Restorative Neurology and Neuroscience

Purpose: Explore in healthy subjects whether the differences in cortical activation between ankle and finger movements observed in earlier studies with no attention to accuracy also apply to accuracy-demanding tasks.

Methods: Twenty healthy subjects performed right-finger and right-ankle tracking tasks during functional magnetic resonance imaging. Cortical activation was analyzed in the primary motor area (M1), primary somatosensory area (S1), supplementary motor area (SMA), and premotor cortex (PMC). The blood-oxygen-level-dependent signal intensity of active voxels was compared between the finger and ankle conditions.

Results: The results indicated that finger tracking exhibited greater intensity in contralateral (left) M1 and S1 compared to ankle tracking, and ankle tracking exhibited greater intensity in both contralateral and ipsilateral SMA compared to finger tracking. Both M1 and S1 showed more lateralized (contralaterally) organization during finger tracking compared to ankle tracking. Activation maps derived from contrasting the two tracking conditions to each other suggested that ipsilateral areas serve an important role in accuracy-demanding tasks. Regression analysis revealed signal intensity in contralateral M1 and ipsilateral S1 as significant variables in predicting tracking accuracy, but only for finger tracking.

Conclusions: Differences in lateralization between finger and ankle performance found earlier (Kapreli et al., 2006) during simple movement tasks also hold for accuracy-demanding tasks, but dissimilarity in brain organization exists in that the latter show greater ipsilateral activity for both joints. These results invite further research on controlled movements in the lower limb to promote favorable brain reorganization and recovery in people with brain injury.

Purpose: Traumatic brain injury (TBI) -induced brain edema can be reduced by acute progesterone (PROG) treatment in young adult males and females, and in aged males. To extend these findings we tested these hypotheses: 1. Acute PROG treatment post-TBI will reduce cortical edema in aged females as much as in young adults. 2. TBI will induce edema in sub-cortical structures (SCS): the thalamus (TH), hypothalamus (HT), brain stem (BS) and anterior pituitary (AP). 3. Acute, systemic PROG treatment post-TBI will reduce edema in SCS.

Methods: Young adult (n=42) and aged (n=40), bilaterally ovariectomized rats were given medial frontal cortical (MFC) contusion injury, treated with PROG (16 mg/kg body weight) or vehicle at 1, 6 and 24 hours post-injury and killed at 6, 24 and 48 hours post-injury. Their brains were removed and the target areas isolated and measured for water content.

Results: TBI induced cortical and delayed sub-cortical edema. Acute PROG treatment decreased this edema. At 6 hours post-TBI serum PROG levels were substantially elevated in both young and aged, PROG-treated, groups, but were higher in the latter.

Conclusion: Acute PROG treatment post-TBI could prove an effective intervention to prevent or attenuate systemic, post-injury cortical and sub-cortical edema in young and aged females.

Purpose: Promising new rehabilitative approaches to improve the substantial motor disability associated with chronic stroke include pharmacotherapy to enhance motor recovery. We conducted a single-blind placebo-controlled crossover pilot study to investigate the effects of prolonged treatment with L-DOPA in stroke patients.

Methods: Ten chronic (10–48 months) stroke patients received placebo or L-DOPA 100 mg daily for 5 weeks. During drug's treatment patients suspended physiotherapy. Patients underwent clinical evaluation (Rivermead Motor Assessment, Nine Hole Peg Test, and 10 meter walking test) and transcranial magnetic stimulation recordings from the affected and unaffected hemisphere (resting motor threshold, motor evoked potential amplitude and cortical silent period) before and after 5 weeks of treatment.

Results: After L-DOPA treatment patients improved their walking speed (p< 0.01) and manual dexterity (p< 0.01) with the affected hand, the cortical silent period over the affected hemisphere lengthened (p< 0.01), while no changes were found in placebo-group.

Conclusion: A 5-week course of oral L-DOPA in a single daily dose substantially improves motor performance in patients with chronic stroke and could do so by modulating motor cortical excitability (cortical silent period lengthening) suggesting that cortical inhibitory mechanisms have a role in motor recovery after stroke. Pharmacotherapy could be a useful therapeutic approach for chronic stroke patients.

Purpose: To evaluate whether a standardized nerve graft bridging of a spinal cord injury gap in a device with white to grey matter projections allows regeneration and electrophysiological contact from supraspinal centers to the leg, since previous studies using nerve grafts in combination with acidic fibroblast growth factor have been difficult to microsurgically reproduce.

Methods: A moulded prosthesis containing twelve peripheral nerve grafts with estimated pre-set projections guiding white tracts to grey matter replaced a spinal cord resection gap at level T11 in adult rats. The animals were evaluated with electrophysiology, morphology, immunohistochemistry and functional scoring.

Results: At six months postoperatively the grafts were found to be positioned at the desired locations, motor evoked potentials were detected in 80% of the animals subjected to nerve graft bridging. Morphology and immunohistochemical analysis revealed numerous de novo axons in the grafts, which possibly reached into the spinal cord on the other side.

Conclusion: Replacing the resected spinal cord with oblique peripheral nerve grafts results in spinal cord regeneration and positive motor evoked potential. Transplantation of nerve grafts organized in a prosthesis is re-producible, sustains desired projections and ensures transverse ends towards the spinal cord surface and may be useful in the future.

Purpose: nerve regeneration and functional recovery are often incomplete after peripheral nerve damage. The aim of this study was to determine if chondroitinase ABC injection at the lesion site, 1 hour of electrical stimulation and the combination of these treatments at the time of repair could be effective in promoting muscle reinnervation.

Methods: the right sciatic nerve was completely sectioned in 32 female Sprague-Dawley rats. End-to-end microsuture repair was undertaken and fibrin glue was added. Five groups were studied: 1) suture (S) + fibrin glue (F) only; 2) S + F + chondroitinase ABC; 3) S + F + electrical stimulation; 4) S + F + chondroitinase ABC + electrical stimulation; 5) uninjured nerve.

Results: post recovery kinematics showed larger excursion of the hip-ankle-toe angle during walking in groups 2, 3 and 4 than in group 1 (p< 0.05). In vivo electromyographic activity and maximal muscle force were similar between groups 2, 3, 4 and 5, with higher values in all of them compared to group 1 (p< 0.05). Also, the distal stump of the sciatic nerve was excised, and cross-sectioning revealed that the number of axons were similar in all groups.

Conclusions: 150 days after nerve transection, recovery was incomplete with S and F only. Chondroitinase ABC injection at the lesion site and/or 1 hour of electrical stimulation of the proximal nerve stump were beneficial in promoting nerve regeneration and functional muscle reinnervation.

Purpose: Previous reports established that after a contusion injury to the rat spinal cord, locomotor function was enhanced by the transplantation of cells from bone marrow referred to as either mesenchymal stem cells or multipotent mesenchymal stromal cells (MSCs). It has also been established that neural stem cells (NSCs) enhance locomotor function after transplantation into the injured rat spinal cord. However, the beneficial effects of NSCs are limited by graft-induced allodynia-like responses. Little is known about the effects of MSCs on sensory function in spinal cord injury. Therefore, the objective of this research was to determine whether transplantation of MSCs into the injured rat spinal cord induces allodynia-like responses.

Methods: Contusion injuries of two different severities were induced in rats to examine the effects of transplantation with MSCs on sensorimotor deficits. The effects of MSCs on chronic inflammation were investigated, since inflammation is reported to have a role in the sensorimotor deficits associated with spinal cord injury. In addition, observations in other models suggest that MSCs possess immunosuppressive effects.

Results: We found that in contrast to previous observations with the transplantation of neural stem cells, transplantation of MSCs did not induce allodynia. MSCs attenuated injury-induced sensitivity to mechanical stimuli but had no effect on injury-induced sensitivity to cold stimuli. MSCs also significantly attenuated the chronic inflammatory response as assayed by GFAP immunoreactivity for reactive astrocytes and ED1 immunoreactivity for activated macrophages/microglia. In addition, transplantation of MSCs increased white matter volumes and decreased cyst size in sections of the cord containing the lesion.

Conclusion: The results suggest that the sensorimotor enhancements produced by MSCs can at least in part be explained by anti-inflammatory/immunosuppressive effects of the cells, similar to such effects of these cells observed in other experimental models.

Purpose: Neural stem and progenitor cells (NSPC) generate neurons and glia, a feature that makes them attractive for cell replacement therapies. However, efforts to transplant neural progenitors in animal models of brain injury typically result in high cell mortality and poor neuronal differentiation.

Methods: In an attempt to improve the outcome for grafted NSPC after controlled cortical impact we transplanted Enhanced Green Fluorescent Protein (EGFP)-positive NSPC into the contra lateral ventricle of mice one week after injury.

Results: Grafted EGFP-NSPC readily migrated to the injured hemisphere where we analyzed the proportion of progenitors and differentiated progeny at different time points. Transplantation directly into the injured parenchyma, resulted in few brains with detectable EGFP-NSPC. On the contrary, in more than 90% of the mice that received a transplant into the lateral ventricle detectable EGFP-positive cells were found. The cells were integrated into the lateral ventricle wall of the un-injured hemisphere, throughout the corpus callosum, and in the cortical perilesional area. At one-week post transplantation, grafted cells that had migrated to the perilesion area mainly expressed markers of neural progenitors and neurons, while in the corpus callosum and the ventricular lining, grafted cells with a glial fate were more abundant. After 3 months, grafted cells in the perilesion area were less abundant whereas cells that had migrated to the walls of the third- and lateral- ventricle of the injured hemisphere were still detectable, suggesting that the injury site remained a hostile environment.

Conclusion: Transplantation to the lateral ventricle, presumably for being a neurogenic region, provides a favorable environment improving the outcome for grafted NSPC both in term of their appearance at the cortical site of injury, and their acquisition of neural markers.

Purpose: Damage to segmental motoneurons and to spinal cord parenchyma cause denervation atrophy to the muscles, contributing to the chronic disability originated by spinal cord injury (SCI) and spinal motor neuron diseases. After SCI, damage is promoted by several underlying mechanisms, including release of glutamate and consequent over-activation of glutamate receptors, mainly NMDA receptors, that lead to neuronal death. Due to the lack of effective treatments for such conditions, new alternatives need to be explored.

Methods: In order to perform a relatively quick and high-fidelity drug screening, we optimized a postnatal rat organotypic spinal cord culture. By using a glutamate excitotoxic model of spinal cord damage on the explants, we compared the neuroprotective efficacy of four agents: methylprednisolone, erythropoietin, riluzole and rolipram. We evaluated the number of surviving ventral motor neurons stained with the SMI32 antibody and estimated the cord tissue preservation by quantifying the amount of EthD fluorescent probe incorporated into the cells.

Results: The best tissue protection was achieved with riluzole (98%) whereas the highest motoneuron preservation was obtained with methylprednisolone (92%).

Conclusion: The in vitro model used may serve to initiate comparative analyses of new compounds to narrow the choice for future neuroprotective agents to be tested in vivo.

Purpose: To determine the feasibility of restoring electroretinogram (ERG) activity of exenterated swine eyes following in vivo arterial anastomosis.

Methods: The carotid artery was exposed and cannulated. The eye was exenterated along with the extraocular muscles and surrounding connective tissue. Prior to eye transplantation, the ophthalmic artery was identified and anastomosed to the carotid artery. Perfusion was confirmed by injecting FITC-conjugated tomato lectin into the anastomotic tubing and performing confocal microscopy of retinal flat-mounts. Dark-adapted ERG and optic nerve responses were analyzed to assess retinal function, and dilated eye examination and retinal imaging were performed.

Results: Arterial anastomosis resulted in perfusion of blood from the carotid artery through the anastomosis and into the ophthalmic artery. Arterial perfusion was confirmed by the presence of tomato lectin-stained retinal vessels. Immediately following the anastomosis, ERG and optic nerve activities were minimal. However, an "a" wave (representing photoreceptor activity), "b" wave (representing bipolar cell activity), and optic nerve responses (representing RGC activity) were detected 30 min after reperfusion.

Conclusions: Electroretinographic function is partially recovered following re-anastomosis of exenterated swine eyes. This model would be useful for further studies on eye transplantation.

Purpose: human embryonic stem cells (hESCs) which express a reporter gene consistently during all phases of differentiation would be valuable for basic research on cell transplantation. In this study, we describe karyotypically-abnormal variant hESCs, BGO1V2-EFG, which express hrGFP driven by the EF1 promoter.

Methods: BGO1V2-EFG cells were analyzed by using immunocytochemistry, single cell-based confocal image, and in vitro differentiation, including dopaminergic differentiation.

Results: Undifferentiated BGO1V2-EFG cells expressed pluripotent ESC markers and retained the ability to differentiate into cell types of all three germ layers. BGO1V2-EFG cells maintained stable and robust hrGFP expression in vitro in the undifferentiated state and during differentiation. The EF1 promoter retained activity during dopaminergic differentiation, as 76% of tyrosine hydroxlase (TH)-positive cells co-expressed hrGFP by confocal analysis. Treated with sodium butyrate (0.02 mM to 2.0 mM), an inhibitor of histone deacetylase (HDAC), during differentiation did not affect hrGFP expression, although TH expression was reduced by higher concentrations of sodium butyrate.

Conclusion: BGO1V2-EFG cells maintain stable and robust hrGFP expression in the undifferentiated state and during neural differentiation. Especially, the EF1 promoter was effective in driving hrGFP expression during dopaminergic differentiation. BGO1V2-EFG cells may be useful for transplantation studies in Parkinson disease animal models.