Browsing by Author "Weidner, Norbert"

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    Depolarization and electrical stimulation enhance in vitro and in vivo sensory axon growth after spinal cord injury
    (Elsevier, 2018-02) Goganau, Ioana; Sandner, Beatrice; Weidner, Norbert; Fouad, Karim; Blesch, Armin; Neurological Surgery, School of Medicine
    Activity dependent plasticity is a key mechanism for the central nervous system (CNS) to adapt to its environment. Whether neuronal activity also influences axonal regeneration in the injured CNS, and whether electrical stimulation (ES) can activate regenerative programs in the injured CNS remains incompletely understood. Using KCl-induced depolarization, in vivo ES followed by ex-vivo neurite growth assays and ES after spinal cord lesions and cell grafting, we aimed to identify parameters important for ES-enhanced neurite growth and axonal regeneration. Using cultures of sensory neurons, neurite growth was analyzed after KCl-induced depolarization for 1-72h. Increased neurite growth was detected after short-term stimulation and after longer stimulation if a sufficient delay between stimulation and growth measurements was provided. After in vivo ES (20Hz, 2× motor threshold, 0.2ms, 1h) of the intact sciatic nerve in adult Fischer344 rats, sensory neurons showed a 2-fold increase in in vitro neurite length one week later compared to sham animals, an effect not observed one day after ES. Longer ES (7h) and repeated ES (7days, 1h each) also increased growth by 56-67% one week later, but provided no additional benefit. In vivo growth of dorsal column sensory axons into a graft of bone marrow stromal cells 4weeks after a cervical spinal cord lesion was also enhanced with a single post-injury 1h ES of the intact sciatic nerve and was also observed after repeated ES without inducing pain-like behavior. While ES did not result in sensory functional recovery, our data indicate that ES has time-dependent influences on the regenerative capacity of sensory neurons and might further enhance axonal regeneration in combinatorial approaches after SCI.
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    Early-onset treadmill training reduces mechanical allodynia and modulates calcitonin gene-related peptide fiber density in lamina III/IV in a mouse model of spinal cord contusion injury
    (Lippincott, Williams, and Wilkins, 2016-03) Nees, Timo A.; Tappe-Theodor, Anke; Sliwinski, Christopher; Motsch, Melanie; Rupp, Rüdiger; Kuner, Rohini; Weidner, Norbert; Blesch, Armin; Department of Neurological Surgery, IU School of Medicine
    Abstract: Below-level central neuropathic pain (CNP) affects a large proportion of spinal cord injured individuals. To better define the dynamic changes of the spinal cord neural network contributing to the development of CNP after spinal cord injury (SCI), we characterized the morphological and behavioral correlates of CNP in female C57BL/6 mice after a moderate T11 contusion SCI (50 kdyn) and the influence of moderate physical activity. Compared with sham-operated animals, injured mice developed mechanical allodynia 2 weeks post injury when tested with small-diameter von Frey hair filaments (0.16 g and 0.4 g filament), but presented hyporesponsiveness to noxious mechanical stimuli (1.4 g filament). The mechano-sensory alterations lasted up to 35 days post injury, the longest time point examined. The response latency to heat stimuli already decreased significantly 10 days post injury reaching a plateau 2 weeks later. In contrast, injured mice developed remarkable hyposensitivity to cold stimuli. Animals that underwent moderate treadmill training (2 × 15 minutes; 5 d/wk) showed a significant reduction in the response rate to light mechanical stimuli as early as 6 days after training. Calcitonin gene-related peptide (CGRP) labeling in lamina III-IV of the dorsal horn revealed significant increases in CGRP-labeling density in injured animals compared with sham control animals. Importantly, treadmill training reduced CGRP-labeling density by about 50% (P < 0.01), partially reducing the injury-induced increases. Analysis of IB4-labeled nonpeptidergic sensory fibers revealed no differences between experimental groups. Abnormalities in temperature sensation were not influenced by physical activity. Thus, treadmill training partially resolves signs of below-level CNP after SCI and modulates the density of CGRP-labeled fibers.
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    An Inducible Tyrosine Kinase Receptor for Axonal Regeneration
    (2016) Deng, Ming; McCall, Julianne; Goganau, Ioana; Motsch, Melanie; Weidner, Norbert; Blesch, Armin; Department of Neurological Surgery, IU School of Medicine
    The prevention or reduction of neuronal degeneration remains a challenge in neurotrophins therapy. An inducible trkA (ItrkA) system has been shown to regulate embryonic dorsal root ganglion (DRG) neuronal survival and neurite outgrowth in vitro. A new ItrkA plasmid ItrkA-membrane (ItrkAmemb) with one adenine at 3’ terminal was established by correcting the sequence of the original plasmid ItrkA-cytosol (ItrkAcyto). Adult DRGs were dissected from adult Fischer 344 rats (8-14 weeks) for the treatment with AP20187 (membrane-permeable small-molecule ligand), vehicle or NGF (Nerve Growth Factor). Neurite outgrowth assessments were done by manually tracing the longest neurite of each neuron. Cell diameters were also measured and averaged for each well. Protein expression after ItrkAmemb transfection and trkA downstream signaling were investigated by Western-blotting. Neurite length of ItrkAmemb transfected DRGs was not influenced by AP20187 or NGF but cells displayed shorter neurites compared to GFP control groups. While ItrkAcyto transfected DRGs cultured with AP20187 had the longest neurite growth compared to ItrkAmemb transfected neurons and ItrkAcyto transfected cells treated with vehicle or NGF, no significant difference to GFP controls was detected. Quantification of the mean diameter of transfected DRGs demonstrated that ItrkAmemb electroporation significantly increased cell diameter, while the diameter of ItrkAcyto transfected neurons and GFP controls were almost the same as naïve neurons. In contrast to electroporated adult DRG neurons, ItrkAmemb virus transfection did not affect the diameter of infected adult DRG Neurons. No obvious difference was observed between the ItrkAmemb and GFP electroporated cells, and only cells transduced with ItrkAmemb treated with AP20187 seemed to show higher phosphorylation both of Akt and Erk1/2. The effect of adult DRG neurons after ItrkA transfection differs, which depends on the change of cell soma size and/or neurite growth, gene delivery technique, expression level and the localization of ItrkA.
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    Neuropathic pain after spinal cord injury: the impact of sensorimotor activity
    (Wolters Kluwer, 2017-03) Nees, Timo A.; Finnerup, Nanna B.; Blesch, Armin; Weidner, Norbert; Neurological Surgery, School of Medicine
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    Regulated Viral BDNF Delivery in Combination with Schwann Cells Promotes Axonal Regeneration through Capillary Alginate Hydrogels after Spinal Cord Injury
    (Elsevier, 2017-09) Liu, Shengwen; Sandner, Beatrice; Schackel, Thomas; Nicholson, LaShae; Chtarto, Abdelwahed; Tenenbaum, Liliane; Puttagunta, Radhika; Müller, Rainer; Weidner, Norbert; Blesch, Armin; Department of Neurological Surgery, School of Medicine
    Grafting of cell-seeded alginate capillary hydrogels into a spinal cord lesion site provides an axonal bridge while physically directing regenerating axonal growth in a linear pattern. However, without an additional growth stimulus, bridging axons fail to extend into the distal host spinal cord. Here we examined whether a combinatory strategy would support regeneration of descending axons across a cervical (C5) lateral hemisection lesion in the rat spinal cord. Following spinal cord transections, Schwann cell (SC)-seeded alginate hydrogels were grafted to the lesion site and AAV5 expressing brain-derived neurotrophic factor (BDNF) under control of a tetracycline-regulated promoter was injected caudally. In addition, we examined whether SC injection into the caudal spinal parenchyma would further enhance regeneration of descending axons to re-enter the host spinal cord. Our data show that both serotonergic and descending axons traced by biotinylated dextran amine (BDA) extend throughout the scaffolds. The number of regenerating axons is significantly increased when caudal BDNF expression is activated and transient BDNF delivery is able to sustain axons after gene expression is switched off. Descending axons are confined to the caudal graft/host interface even with continuous BDNF expression for 8 weeks. Only with a caudal injection of SCs, a pathway facilitating axonal regeneration through the host/graft interface is generated allowing axons to successfully re-enter the caudal spinal cord.
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    Sensorimotor Activity Partially Ameliorates Pain and Reduces Nociceptive Fiber Density in the Chronically Injured Spinal Cord
    (Mary Ann Liebert, 2018-09-15) Sliwinski, Christopher; Nees, Timo A.; Puttagunta, Radhika; Weidner, Norbert; Blesch, Armin; Neurological Surgery, School of Medicine
    A large proportion of patients suffering from spinal cord injury (SCI) develop chronic central neuropathic pain. Previously, we and others have shown that sensorimotor training early after SCI can prevent the development of mechanical allodynia. To determine whether training initiated in the subchronic/chronic phase remains effective, correlates of below-level neuropathic pain were analyzed in the hindpaws 5-10 weeks after a moderate T11 contusion SCI (50 kDyn) in adult female C57BL/6 mice. In a comparison of SCI and sham mice 5 weeks post-injury, about 80% of injured animals developed mechanical hypersensitivity to light mechanical stimuli, whereas testing of noxious stimuli revealed hypo-responsiveness. Thermal sensitivity testing showed a decreased response latency after injury. Without intervention, mechanical and thermal hyper-responsiveness were evident until the end of the experiment (10 weeks). In contrast, treadmill training (2 × 15 min/day; 5 × /week) initiated 6 weeks post-injury resulted in partial amelioration of pain behavior and this effect remained stable. Analysis of calcitonin gene-related peptide (CGRP)-labeled fibers in lamina III-IV of the lumbar dorsal horn revealed an increase in labeling density after SCI. This was not due to changes in the number or size distribution of CGRP-labeled lumbar dorsal root ganglion neurons. Treadmill training reduced the CGRP-labeling density in the spinal cord of injured mice, whereas the density of non-peptidergic isolectin-B4 (IB4)+ fibers showed no changes in lamina IIi and a slight reduction of sparse IB4 labeling in laminae III-IV. Thus, sensorimotor activity initiated in the subchronic/chronic phase of SCI remains effective in ameliorating pain behavior and influencing structural changes of the nociceptive system.
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    Systemic epothilone D improves hindlimb function after spinal cord contusion injury in rats
    (Elsevier, 2018) Sandner, Beatrice; Puttagunta, Radhika; Motsch, Melanie; Bradke, Frank; Ruschel, Jörg; Blesch, Armin; Weidner, Norbert; Neurological Surgery, School of Medicine
    Following a spinal cord injury (SCI) a growth aversive environment forms, consisting of a fibroglial scar and inhibitory factors, further restricting the already low intrinsic growth potential of injured adult central nervous system (CNS) neurons. Previous studies have shown that local administration of the microtubule-stabilizing drug paclitaxel or epothilone B (Epo B) reduce fibrotic scar formation and axonal dieback as well as induce axonal growth/sprouting after SCI. Likewise, systemic administration of Epo B promoted functional recovery. In this study, we investigated the effects of epothilone D (Epo D), an analog of Epo B with a possible greater therapeutic index, on fibrotic scarring, axonal sprouting and functional recovery after SCI. Delayed systemic administration of Epo D after a moderate contusion injury (150 kDyn) in female Fischer 344 rats resulted in a reduced number of footfalls when crossing a horizontal ladder at 4 and 8 weeks post-injury. Hindlimb motor function assessed with the BBB open field locomotor rating scale and Catwalk gait analysis were not significantly altered. Moreover, formation of laminin positive fibrotic scar tissue and 5-HT positive serotonergic fiber length caudal to the lesion site were not altered after treatment with Epo D. These findings recapitulate a functional benefit after systemic administration of a microtubule-stabilizing drug in rat contusion SCI.