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Unknown,Transcriptomics,Genomics,Proteomics

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Transcription profiling of mouse rat cortical axons from un-injured and injured animals isolated after 13 days in culture

ABSTRACT: Efficient growth cone regeneration requires protein synthesis in the adult mammalian brain and spinal cord. Recent evidence suggests that the local availability of protein synthesis machinery in adult mammalian axons may be an indicator of their regenerative capacity. Here we investigated the local protein synthesis capacity in matured cortical axons, which have poor regenerative capacity, yet are critical for recovery following injury due to traumatic brain injury and stroke. This work is the first to biochemically isolate and identify mRNA from mammalian cortical axons, making use of a unique microfluidic platform to isolate axons free of other cellular debris. We first sought to identify mRNA in naÃ¯ve axons that makes up the pool of mRNA available for translation initiated following axotomy. Next, we investigated changes in the mRNA population localized to axons 2 days following axotomy and growth cone regeneration. Experiment Overall Design: Cortical axons were harvested using the compartmentalized microfluidic platform after 13 days in culture at a time when they express mature synaptic proteins. A total of 7 Genechips were used for the uninjured cortical axons from 3 different culture batches. 3 Genechips were used for neurons isolated from the neuronal compartment of the microfluidic platfrom. The neuronal Genechip were used to compare mRNA populations with axonal Genechips and for quality control purposes. 3 Genechips were used for regenerating axons; for these, axons were axotomized at 11 days in culture, then allowed to regrow for 2 days before harvesting.

ORGANISM(S): Rattus norvegicus

SUBMITTER: Anne Taylor

PROVIDER: E-GEOD-11730 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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mRNAs in un-injured and injured rat cortical axons isolated after 13 days in culture

Project description:Efficient growth cone regeneration requires protein synthesis in the adult mammalian brain and spinal cord. Recent evidence suggests that the local availability of protein synthesis machinery in adult mammalian axons may be an indicator of their regenerative capacity. Here we investigated the local protein synthesis capacity in matured cortical axons, which have poor regenerative capacity, yet are critical for recovery following injury due to traumatic brain injury and stroke. This work is the first to biochemically isolate and identify mRNA from mammalian cortical axons, making use of a unique microfluidic platform to isolate axons free of other cellular debris. We first sought to identify mRNA in naïve axons that makes up the pool of mRNA available for translation initiated following axotomy. Next, we investigated changes in the mRNA population localized to axons 2 days following axotomy and growth cone regeneration. Keywords: axonal expression analysis; injury state analysis

2009-01-01 | GSE11730 | GEO

Distal axotomy enhances retrograde presynaptic excitability onto injured pyramidal neurons via trans-synaptic signaling

Project description:Injury of descending motor tracts remodels cortical circuitry and leads to enhanced neuronal excitability, thus influencing recovery following injury. The neuron-specific contributions remain unclear due to the complex cellular composition and connectivity of the CNS. We developed a microfluidics-based in vitro model system to examine intrinsic synaptic remodeling following axon damage. We found that distal axotomy of cultured rat pyramidal neurons caused dendritic spine loss at synapses onto the injured neurons followed by a persistent retrograde enhancement in presynaptic excitability over days. These in vitro results mirrored hyper-activity of directly injured corticospinal neurons in hindlimb motor cortex layer Vb following spinal cord contusion. In vitro axotomy-induced hyper-excitability coincided with elimination of inhibitory presynaptic terminals, including those formed onto dendritic spines. We identified netrin-1 as downregulated following axotomy and exogenous netrin-1 applied 2 days after injury normalized spine density, presynaptic excitability, and the fraction of inhibitory inputs onto injured neurons. These findings demonstrate a novel model system for studying the response of pyramidal circuitry to axotomy and provide new insights of neuron-specific mechanisms that contribute to synaptic remodeling.

2016-11-02 | GSE89407 | GEO

Axonal mRNA in human embryonic stem cell derived neurons

Project description:The identification of axonal mRNAs in model organisms has led to the discovery of many axonally translated proteins required for axon guidance and injury response. The extent to which these axonal mRNAs are conserved in humans is unknown. Here we report on the axonal transcriptome of glutamatergic neurons derived from human embryonic stem cells (hESC-neurons) grown in axon isolating microfluidic chambers. We identified mRNAs enriched in axons, representing a functionally unique local transcriptome as compared to the whole neuron transcriptome. Further, we found that the enriched functional categories within high confidence axonal transcripts resemble those in the axonal transcriptome of rat cortical neurons. Comparing our list of human axonal transcripts to similar datasets generated from embryonic and adult rat dorsal root ganglia and rat cortical neurons we found 60 mRNAs common to all four neuron types. We found that over half of these genes are associated with neurological phenotypes or diseases in model organisms and human. This data provides an important resource for studying local mRNA translation in human axons and has the potential to reveal both conserved and unique axonal mechanisms across species and neuronal types. We analyzed the axonal and whole hESC-neuron transcriptome in triplicate using the Affymetrix Human Gene 2.0 ST Array platform.

2016-07-29 | E-GEOD-84975 | biostudies-arrayexpress

Changes of axonal transcriptomes in response to local Abeta

Project description:We report that local application of Abeta(1-42) for 24h triggers changes in the localized transcriptome, while the transcriptome in cell bodies remains largely unchanged. Axonal and somatic mRNA profiles of rat embryonic hippocampal neurons grown in microfluidic chambers were generated by RNA-Seq in duplicates using an Illumina MiSeq. Axons only were treated for 24 h with soluble oligomeric Abeta(1-42) or vehicle control.

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miR-182 regulates Slit2-mediated axon guidance by modulating the local translation of a specific mRNA

Project description:During brain wiring, mRNAs are trafficked into axons and growth cones where they are differentially translated in response to extrinsic signals. Differential control of local protein synthesis mediates neuronal compartment-specific behaviors that aid axon guidance. Yet little is understood about how specific mRNAs are selected for translation. Here we have investigated the local role of microRNAs (miRNAs) in mRNA-specific translation during axon pathfinding of Xenopus laevis retinal ganglion cell (RGC) axons. Profiling experiments revealed a rich repertoire of axonal miRNAs in developing RGC axons and identified miR-182 as one of the most abundant. Loss of miR182 impairs Slit2-induced growth cone repulsion and causes RGC axon targeting defects in vivo. To aid miRNA target prediction, we also profiled mRNA expression in RGC axons. Our results show that miR-182 targets cofilin1 mRNA in RGC growth cones and modulates its local translation in response to Slit2.

2017-01-23 | GSE86883 | GEO

Axonal mRNA in human embryonic stem cell derived neurons

2016-07-29 | GSE84975 | GEO

Peripheral Nerve Single-Cell Analysis Identifies Mesenchymal Ligands that Promote Axonal Growth (injured sciatic nerve microarray data)

Project description:Peripheral nerves provide a supportive growth environment for developing and regenerating axons and are essential for maintenance and repair of many non-neural tissues. This capacity has largely been ascribed to paracrine factors secreted by nerve-resident Schwann cells. Here, we used single-cell transcriptional profiling to identify ligands made by different injured rodent nerve cell types and have combined this with cell-surface mass spectrometry to computationally model potential paracrine interactions with peripheral neurons. These analyses show that peripheral nerves make many ligands predicted to act on peripheral and CNS neurons, including known and previously uncharacterized ligands. While Schwann cells are an important ligand source within injured nerves, more than half of the predicted ligands are made by nerve-resident mesenchymal cells, including the endoneurial cells most closely associated with peripheral axons. At least three of these mesenchymal ligands, ANGPT1, CCL11, and VEGFC, promote growth when locally applied on sympathetic axons. These data therefore identify an unexpected paracrine role for nerve mesenchymal cells and suggest that multiple cell types contribute to creating a highly pro-growth environment for peripheral axons.

2020-05-06 | GSE146883 | GEO

Transcriptional Analysis of the Effects of Integrin-driven Axon Regeneration in the Spinal Cord

Project description:To elucidate the mechanisms and down-stream pathways affected by activated integrin expression, we performed transcriptomic analyses of dorsal root gangliona (DRG) sensory neurons transduced with α9 integrin and its activator kindlin-1(α9k1), which enable axons to interact with tenascin-C, or controls, with and without dorsal root/central axotomy.

2022-11-30 | GSE188775 | GEO

Overlapping transcriptional programs promote survival and axonal regeneration of injured retinal ganglion cells [2]

Project description:Neurons of the central nervous system (CNS) display only a limited ability to survive and regenerate their axons after an injury. In mice, 85% of retinal ganglion cells (RGCs) die within 2 weeks of axotomy by optic nerve crush (ONC) and only few survivors regenerate axons. In the past years, a multitude of interventions have been identified to improve RGC survival and regeneration after an injury, however, each only protects a subset of neurons and stimulates axon regrowth in an even smaller set.. Here, we sought out to elucidate the molecular mechanisms underlying this selective responsiveness and investigated genes regulated by three well established survival and regeneration-promoting interventions – activation of the MTOR pathway via deletion of its inhibitor Pten, activation of the Jak/Stat-pathway by deletion of its endogenous inhibitor Socs3, and overexpression of the neurotrophic cytokine CNTF. Analysis of the transcriptomes from >125,000 single RGCs at various time points after ONC showed that while broad survival of all RGC types could be induced with each intervention, type-independent axon regeneration was only overcome with the manipulation of multiple pathways. Those RGCs were able to mitigate the injury response and simultaneously upregulated survival and regeneration associated programs (prior and after injury). Four independent ways of analysis identified these programs to be differentially regulated among RGCs, with distinct signatures for degenerating, surviving and regenerating cells. Finally, testing some genes associated with the regeneration-program in vivo identified potential future therapeutic targets to promote neuroprotection and axonal regeneration.

2022-06-28 | GSE202154 | GEO

Overlapping transcriptional programs promote survival and axonal regeneration of injured retinal ganglion cells [1]

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