Dissertations / Theses on the topic 'Spinal cord Axons Oligodendroglia'

要旨pdfファイル:タイトル「幼若ラットにおける脊髄損傷後の索路の再生」
Kyoto University (京都大学)
0048
新制・課程博士
博士(医学)
甲第7731号
医博第2084号
新制||医||708(附属図書館)
UT51-99-G325
京都大学大学院医学研究科脳統御医科学系専攻
(主査)教授 金子 武嗣, 教授 柴崎 浩, 教授 川口 三郎
学位規則第4条第1項該当

Thesis (Ph.D)--Biomedical Engineering, Georgia Institute of Technology, 2009.
Committee Co-Chair: Shawn Hochman; Committee Co-Chair: Stephen P. DeWeerth; Committee Member: Lena Ting; Committee Member: Robert J. Butera; Committee Member: Robert Lee; Committee Member: Vivian K. Mushahwar. Part of the SMARTech Electronic Thesis and Dissertation Collection.

Les mouvements locomoteurs rythmiques nécessitent l’intervention de circuits neuronaux qui coordonnent l’activité motrice des deux parties du corps. Ces circuits sont formés majoritairement par les projections des interneurones commissuraux de la moelle épinière. Des facteurs de guidage comme la Nétrine, les Slits jouent un rôle fondamental dans la mise en place de ces projections. Une étude a également montré qu’une signalisation impliquant le récepteur Neuropiline2 (Nrp2) des signaux Sémaphorines de la classe 3 (Sema3), participe au guidage de ces projections et cela uniquement après la traversée de la ligne médiane (Zou et al. 2000). Ma thèse porte sur l’étude fonctionnelle d’un ligand de la Nrp2, la Sema3B dans le développement de ce système de projections. J’ai analysé une souris invalidée pour Sema3B et observé de nombreuses erreurs de trajectoires après la traversée de la ligne médiane. Je me suis ensuite intéressée aux mécanismes sous-jacents au gain de réponse : par une approche pharmacologique et biochimique j’ai pu montrer que le signal de la plaque du plancher inhibe une activité de dégradation dépendante de la calpaine1. L’inhibition de cette voie conduit à la stabilisation d’un co-récepteur de la Nrp2, la Plexine A1 dont l’expression est très faible dans les axones n’ayant pas encore traversé la ligne médiane. Cette régulation permet alors l’assemblage d’un complexe récepteur fonctionnel de Sema3B, comprenant cette Plexine associée à la Nrp2 au niveau des cônes de croissance. J’ai identifié la molécule d’adhérence NrCAM, et le facteur neurotrophique GDNF comme étant les facteurs de la plaque du plancher déclencheurs de la réponse
Rhythmic locomotor movements require neuronal circuits ensuring left-right coordination. Spinal commissural projections participate to left-right coordination of limb movements by mediating reciprocal inhibition in synchrony. Extensive research of the mechanisms governing the formation of commissural pathways focused on dorsally-located spinal commissural neurons, establishing a fundamental role for multiple guidance cues derived for the midline and surrounding tissues, including Netrins, Slits and various morphogens. Semaphorin (Sema2)/Neuropilin-2 (Nrp2) signaling has been proposed to contribute to the guidance of commissural projections in the spinal cord at the post- but not pre-crossing stage (Zou et al, 2000). My PhD project aimed at analyzing the role of a Nrp2 ligand, Sema3B, in the guidance of spinal commissural projections, whose expression is dynamic and restricted to some territories, including the floor plate in which axons cross the midline. Analysis of Sema3B null mice showed that the loss of Sema3B induces a range of guidance defects of post-crossing commissural pathways. I investigated the underlying mechanisms and found that the floor plate signal induces through blockade of a calpain 1-dependant pathway the stabilization of the Nrp2 co-receptor Plexin-A1, and enable the assembly of Nrp2/Plexin-A1 sub-units into functional complexes for Sema3B in post-crossing commissural growth cones. I identified the cell adhesion molecule NrCAM and the neurotrophic factor GDNF as being the floor-platederived signals triggering the gain of response

The control of cellular tyrosine phosphorylation levels is of great importance in many biological systems. Among the kinases and phosphatases that modulate these levels, the LAR-RPTPs have been suggested to act in several key aspects of neural development, and in a dysfunctional manner in various pathologies from diabetes to cancer. The aim of this thesis is to describe the physiological functions of one of the members of this subfamily of RPTPs, namely RPTPsigma. First, we showed that glucose homeostasis is altered in RPTPsigma null mice. They are hypoglycemic and more sensitive to exogenous insulin and we proposed that the insulin hypersensitivity observed in RPTPsigma-null mice is likely secondary to their neuroendocrine dysplasia and GH/IGF-1 deficiency. In addition to regulating nervous system development, RPTPsigma was previously shown to regulate axonal regeneration after injury. In the absence of RPTPsigma, axonal regeneration in the sciatic, facial and optical nerves was enhanced following nerve crush. However, myelin-associated growth inhibitory proteins and components of the glial scar such as CSPGs (chondroitin sulfate proteoglycans) have long been known to inhibit axonal regeneration in the CNS, making spinal cord injury irreversible. In collaboration with Dr Samuel David, we unveiled that RPTPsigma null mice are able to regenerate their corticospinal tract following spinal cord hemisections as opposed to their WT littermates. We then isolated primary neurons from both sets of animals and found that the absence of RPTPsigma promotes the ability of the neurons to adhere to certain inhibitory substrates. Finally, in order to better understand the physiological role of RPTPsigma, we used a yeast substrate-trapping approach, to screen a murine embryonic library for new substrates. This screen identified the RhoGAP p250GAP as a new substrate, suggesting a downstream role for RPTPsigma in RhoGTPase signaling. We also identified p130Cas and Fyn as new binding partners. All these proteins have clear functional links to neurite extension. The characterization of RPTPsigma and its signaling partners is essential for understanding its role in neurological development and may one day translate into treatments of neural diseases and injuries.

Axonal degeneration is a major pathological feature of the central nervous system (CNS) inflammatory demyelinating disease multiple sclerosis (MS). This axonal degeneration has major consequences, as functional axonal regeneration in the CNS is largely absent. Cumulative axonal degeneration is the likely cause of the majority of progressive MS-related disability, and therefore, the need for novel neuroprotective therapies for MS exists. Experimental autoimmune encephalomyelitis (EAE), an animal model of MS pathology, also produces axonal injury. In particular, the optic nerve and spinal cord are key sites of neuroinflammation in mouse EAE. By utilizing this model, the short term and long term effects of the putative neuroprotective cytokine, leukaemia inhibitory factor (LIF), were investigated in the optic nerve and spinal cord utilising a number of outcome measures of axonal dysfunction. These included MRI measures of water diffusivity along (ADC ||) and across (ADC┴) the optic nerves, serum levels of phosphorylated neurofilament heavy chain subunit (pNF-H) and histological morphometric measures. LIF treatment reduced EAE grade and pNF-H plasma levels, decreased ADC┴, but had no effect on ADC ||, axon counts or inflammatory infiltration.
In contrast, genetic deletion of LIF and its sister cytokine ciliary neurotrophic factor (CNTF), not only increased EAE grade and pNF-H levels, but also decreased optic nerve ADC|| and optic nerve and spinal cord axon densities. After reviewing current literature, we hypothesize that the target cell for endogenously upregulated LIF in EAE may be the neuron or axon, whereas the target cell for exogenously administered therapeutic LIF may be another cell type, possibly infiltrating macrophages and activated microglial cells. LIF antagonist treatment did not have any affect on EAE grade, pNF-H levels or MRI parameters. This lack of effect may be due to the inability of the LIF antagonist to enter the CNS, supporting the hypothesis that endogenous LIF has a centrally acting mechanism.

博士
長庚大學
臨床醫學研究所
102
Neurons are absent of re-growth ability or re-establishment of functional connections following spinal cord injury (SCI) in humans, which is due to lack of neurotrophic support or increased production of molecules that inhibit neuronal re-growth in the scar after SCI. Therefore, to understand molecular functions of these neurotrophic factors would help us to further insight into plasticity and regeneration ability of neuron. Among these neurotrophic factors, the nerve growth factor (NGF) and Artemin were chosen to study in this thesis to reveal their ability on neuronal plasticity and regeneration. NGF, a neurotrophin contributes to sensory neurons development in the prenatal stage, but it switches to its functions to neural plasticity and regeneration in mature neurons; whereas Artemin is a member of the glial cell line-derived neurotrophic factor (GDNF) family and has been proved to support the survival of sensory neurons. Nociceptors are the receptors of sensory neurons that detect painful or potentially damage stimulation, and have been shown to be dependent on target-derived NGF for survival during development. To reveal the NGF/Artemin-mediated neuronal plasticity and regeneration ability, the adenovirus expression vector encoding NGF or Artemin was infected glial cells in the dorsal horn of spinal cord of normal and injured groups to examine their protein expression, animal behavioral responses of thermal/mechanical nociception, the expression of pre-synaptic and post-synaptic markers (synaptophysin and cFos) by immunohistochemistry, and neuronal regeneration. At first, the expression of NGF induced robust sprouting (plasticity) calcitonin gene-related peptide (CGRP) axons in the dorsal horn and caused behavioral responses of thermal/mechanical hyperalgesia in uninjured animals. On the other hand, injecting adenovirus encoding NGF 2 weeks after a dorsal root crush injury also induced robust regenerating CGRP axons throughout the dorsal horn 4 weeks after injection but caused different behavioral responses of protective pain, in which molecular mechanisms remained poorly understood. Moreover, no restoration of their appropriate target areas within the spinal cord by sprouting or regenerated CGRP axons was also observed. The expression of synaptophysin and cFos were used to evaluate the relationship between synaptic connections and behavioral responses, which revealed that NGF-induced sprouting of CGRP axons showed in a significant redistribution of synapses and the expression pattern cFos in the deeper dorsal horn. Regeneration of only the CGRP axons showed a general reduction in synapses and cFos expression within laminae I and II; however, inflammation of the hind paw induced peripheral sensitization. This data suggested that although NGF-induced sprouting of peptidergic axons induces robust chronic pain and the expression of cFos throughout the entire dorsal horn, regeneration of the same axons resulted in normal protective pain with a synaptic and cFos distribution similar, albeit significantly less than that shown by the sprouting of CGRP axons. The second part of this thesis, we examined the effect of Artemin on the sensory neurons and investigated if the regenerated nociceptive axons can project back to their specific targets to remake appropriate functional synaptic connections and behavioral responses. Our preliminary data showed that injecting virus encoding Artemin 2 weeks after a dorsal root crush injury only induced regenerated CGRP axons project to the appropriate site within the dorsal horn of spinal cord 4 weeks after injection and restore the behavior function of protective pain. In conclusion, NGF-mediated spouting of CGRP axons induces hyperalgesia and NGF-mediated regeneration of only CGRP axons after rhizotomy induces protective pain. Peripheral inflammation can induce tactile hyperalgesia after NGF-mediated regeneration. Synaptic numbers, cFos expression and localization are different between the 2 treatments. Artemin-mediated only regenerated CGRP axons to the specific site after rhizotomy and recovery of pain. These results provide us a better understanding of how the guidance molecules influence sensory neurons in spinal cord and mediate nociceptive axonal plasticity and regeneration within the spinal cord after injury.

Acquiring knowledge of the morphological, molecular, and functional changes that occur to neurons following axotomy is a key step for a comprehensive understanding of the nervous system and how it reacts to injury. Propriospinal commissural interneurons (PCIs or CINs) are a class of neuron with axons that project through the ventral commissure to the contralateral spinal cord. My goal was to examine the morphological, molecular, and functional changes that occur to adult feline PCIs following a proximal axotomy. We first determined whether proximally axotomized PCIs develop de novo axons from their dendrites. C3 PCIs were proximally axotomized and several weeks later we stained PCIs and prepared the tissue for histological evaluation. Two primary classes of axotomized PCI were identified: those with a very short axon (called permanently axotomized) and those with an axon that projected across the injury site. Permanently axotomized PCIs had processes with morphological features typical of axons that emerged from their distal dendrites. These axonal processes of the distal dendrites also had GAP-43 (an axonal marker) and lacked MAP2a/b (a dendritic marker). We concluded that permanently axotomized PCIs develop de novo axons from distal dendrites. We then determined whether the axons that crossed the lesion site were representative of spontaneous functional regeneration. First, we showed that PCI axons regenerate through an environment that is typically highly inhibitory to regenerating axons. Second, we established that the regenerated axons conduct action potentials. Finally, we found that regenerated PCI axons form functional synaptic connections with neurons in the contralateral spinal cord. Collectively, these data indicated that spinal interneurons are capable of spontaneous functional regeneration through an injured spinal cord. PCI growth cones are complex and unlike growth cones previously described in the literature. The final study of the thesis examines the morphologies of PCI growth cones within spinal cord injury sites. We found that PCI growth cones have a wide range of morphologies that is independent of their location within the lesion site. Taken together, these data indicate that PCIs have a remarkable capacity for axonal elongation and contribute to remodelling of spinal circuitry following spinal injury.
Thesis (Ph.D, Physiology) -- Queen's University, 2009-12-07 11:21:47.036

Indiana University-Purdue University Indianapolis (IUPUI)
After spinal cord injury (SCI), poor axonal regeneration of the central nervous system, which mainly attributed to glial scar and low intrinsic regenerating capacity of severely injured neurons, causes limited functional recovery. Combinatory strategy has been applied to target multiple mechanisms. Schwann cells (SCs) have been explored as promising donors for transplantation to promote axonal regeneration. Among the central neurons, descending propriospinal neurons (DPSN) displayed the impressive regeneration response to SCs graft. Glial cell line-derived neurotrophic factor (GDNF), which receptor is widely expressed in nervous system, possesses the ability to promote neuronal survival, axonal regeneration/sprouting, remyelination, synaptic formation and modulate the glial response. We constructed a novel axonal permissive pathway in rat model of thoracic complete transection injury by grafting SCs over-expressing GDNF (SCs-GDNF) both inside and caudal to the lesion gap. Behavior evaluation and histological analyses have been applied to this study. Our results indicated that tremendous DPSN axons as well as brain stem axons regenerated across the lesion gap back to the caudal spinal cord. In addition to direct promotion on axonal regeneration, GDNF also significantly improved the astroglial environment around the lesion. These regenerations caused motor functional recovery. The dendritic plasticity of axotomized DPSN also contributed to the functional recovery. We applied a G-mutated rabies virus (G-Rabies) co-expressing green fluorescence protein (GFP) to reveal Golgi-like dendritic morphology of DPSNs and its response to axotomy injury and GDNF treatment. We also investigated the neurotransmitters phenotype of FluoroGold (FG) labeled DPSNs. Our results indicated that over 90 percent of FG-labeled DPSNs were glutamatergic neurons. DPSNs in sham animals had a predominantly dorsal-ventral distribution of dendrites. Transection injury resulted in alterations in the dendritic distribution, with dorsal-ventral retraction and lateral-medial extension of dendrites. Treatment with GDNF significantly increased the terminal dendritic length of DPSNs. The density of spine-like structures was increased after injury and treatment with GDNF enhanced this effect.