Treatment of pain has not seen much improvement over the past decade, likely due to species differences in preclinical models. Thus, iPS-cell derived sensory neurons offer a highly welcome translational approach for research and drug development. Although central neuronal differentiation is relatively straightforward, the successful and reliable generation of peripheral neurons requires more complex measures. Here, we describe a small molecule-based protocol for the differentiation of human sensory neurons from iPS-cells which renders functional nociceptor-like cells within several weeks.

HMGB1 released from nociceptors mediates inflammation

Inflammation, the body's primary defensive response system to injury and infection, is triggered by molecular signatures of microbes and tissue injury. These molecules also stimulate specialized sensory neurons, termed nociceptors. Activation of nociceptors mediates inflammation through antidromic release of neuropeptides into infected or injured tissue, producing neurogenic inflammation. Because HMGB1 is an important inflammatory mediator that is synthesized by neurons, we reasoned nociceptor release of HMGB1 might be a component of the neuroinflammatory response. In support of this possibility, we show here that transgenic nociceptors expressing channelrhodopsin-2 (ChR2) directly release HMGB1 in response to light stimulation. Additionally, HMGB1 expression in neurons was silenced by crossing synapsin-Cre (Syn-Cre) mice with floxed HMGB1 mice (HMGB1f/f). When these mice undergo sciatic nerve injury to activate neurogenic inflammation, they are protected from the development of cutaneous inflammation and allodynia as compared to wild-type controls. Syn-Cre/HMGB1fl/fl mice subjected to experimental collagen antibody-induced arthritis, a disease model in which nociceptor-dependent inflammation plays a significant pathological role, are protected from the development of allodynia and joint inflammation. Thus, nociceptor HMGB1 is required to mediate pain and inflammation during sciatic nerve injury and collagen antibody-induced arthritis.

Selective disruption of trigeminal sensory neurogenesis and differentiation in a mouse model of 22q11.2 deletion syndrome.

ABSTRACT22q11.2 Deletion Syndrome (22q11DS) is a neurodevelopmental disorder associated with cranial nerve anomalies and disordered oropharyngeal function, including pediatric dysphagia. Using the LgDel 22q11DS mouse model, we investigated whether sensory neuron differentiation in the trigeminal ganglion (CNgV), which is essential for normal orofacial function, is disrupted. We did not detect changes in cranial placode cell translocation or neural crest migration at early stages of LgDel CNgV development. However, as the ganglion coalesces, proportions of placode-derived LgDel CNgV cells increase relative to neural crest cells. In addition, local aggregation of placode-derived cells increases and aggregation of neural crest-derived cells decreases in LgDel CNgV. This change in cell-cell relationships was accompanied by altered proliferation of placode-derived cells at embryonic day (E)9.5, and premature neurogenesis from neural crest-derived precursors, reflected by an increased frequency of asymmetric neurogenic divisions for neural crest-derived precursors by E10.5. These early differences in LgDel CNgV genesis prefigure changes in sensory neuron differentiation and gene expression by postnatal day 8, when early signs of cranial nerve dysfunction associated with pediatric dysphagia are observed in LgDel mice. Apparently, 22q11 deletion destabilizes CNgV sensory neuron genesis and differentiation by increasing variability in cell-cell interaction, proliferation and sensory neuron differentiation. This early developmental divergence and its consequences may contribute to oropharyngeal dysfunction, including suckling, feeding and swallowing disruptions at birth, and additional orofacial sensory/motor deficits throughout life.

Functional validation of epitope-tagged ATF5 knock-in mice generated by improved genome editing of oviductal nucleic acid delivery (i-GONAD).

Activating transcription factor 5 (ATF5) is a stress-responsive transcription factor that belongs to the cAMP response element-binding protein (CREB)/ATF family, and is essential for the differentiation and survival of sensory neurons in murine olfactory organs. However, the study of associated proteins and target genes for ATF5 has been hampered due to the limited availability of immunoprecipitation-grade ATF5 antibodies. To overcome this issue, we generated hemagglutinin (HA)-tag knock-in mice for ATF5 using CRISPR/Cas9-mediated genome editing with one-step electroporation in oviducts (i-GONAD). ATF5-HA fusion proteins were detected in the nuclei of immature and some mature olfactory and vomeronasal sensory neurons in the main olfactory epithelium and vomeronasal organ, respectively, as endogenous ATF5 proteins were expressed, and some ATF5-HA proteins were found to be phosphorylated. Chromatin immunoprecipitation (ChIP) experiments revealed that ATF5-HA bound to the CCAAT/enhancer-binding protein (C/EBP)-ATF response element site in the promotor region of receptor transporting protein 1 (Rtp1), a chaperone gene responsible for proper olfactory receptor expression. These knock-in mice may be used to examine the expression, localization, and protein-protein/-DNA interactions of endogenous ATF5 and, ultimately, the function of ATF5 in vivo.

Stress and odorant receptor feedback during a critical period after hatching regulates olfactory sensory neuron differentiation in Drosophila.

Here, we reveal that the regulation of Drosophila odorant receptor (OR) expression during the pupal stage is permissive and imprecise. We found that directly after hatching an OR feedback mechanism both directs and refines OR expression. We demonstrate that, as in mice, dLsd1 and Su(var)3-9 balance heterochromatin formation to direct OR expression. We show that the expressed OR induces dLsd1 and Su(var)3-9 expression, linking OR level and possibly function to OR expression. OR expression refinement shows a restricted duration, suggesting that a gene regulatory critical period brings olfactory sensory neuron differentiation to an end. Consistent with a change in differentiation, stress during the critical period represses dLsd1 and Su(var)3-9 expression and makes the early permissive OR expression permanent. This induced permissive gene regulatory state makes OR expression resilient to stress later in life. Hence, during a critical period OR feedback, similar to in mouse OR selection, defines adult OR expression in Drosophila.

Biophysics and Modeling of Mechanotransduction in Neurons: A Review

Mechanosensing is a key feature through which organisms can receive inputs from the environment and convert them into specific functional and behavioral outputs. Mechanosensation occurs in many cells and tissues, regulating a plethora of molecular processes based on the distribution of forces and stresses both at the cell membrane and at the intracellular organelles levels, through complex interactions between cells’ microstructures, cytoskeleton, and extracellular matrix. Although several primary and secondary mechanisms have been shown to contribute to mechanosensation, a fundamental pathway in simple organisms and mammals involves the presence of specialized sensory neurons and the presence of different types of mechanosensitive ion channels on the neuronal cell membrane. In this contribution, we present a review of the main ion channels which have been proven to be significantly involved in mechanotransduction in neurons. Further, we discuss recent studies focused on the biological mechanisms and modeling of mechanosensitive ion channels’ gating, and on mechanotransduction modeling at different scales and levels of details.

SARM1 is required in human derived sensory neurons for injury-induced and neurotoxic axon degeneration

Axonal degeneration contributes to the pathogenesis of many neurodegenerative disorders, motivating efforts to dissect the mechanism of pathological axon loss in order to develop therapies for axonal preservation. SARM1 is a particularly attractive therapeutic target, as it is an inducible NAD+ cleaving enzyme that is required for axon loss in multiple mouse models of traumatic and degenerative neurological disease. However, it is essential to establish whether SARM1 triggers axon degeneration in human neurons before proceeding with the development of SARM1-directed therapeutics. Here we combine genome engineering with the production of human stem cell-derived neurons to test the role of human SARM1 in traumatic and neurotoxic axon degeneration. We have generated two independent SARM1 knockout human iPSC lines that do not express SARM1 protein upon differentiation into neurons. We have developed a modified sensory neuron differentiation protocol that generates human sensory neurons with high yield and purity. We find that SARM1 is required for axon degeneration in response to both physical trauma and in a cellular model of chemotherapy-induced peripheral neuropathy. Finally, we identify cADPR as a biomarker of SARM1 enzyme activity in both healthy and injured human sensory neurons. These findings are consistent with prior molecular and cellular studies in mouse neurons, and highlight the therapeutic potential of SARM1 inhibition for the prevention and treatment of human neurological disease.

Development of specialized sensory neurons engages a nuclear receptor required for functional plasticity.

During development, the nervous system generates neurons that serve highly specialized roles and, accordingly, possess unique functional attributes. The chemosensory BAG neurons of C. elegans are striking exemplars of this. BAGs sense the respiratory gas carbon dioxide (CO2) and, in a context-dependent manner, switch from mediating avoidance of CO2 to supporting CO2 attraction. To determine mechanisms that support the physiology and plasticity of BAG neurons, we used tandem ChIP-seq and cell targeted RNA-seq to identify gene targets of the transcription factor ETS-5, which is required for BAG development. A functional screen of ETS-5 targets revealed that NHR-6, the sole C. elegans NR4A-type nuclear receptor, is required for BAG-mediated avoidance of CO2 and regulates expression of a subset of BAG-specific genes. Unlike ets-5 mutants, which are defective for both attraction to and avoidance of CO2, nhr-6 mutants are fully competent for attraction. These data indicate that the remarkable ability of BAGs to adaptively assign positive or negative valence to a chemosensory stimulus requires a gene-regulatory program supported by an evolutionarily conserved type of nuclear receptor. We suggest that NHR-6 might be an example of a developmental mechanism for modular encoding of functional plasticity in the nervous system.

Microglia Activation and Inflammation During the Death of Mammalian Photoreceptors.

Photoreceptors are highly specialized sensory neurons with unique metabolic and physiological requirements. These requirements are partially met by Müller glia and cells of the retinal pigment epithelium (RPE), which provide essential metabolites, phagocytose waste, and control the composition of the surrounding microenvironment. A third vital supporting cell type, the retinal microglia, can provide photoreceptors with neurotrophic support or exacerbate neuroinflammation and hasten neuronal cell death. Understanding the physiological requirements for photoreceptor homeostasis and the factors that drive microglia to best promote photoreceptor survival has important implications for the treatment and prevention of blinding degenerative diseases like retinitis pigmentosa and age-related macular degeneration.

Sensory Neuroblast Quiescence Depends on Vascular Cytoneme Contacts and Sensory Neuronal Differentiation Requires Initiation of Blood Flow

In many organs, stem cell function depends on communication with their niche partners. Cranial sensory neurons develop in close proximity to blood vessels; however, whether vasculature is an integral component of their niches is yet unknown. Here, two separate roles for vasculature in cranial sensory neurogenesis in zebrafish are uncovered. The first involves precise spatiotemporal endothelial-neuroblast cytoneme contacts and Dll4-Notch signaling to restrain neuroblast proliferation. The second, instead, requires blood flow to trigger a transcriptional response that modifies neuroblast metabolic status and induces sensory neuron differentiation. In contrast, no role of sensory neurogenesis in vascular development is found, suggesting unidirectional signaling from vasculature to sensory neuroblasts. Altogether, we demonstrate that the cranial vasculature constitutes a niche component of the sensory ganglia that regulates the pace of their growth and differentiation dynamics.

Nerve Growth Factor Signaling and Its Contribution to Pain.

Nerve growth factor (NGF) is a neurotrophic protein essential for the growth, differentiation, and survival of sympathetic and sensory afferent neurons during development. A substantial body of evidence, based on both animal and human studies, demonstrates that NGF plays a pivotal role in modulation of nociception in adulthood. This has spurred development of a variety of novel analgesics that target the NGF signaling pathway. Here, we present a narrative review designed to summarize how NGF receptor activation and downstream signaling alters nociception through direct sensitization of nociceptors at the site of injury and changes in gene expression in the dorsal root ganglion that collectively increase nociceptive signaling from the periphery to the central nervous system. This review illustrates that NGF has a well-known and multifunctional role in nociceptive processing, although the precise signaling pathways downstream of NGF receptor activation that mediate nociception are complex and not completely understood. Additionally, much of the existing knowledge derives from studies performed in animal models and may not accurately represent the human condition. However, available data establish a role for NGF in the modulation of nociception through effects on the release of inflammatory mediators, nociceptive ion channel/receptor activity, nociceptive gene expression, and local neuronal sprouting. The role of NGF in nociception and the generation and/or maintenance of chronic pain has led to it becoming a novel and attractive target of pain therapeutics for the treatment of chronic pain conditions.

Renewal and Differentiation of GCD Necklace Olfactory Sensory Neurons.

Both canonical olfactory sensory neurons (OSNs) and sensory neurons belonging to the guanylate cyclase D (GCD) "necklace" subsystem are housed in the main olfactory epithelium, which is continuously bombarded by toxins, pathogens, and debris from the outside world. Canonical OSNs address this challenge, in part, by undergoing renewal through neurogenesis; however, it is not clear whether GCD OSNs also continuously regenerate and, if so, whether newborn GCD precursors follow a similar developmental trajectory to that taken by canonical OSNs. Here, we demonstrate that GCD OSNs are born throughout adulthood and can persist in the epithelium for several months. Phosphodiesterase 2A is upregulated early in the differentiation process, followed by the sequential downregulation of β-tubulin and the upregulation of CART protein. The GCD and MS4A receptors that confer sensory responses upon GCD neurons are initially expressed midway through this process but become most highly expressed once CART levels are maximal late in GCD OSN development. GCD OSN maturation is accompanied by a horizontal migration of neurons toward the central, curved portions of the cul-de-sac regions where necklace cells are concentrated. These findings demonstrate that-like their canonical counterparts-GCD OSNs undergo continuous renewal and define a GCD-specific developmental trajectory linking neurogenesis, maturation, and migration.

The genetic program to specify ectodermal cells in ascidian embryos.

The ascidian belongs to the sister group of vertebrates and shares many features with them. The gene regulatory network (GRN) controlling gene expression in ascidian embryonic development leading to the tadpole larva has revealed evolutionarily conserved gene circuits between ascidians and vertebrates. These conserved mechanisms are indeed useful to infer the original developmental programs of the ancestral chordates. Simultaneously, these studies have revealed which gene circuits are missing in the ascidian GRN; these gene circuits may have been acquired in the vertebrate lineage. In particular, the GRN responsible for gene expression in ectodermal cells of ascidian embryos has revealed the genetic programs that regulate the regionalization of the brain, formation of palps derived from placode-like cells, and differentiation of sensory neurons derived from neural crest-like cells. We here discuss how these studies have given insights into the evolution of these traits.

Atypical Myosin Tunes Dendrite Arbor Subdivision

Dendrite arbor pattern determines the functional characteristics of a neuron. It is founded on primary branch structure, defined through cell intrinsic and transcription-factor-encoded mechanisms. Developing arbors have extensive acentrosomal microtubule dynamics, and here, we report an unexpected role for the atypical actin motor Myo6 in creating primary branch structure by specifying the position, polarity,and targeting of these events. We carried out invivo time-lapse imaging of Drosophila adult sensory neuron differentiation, integrating machine-learning-based quantification of arbor patterning with molecular-level tracking of cytoskeletal remodeling. This revealed that Myo6 and the transcription factor Knot regulate transient surges of microtubule polymerization at dendrite tips; they drive retrograde extension of an actin filament array that specifies anterograde microtubule polymerization and guides these microtubules to subdivide the tip into multiple branches. Primary branches delineate functional compartments; this tunable branching mechanism is key to define and diversify dendrite arbor compartmentalization.

Vasoinhibin generation and effect on neuronal apoptosis in the hippocampus of late mouse embryos.

Morphological and behavioral evidence suggests that vasoinhibin is present in the central nervous system (CNS), triggering neuroendocrine and behavioral responses to stress. Moreover, vasoinhibin reduces neuronal survival and differentiation of primary sensory neurons of the peripheral nervous system. To address the functional role played by vasoinhibin at the CNS, and to better understand the underlying mechanisms involved in its actions, we treated primary cultured hippocampal neurons obtained from embryonic day 16 (E16) mice with a human recombinant vasoinhibin. We examined the resulting cellular changes, focusing on neuronal cell death, and explored the local generation of vasoinhibin within the hippocampus. Our results show that vasoinhibin significantly reduced neuronal cell density and increased immunoreactive activated caspase-3 and TUNEL-positive staining at 72, 16, and 24 h, respectively. Furthermore, vasoinhibin increased the expression of proapoptotic genes BAX, BAD, BIM, and PUMA and decreased that of the antiapoptotic gene BCL-2 at 24 h, as assessed by quantitative real-time reverse transcription-polymerase chain reaction. Vasoinhibin effects were blocked by coincubation with a vasoinhibin antibody or with prolactin. Immunoreactive bands consistent with vasoinhibin were observed in hippocampal extracts by Western blot analysis, and a prolactin standard was cleaved to vasoinhibin by a hippocampal lysate in a heat- and cathepsin D inhibitor pepstatin A-dependent fashion. Taken together, these data support the notion that vasoinhibin is locally produced by cathepsin D within the embryonic mouse hippocampus, a brain region that plays a critical role in emotional regulation, resulting in decreased neuronal cell viability via the activation of the intrinsic apoptosis pathway.

Olig2 regulates terminal differentiation and maturation of peripheral olfactory sensory neurons.

The bHLH transcription factor Olig2 is required for sequential cell fate determination of both motor neurons and oligodendrocytes and for progenitor proliferation in the central nervous system. However, the role of Olig2 in peripheral sensory neurogenesis remains unknown. We report that Olig2 is transiently expressed in the newly differentiated olfactory sensory neurons (OSNs) and is down-regulated in the mature OSNs in mice from early gestation to adulthood. Genetic fate mapping demonstrates that Olig2-expressing cells solely give rise to OSNs in the peripheral olfactory system. Olig2 depletion does not affect the proliferation of peripheral olfactory progenitors and the fate determination of OSNs, sustentacular cells, and the olfactory ensheathing cells. However, the terminal differentiation and maturation of OSNs are compromised in either Olig2 single or Olig1/Olig2 double knockout mice, associated with significantly diminished expression of multiple OSN maturation and odorant signaling genes, including Omp, Gnal, Adcy3, and Olfr15. We further demonstrate that Olig2 binds to the E-box in the Omp promoter region to regulate its expression. Taken together, our results reveal a distinctly novel function of Olig2 in the periphery nervous system to regulate the terminal differentiation and maturation of olfactory sensory neurons.

Irrational behavior in C. elegans arises from asymmetric modulatory effects within single sensory neurons

C. elegans worms exhibit a natural chemotaxis towards food cues. This provides a potential platform to study the interactions between stimulus valence and innate behavioral preferences. Here we perform a comprehensive set of choice assays to measure worms’ relative preference towards various attractants. Surprisingly, we find that when facing a combination of choices, worms’ preferences do not always follow value-based hierarchy. In fact, the innate chemotaxis behavior in worms robustly violates key rationality paradigms of transitivity, independence of irrelevant alternatives and regularity. These violations arise due to asymmetric modulatory effects between the presented options. Functional analysis of the entire chemosensory system at a single-neuron resolution, coupled with analyses of mutants, defective in individual neurons, reveals that these asymmetric effects originate in specific sensory neurons.

Co-expression of C/EBPγ and ATF5 in mouse vomeronasal sensory neurons during early postnatal development.

The differentiation of sensory neurons involves gene expression changes induced by specific transcription factors. Vomeronasal sensory neurons (VSNs) in the mouse vomeronasal organ (VNO) consist of two major subpopulations of neurons expressing vomeronasal 1 receptor (V1r)/Gαi2 or vomeronasal 2 receptor (V2r)/Gαo, which differentiate from a common neural progenitor. We previously demonstrated that the differentiation and survival of VSNs were inhibited in ATF5 transcription factor-deficient mice (Nakano et al. Cell Tissue Res 363:621-633, 2016). These defects were more prominent in V2r/Gαo-type than in V1r/Gαi2-type VSNs; however, the molecular mechanisms responsible for the differentiation of V2r/Gαo-type VSNs by ATF5 remain unclear. To identify a cofactor involved in ATF5-regulated VSN differentiation, we investigated the expression and function of CCAAT/enhancer-binding protein gamma (C/EBPγ, Cebpg), which is a major C/EBP family member expressed in the mouse VNO and dimerizes with ATF5. The results obtained showed that C/EBPγ mRNAs and proteins were broadly expressed in the postmitotic VSNs of the neonatal VNO, and their expression decreased by the second postnatal week. The C/EBPγ protein was expressed in the nuclei of approximately 70% of VSNs in the neonatal VNO, and 20% of the total VSNs co-expressed C/EBPγ and ATF5 proteins. We examined the trans-acting effects of C/EBPγ and ATF5 on V2r transcription and found that the co-expression of C/EBPγ and ATF5, but not C/EBPγ or ATF5 alone, increased Vmn2r66 promoter reporter activity via the C/EBP:ATF response element (CARE) in Neuro2a cells. These results suggest the role of C/EBPγ on ATF5-regulated VSN differentiation in early postnatal development.

Alternative polyadenylation produces multiple 3\u2019 untranslated regions of odorant receptor mRNAs in mouse olfactory sensory neurons

BackgroundOdorant receptor genes constitute the largest gene family in mammalian genomes and this family has been extensively studied in several species, but to date far less attention has been paid to the characterization of their mRNA 3′ untranslated regions (3’UTRs). Given the increasing importance of UTRs in the understanding of RNA metabolism, and the growing interest in alternative polyadenylation especially in the nervous system, we aimed at identifying the alternative isoforms of odorant receptor mRNAs generated through 3’UTR variation.ResultsWe implemented a dedicated pipeline using IsoSCM instead of Cufflinks to analyze RNA-Seq data from whole olfactory mucosa of adult mice and obtained an extensive description of the 3’UTR isoforms of odorant receptor mRNAs. To validate our bioinformatics approach, we exhaustively analyzed the 3’UTR isoforms produced from 2 pilot genes, using molecular approaches including northern blot and RNA ligation mediated polyadenylation test. Comparison between datasets further validated the pipeline and confirmed the alternative polyadenylation patterns of odorant receptors. Qualitative and quantitative analyses of the annotated 3′ regions demonstrate that 1) Odorant receptor 3’UTRs are longer than previously described in the literature; 2) More than 77% of odorant receptor mRNAs are subject to alternative polyadenylation, hence generating at least 2 detectable 3’UTR isoforms; 3) Splicing events in 3’UTRs are restricted to a limited subset of odorant receptor genes; and 4) Comparison between male and female data shows no sex-specific differences in odorant receptor 3’UTR isoforms.ConclusionsWe demonstrated for the first time that odorant receptor genes are extensively subject to alternative polyadenylation. This ground-breaking change to the landscape of 3’UTR isoforms of Olfr mRNAs opens new avenues for investigating their respective functions, especially during the differentiation of olfactory sensory neurons.