Experience-dependent structural plasticity in the adult brain: How the learning brain grows.

Volumetric magnetic resonance imaging studies have shown that intense learning can be associated with grey matter volume increases in the adult brain. The underlying mechanisms are poorly understood. Here we used monocular deprivation in rats to analyze the mechanisms underlying use-dependent grey matter increases. Optometry for quantification of visual acuity was combined with volumetric magnetic resonance imaging and microscopic techniques in longitudinal and cross-sectional studies. We found an increased spatial vision of the open eye which was associated with a transient increase in the volumes of the contralateral visual and lateral entorhinal cortex. In these brain areas dendrites of neurons elongated, and there was a strong increase in the number of spines, the targets of synapses, which was followed by spine maturation and partial pruning. Astrocytes displayed a transient pronounced swelling and underwent a reorganization of their processes. The use-dependent increase in grey matter corresponded predominantly to the swelling of the astrocytes. Experience-dependent increase in brain grey matter volume indicates a gain of structure plasticity with both synaptic and astrocyte remodeling.

Altered Cerebral Blood Flow and Potential Neuroprotective Effect of Human Relaxin-2 (Serelaxin) During Hypoxia or Severe Hypovolemia in a Sheep Model.

Specific neuroprotective strategies to minimize cerebral damage caused by severe hypoxia or hypovolemia are lacking. Based on previous studies showing that relaxin-2/serelaxin increases cortical cerebral blood flow, we postulated that serelaxin might provide a neuroprotective effect. Therefore, we tested serelaxin in two emergency models: hypoxia was induced via inhalation of 5% oxygen and 95% nitrogen for 12 min; thereafter, the animals were reoxygenated. Hypovolemia was induced and maintained for 20 min by removal of 50% of the total blood volume; thereafter, the animals were retransfused. In each damage model, the serelaxin group received an intravenous injection of 30 µg/kg of serelaxin in saline, while control animals received saline only. Blood gases, shock index values, heart frequency, blood pressure, and renal blood flow showed almost no significant differences between control and treatment groups in both settings. However, serelaxin significantly blunted the increase of lactate during hypovolemia. Serelaxin treatment resulted in significantly elevated cortical cerebral blood flow (CBF) in both damage models, compared with the respective control groups. Measurements of the neuroproteins S100B and neuron-specific enolase in cerebrospinal fluid revealed a neuroprotective effect of serelaxin treatment in both hypoxic and hypovolemic animals, whereas in control animals, neuroproteins increased during the experiment. Western blotting showed the expression of relaxin receptors and indicated region-specific differences in relaxin receptor-mediated signaling in cortical and subcortical brain arterioles, respectively. Our findings support the hypothesis that serelaxin is a potential neuroprotectant during hypoxia and hypovolemia. Due to its preferential improvement of cortical CBF, serelaxin might reduce cognitive impairments associated with these emergencies.

Botulinum Neurotoxin Application to the Severed Femoral Nerve Modulates Spinal Synaptic Responses to Axotomy and Enhances Motor Recovery in Rats.

Botulinum neurotoxin A (BoNT) and brain-derived neurotrophic factor (BDNF) are known for their ability to influence synaptic inputs to neurons. Here, we tested if these drugs can modulate the deafferentation of motoneurons following nerve section/suture and, as a consequence, modify the outcome of peripheral nerve regeneration. We applied drug solutions to the proximal stump of the freshly cut femoral nerve of adult rats to achieve drug uptake and transport to the neuronal perikarya. The most marked effect of this application was a significant reduction of the axotomy-induced loss of perisomatic cholinergic terminals by BoNT at one week and two months post injury. The attenuation of the synaptic deficit was associated with enhanced motor recovery of the rats 2-20 weeks after injury. Although BDNF also reduced cholinergic terminal loss at 1 week, it had no effect on this parameter at two months and no effect on functional recovery. These findings strengthen the idea that persistent partial deafferentation of axotomized motoneurons may have a significant negative impact on functional outcome after nerve injury. Intraneural application of drugs may be a promising way to modify deafferentation and, thus, elucidate relationships between synaptic plasticity and restoration of function.

The extracellular matrix glycoprotein tenascin-R regulates neurogenesis during development and in the adult dentate gyrus of mice.

Abnormal generation of inhibitory neurons that synthesize γ-aminobutyric acid (GABAergic) is characteristic of neuropsychological disorders. We provide evidence that the extracellular matrix molecule tenascin-R (TNR) - which is predominantly expressed by a subpopulation of interneurons - plays a role in the generation of GABAergic and granule neurons in the murine dentate gyrus by regulating fate determination of neural stem or progenitor cells (NSCs). During development, absence of TNR in constitutively TNR-deficient (TNR(-/-)) mice results in increased numbers of dentate gyrus GABAergic neurons, decreased expression of its receptor ß1 integrin, increased activation of p38 MAPK and increased expression of the GABAergic specification gene Ascl1. Postnatally, increased GABAergic input to adult hippocampal NSCs in TNR(-/-) mice is associated not only with increased numbers of GABAergic and, particularly, parvalbumin-immunoreactive neurons, as seen during development, but also with increased numbers of granule neurons, thus contributing to the increased differentiation of NSCs into granule cells. These findings indicate the importance of TNR in the regulation of hippocampal neurogenesis and suggest that TNR acts through distinct direct and indirect mechanisms during development and in the adult.

Increase of cortical cerebral blood flow and further cerebral microcirculatory effects of Serelaxin in a sheep model.

Serelaxin, recombinant human relaxin-2, modulates endothelial vasodilatory functionality and is under evaluation for treatment of acute heart failure. Little is known about acute effects on cerebral perfusion. We tested the hypothesis that Serelaxin might also have effects on the cerebral microcirculation in a sheep model, which resembles human brain structure quite well. We used laser Doppler flowmetry and sidestream dark-field (SDF) imaging techniques, which are reliable tools to continuously assess dynamic changes in cerebral perfusion. Laser Doppler flowmetry shows that bolus injection of 30 µg Serelaxin/kg body wt induces an increase (P = 0.006) to roughly 150% of cortical cerebral blood flow (CBF), whereas subcortical CBF remains unchanged (P = 0.688). The effects on area-dependent CBF were significantly different after the bolus injection (P = 0.042). Effects on cortical CBF were further confirmed by SDF imaging. The bolus injection of Serelaxin increased total vessel density to 127% (P = 0.00046), perfused vessel density to 145% (P = 0.024), and perfused capillary density to 153% (P = 0.024). Western blotting confirmed the expression of relaxin receptors RXFP1 and truncated RXFP2-variants in the respective brain regions, suggesting a possible contribution of RXFP1 on the effects of Serelaxin. In conclusion, the injection of a high dose of Serelaxin exerts quick effects on the cerebral microcirculation. Therefore, Serelaxin might be suitable to improve cortical microcirculation and exert neuroprotective effects in clinically relevant scenarios that involve cortical hypoperfusion. These findings need to be confirmed in relevant experimental settings involving cerebral cortical hypoperfusion and can possibly be translated into clinical practice.

Imaging of lamination patterns of the adult human olfactory bulb and tract: in vitro comparison of standard- and high-resolution 3T MRI, and MR microscopy at 9.4 T.

PURPOSE: Neurological and smelling disorders (e.g. Alzheimer's disease, sinonasal disease) negatively affect the microstructural integrity of the olfactory bulb's (OB) cortical layers. Recovery processes depend on active restoration of this microstructural integrity enabled by neuroneogenesis in the OB. The aim of this study was to evaluate lamination patterns of the OB and adjacent tract (OT) using high resolution MRI at 3 Tesla (T) as well as MR microscopy at 9.4 T in comparison with histological sections. MATERIAL AND METHODS: Twenty-four human OBs were imaged in vitro using standard (2mm slice thickness) and high resolution (0.2mm slice thickness) T1w and T2w MR imaging at 3T. Based on signal intensity differences, the number of OB layers and the OB lamination patterns were assessed by two observers in consensus. Results were compared using Wilcoxon test. Signal intensity profiles were compared to reference Nissl stained histological sections and imaging results of MR microscopy. OT lamination patterns were assessed and different configurations of cross sectional areas were compared to macroscopic results and OB/OT lamination patterns. RESULTS: Standard resolution at 3T identified three layers in 8.3%, two layers in 83.3%, and one layer in 8.3%. High resolution at 3T (4 layers in 91.7%, 3 layers in 8.3%) significantly performed better (P<0.001). Signal intensity profile analysis at 3T and 9.4 T (yielding up to 6 different signal intensities) correlated with histological sections and enabled quantitative evaluation of OB lamination patterns. 3T MRI of the OT revealed two separate signal intensities in T2w in 73%, a hyperintense core and a hypointense sheath, and the number of OT signal intensities positively correlated (ρ=0.541, P=0.006) with the increasing complexity of the OTs' cross sectional area configurations. Additionally, cross sectional area configurations correlated with macroscopic results (ρ=0.558, P=0.002) and OB lamination patterns (ρ=0.446, P=0.022). CONCLUSIONS: 3T MRI and MR-microscopy indicate the possibility to identify the lamination pattern of the human OB/OT and to reflect the histological status. If further development will be able to provide technical equipment that complies with the condition of human in vivo high resolution imaging achieving a good enough signal noise ratio, the method of signal intensity profile analysis could prospectively enable scientists to assess the OB's microstructural status in neurological and smelling disorders.

Improved reversal learning and working memory and enhanced reactivity to novelty in mice with enhanced GABAergic innervation in the dentate gyrus.

The balance between excitation and inhibition controls fundamental aspects of the hippocampal function. Here, we report an increase in the ratio of inhibitory to excitatory neurons in the dentate gyrus, accompanied by γ-aminobutyric acid(A) (GABA(A)) receptor-dependent impairment of synaptic plasticity and enhancement of activity-dependent changes in excitability in anesthetized adult mice deficient for the extracellular matrix glycoprotein tenascin-R (TNR). TNR-deficient mice showed faster reversal learning, improved working memory, and enhanced reactivity to novelty than wild-type littermates. Remarkably, in wild-type and TNR-deficient mice, faster reversal learning rates correlated at the individual animal level with ratios of parvalbumin-positive interneurons to granule cells and densities of parvalbumin-positive terminals on somata of granule cells. Our data demonstrate that modification of the extracellular matrix by ablation of TNR leads to a new structural and functional design of the dentate gyrus, with enhanced GABAergic innervation, that is, enhanced ratio of inhibitory to excitatory cells, and altered plasticity, promoting working memory and reversal learning. In wild-type mice, the enhanced ratio of inhibitory to excitatory cells in the dentate gyrus also positively correlated with reversal learning, indicating that level of inhibition regulates specific aspects of learning independent of the TNR gene.

Polysialic acid glycomimetic promotes functional recovery and plasticity after spinal cord injury in mice.

Regeneration after injury of the central nervous system is poor due to the abundance of molecules inhibiting axonal growth. Here we pursued to promote regeneration after thoracic spinal cord injury in young adult C57BL/6J mice using peptides which functionally mimic polysialic acid (PSA) and human natural killer cell-1 (HNK-1) glycan, carbohydrate epitopes known to promote neurite outgrowth in vitro. Subdural infusions were performed with an osmotic pump, over 2 weeks. When applied immediately after injury, the PSA mimetic and the combination of PSA and HNK-1 mimetics, but not the HNK-1 mimetic alone, improved functional recovery as assessed by locomotor rating and video-based motion analysis over a 6-week observation period. Better outcome in PSA mimetic-treated mice was associated with higher, as compared with control mice, numbers of cholinergic and glutamatergic terminals and monaminergic axons in the lumbar spinal cord, and better axonal myelination proximal to the injury site. In contrast to immediate post-traumatic application, the PSA mimetic treatment was ineffective when initiated 3 weeks after spinal cord injury. Our data suggest that PSA mimetic peptides can be efficient therapeutic tools improving, by augmenting plasticity, functional recovery when applied during the acute phase of spinal cord injury.

The extracellular matrix glycoprotein tenascin-C is beneficial for spinal cord regeneration.

Tenascin-C (TNC), a major component of the extracellular matrix, is strongly upregulated after injuries of the central nervous system (CNS) but its role in tissue repair is not understood. Both regeneration promoting and inhibiting roles of TNC have been proposed considering its abilities to both support and restrict neurite outgrowth in vitro. Here, we show that spontaneous recovery of locomotor functions after spinal cord injury is impaired in adult TNC-deficient (TNC(-/-)) mice in comparison to wild-type (TNC(+/+)) mice. The impaired recovery was associated with attenuated excitability of the plantar Hoffmann reflex (H-reflex), reduced glutamatergic input, reduced sprouting of monaminergic axons in the lumbar spinal cord and enhanced post-traumatic degeneration of corticospinal axons. The degeneration of corticospinal axons in TNC(-/-) mice was normalized to TNC(+/+) levels by application of the alternatively spliced TNC fibronectin type III homologous domain D (fnD). Finally, overexpression of TNC-fnD via adeno-associated virus in wild-type mice improved locomotor recovery, increased monaminergic axons sprouting, and reduced lesion scar volume after spinal cord injury. The functional efficacy of the viral-mediated TNC indicates a potentially useful approach for treatment of spinal cord injury.

Ablation of adhesion molecule L1 in mice favours Schwann cell proliferation and functional recovery after peripheral nerve injury.

The adhesion molecule L1 is one of the few adhesion molecules known to be beneficial for repair processes in the adult central nervous system of vertebrates by promoting axonal growth and neuronal survival. In the peripheral nervous system, L1 is up-regulated by myelination-competent Schwann cells and regenerating axons after nerve damage but its functional role has remained unknown. Here we tested the hypothesis that L1 is, as in the central nervous system, beneficial for nerve regeneration in the peripheral nervous system by performing combined functional and histological analyses of adult L1-deficient mice (L1y/-) and wild-type (L1y/+) littermates. Contrary to our hypothesis, quantitative video-based motion analysis revealed better locomotor recovery in L1y/- than in L1y/+ mice at 4-12 weeks after transection and surgical repair of the femoral nerve. Motoneuron regeneration in L1y/- mice was also enhanced as indicated by attenuated post-traumatic loss of motoneurons, enhanced precision of motor reinnervation, larger cell bodies of regenerated motoneurons and diminished loss of inhibitory synaptic input to motoneurons. In search of mechanisms underlying the observed effects, we analysed peripheral nerves at short time-periods (3-14 days) after transection and found that Schwann cell proliferation is strongly augmented in L1y/- versus L1y/+ mice. L1-deficient Schwann cells showed increased proliferation than wild-type Schwann cells, both in vivo and in vitro. These findings suggest a novel role for L1 in nerve regeneration. We propose that L1 negatively regulates Schwann cell proliferation after nerve damage, which in turn restricts functional recovery by limiting the trophic support for regenerating motoneurons.

Polysialic acid glycomimetics promote myelination and functional recovery after peripheral nerve injury in mice.

alpha2,8 Polysialic acid (PSA) is a carbohydrate attached to the glycoprotein backbone of the neural cell adhesion molecule (NCAM) and implicated in nervous system development and repair. Here, we investigated whether PSA can improve functional recovery after peripheral nerve lesion in adult mice. We applied a functional PSA mimicking peptide or a control peptide in a polyethylene cuff used to surgically reconnect the severed stumps of the femoral nerve before it bifurcates into the motor and sensory branches. Using video-based motion analysis to monitor motor recovery over a 3 month postoperative period, we observed a better functional outcome in the PSA mimetic-treated than in control mice receiving a control peptide or phosphate buffered saline. Retrograde tracing of regenerated motoneurons and morphometric analyses showed that motoneuron survival, motoneuron soma size and axonal diameters were not affected by treatment with the PSA mimetic. However, remyelination of regenerated axons distal to the injury site was considerably improved by the PSA mimetic indicating that effects on Schwann cells in the denervated nerve may underlie the functional effects seen in motor recovery. In line with this notion was the observation that the PSA mimetic enhanced the elongation of Schwann cell processes and Schwann cell proliferation in vitro, when compared with the control peptide. Moreover, Schwann cell proliferation in vivo was enhanced in both motor and sensory branches of the femoral nerve by application of the PSA mimetic. These effects were likely mediated by NCAM through its interaction with the fibroblast growth factor receptor (FGFR), since they were not observed when the PSA mimetic was applied to NCAM-deficient Schwann cells, and since application of two different FGFR inhibitors reduced process elongation from Schwann cells in vitro. Our results indicate the potential of PSA mimetics as therapeutic agents promoting motor recovery and myelination after peripheral nerve injury.

Genetic ablation of tenascin-C expression leads to abnormal hippocampal CA1 structure and electrical activity in vivo.

Despite evidence that the extracellular matrix glycoprotein tenascin-C (TNC) is implicated in brain development and plasticity, its roles in the intact adult brain are unknown. Here we report that spontaneous local field potential (LFP) activity in freely moving adult TNC-deficient mice is abnormal. The power of cortical and hippocampal theta and gamma oscillations was enhanced in comparison to wild-type mice. The alteration in hippocampal gamma rhythm was subfield specific, such that CA1 gamma was accentuated while dentate gyrus gamma was normal. Similar to LFP, synaptic transmission and plasticity at perforant path synapses in the dentate gyrus were unaffected by the mutation. Morphological analyses revealed a subfield-specific reduction in the CA1 volume and a reduction in the numbers of somatostatin-positive interneurons in the hippocampus as potential structural substrates of the observed functional aberrations. These findings indicate a role for tenascin-C in structural organization of the CA1 hippocampal subfield and in shaping neural activity.

Manual stimulation, but not acute electrical stimulation prior to reconstructive surgery, improves functional recovery after facial nerve injury in rats.

UNLABELLED: The outcome of peripheral nerve injuries requiring surgical repair is poor. Recent work suggested that electrical stimulation (ES) of the proximal nerve stump to produce repeated discharges of the parent motoneurons for one hour could be a beneficial therapy if delivered immediately prior to reconstructive surgery of mixed peripheral nerves. PURPOSE: We tested whether ES has a positive influence on functional recovery after repair of a purely motor nerve, the facial nerve. METHODS: Electrical stimulation (20 Hz) was delivered to the proximal nerve stump of the transected facial nerve for 1 hour prior to nerve reconstruction by end-to-end suture (facial-facial anastomosis, FFA). For manual stimulation (MS), animals received daily rhythmic stroking of the whisker pads. Restoration of vibrissal motor performance following ES or MS was evaluated using video-based motion analysis. We also assessed the degree of collateral axonal branching at the lesion site, by counting motoneuronal perikarya after triple retrograde labeling, and estimated the quality of motor end-plate reinnervation in the target musculature. Outcomes at 4 months were compared to animals receiving sham stimulation (SS) or MS. RESULTS: Neither protocol reduced the degree of collateral sprouting. ES did not improve functional outcome and failed to reduce the proportion of polyinnervated motor end-plates. By contrast, MS restored normal whisking function and reduced polyinnervation. CONCLUSION: Whereas acute ES is not beneficial for facial nerve repair, MS provides long-term benefits.

Brief Electrical Stimulation Improves Functional Recovery After Femoral But Not After Facial Nerve Injury in Rats.

Brief low-frequency electrical stimulation (ES, 1 h, 20 Hz) of the proximal nerve stump has emerged as a potential adjunct treatment for nerve injury. Despite available experimental and clinical data, the potentials and limitations of the ES therapy still have to be defined using different animal models, types of nerves, and clinical settings. Here, we show that brief ES of the proximal stump of the transected rat femoral nerve causes, as estimated by motion analysis, enhanced functional recovery reaching preoperative levels within 5 months of injury, in contrast to the incomplete restoration in sham-stimulated (SS) animals. The functional advantage seen in ES rats was associated with higher numbers, as compared with SS, of correctly targeted quadriceps motoneurons. In contrast, ES prior to facial nerve suture did not lead to improvement of whisking compared with SS. Lack of functional effects of the treatment was correlated with lack of changes, as compared with SS, in the precision of muscle reinnervation and frequency of abnormally innervated muscle fibers. These results show that ES is an effective therapy in a spinal nerve injury model leading to complete restoration of function. Although this finding and the safety of the procedure are encouraging, the results for the facial nerve model suggest that brief ES may not be a universal treatment for nerve injuries. Anat Rec, 302:1304-1313, 2019. © 2019 Wiley Periodicals, Inc.

Glial scar expression of CHL1, the close homolog of the adhesion molecule L1, limits recovery after spinal cord injury.

The Ig superfamily adhesion molecule CHL1, the close homolog of the adhesion molecule L1, promotes neurite outgrowth, neuronal migration, and survival in vitro. We tested whether CHL1, similar to its close homolog L1, has a beneficial impact on recovery from spinal cord injury using adult CHL1-deficient (CHL1-/-) mice and wild-type (CHL1+/+) littermates. In contrast to our hypothesis, we found that functional recovery, assessed by locomotor rating and video-based motion analyses, was improved in CHL1-/- mice compared with wild-type mice at 3-6 weeks after compression of the thoracic spinal cord. Better function was associated with enhanced monoaminergic reinnervation of the lumbar spinal cord and altered pattern of posttraumatic synaptic rearrangements around motoneurons. Restricted recovery of wild-type mice was likely related to early and persistent (3-56 d after lesion) upregulation of CHL1 in GFAP-positive astrocytes at the lesion core. In both the intact spinal cord and cultured astrocytes, enhanced expression of CHL1 and GFAP was induced by application of basic fibroblast growth factor, a cytokine involved in the pathophysiology of spinal cord injury. This upregulation was abolished by inhibitors of FGF receptor-dependent extracellular signal-regulated kinase, calcium/calmodulin-dependent kinase, and phosphoinositide-3 kinase signaling pathways. In homogenotypic and heterogenotypic cocultures of neurons and astrocytes, reduced neurite outgrowth was observed only if CHL1 was simultaneously present on both cell types. These findings and novel in vitro evidence for a homophilic CHL1-CHL1 interaction indicate that CHL1 is a glial scar component that restricts posttraumatic axonal growth and remodeling of spinal circuits by homophilic binding mechanisms.

Manual stimulation of the suprahyoid-sublingual region diminishes polynnervation of the motor endplates and improves recovery of function after hypoglossal nerve injury in rats.

BACKGROUND: Using the rat facial nerve axotomy model, the authors recently showed that manual stimulation of denervated whiskerpad muscles reduced the posttransectional polyinnervation at the neuromuscular junctions and promoted full recovery of vibrissal whisking. OBJECTIVE: Prompted by implications for rehabilitation therapy, the authors examined whether manual stimulation of denervated supra- and infrahyoid muscles would also improve recovery after unilateral lesion on the hypoglossal nerve. METHODS: Adult rats underwent transection of the right hypoglossal nerve. Half of the animals received no postoperative treatment, and the other half were subjected to daily manual stimulation of the suprahyoid/sublingual region for 2 months. Recovery was assessed by measuring the angle of tongue-tip deviation from the midline, degree of collateral axonal branching at the lesion site (counts after retrograde labeling with 2 fluorescent dyes), synaptic input to the hypoglossal motoneurons using synaptophysin immunocytochemistry, tongue-muscles motor representation in the cerebral cortex after c-Fos immunocytochemistry, and portion of polyinnervated neuromuscular junctions. RESULTS: In animals receiving manual stimulation, the tongue-tip deviation was 37.0 +/- 49.37 degrees , whereas values in control nonstimulated rats were significantly higher (50.1 +/- 9.01 degrees ; P < .05; mean +/- SD). Improved recovery was not associated with reduced collateral axonal branching; there were also no differences in tongue-muscles representation in the motor cortex. However, manual stimulation restored the total synaptic input to levels in intact animals and reduced the proportion of polyinnervated neuromuscular junctions compared with nonstimulated animals. CONCLUSION: The data show that manual stimulation of denervated muscles improves functional outcome following peripheral nerve injury. This suggests immediate potential for enhancing clinical rehabilitation strategies.

Novelty-induced behavioral traits correlate with numbers of brainstem noradrenergic neurons and septal cholinergic neurons in C57BL/6J mice.

It is generally accepted that different brain regions regulate specific behavioral responses and that structural alterations in these regions may affect behavior. We investigated whether inter-individual variability in novelty-induced behaviors in C57BL/6J mice correlates with numbers of noradrenergic neurons in the locus coeruleus and cholinergic neurons in the septum. We found that exploration of new stimuli correlated negatively with numbers of noradrenergic neurons, whereas anxiety correlated positively with numbers of cholinergic neurons. The observed correlations suggest physiologically plausible links between structure and function and indicate that precise morphological estimates can be predictive for behavioral responses.

Adeno-associated virus-mediated L1 expression promotes functional recovery after spinal cord injury.

Paucity of permissive molecules and abundance of inhibitory molecules in the injured spinal cord of adult mammals prevent axons from successful regeneration and, thus, contribute to the failure of functional recovery. Using an adeno-associated viral (AAV) vector, we expressed the regeneration-promoting cell adhesion molecule L1 in both neurons and glia in the lesioned spinal cord of adult mice. Exogenous L1, detectable already 1 week after thoracic spinal cord compression and immediate vector injection, was expressed at high levels up to 5 weeks, the longest time-period studied. Dissemination of L1-transduced cells throughout the spinal cord was wide, spanning over more than 10 mm rostral and 10 mm caudal to the lesion scar. L1 was not detectable in the fibronectin-positive lesion core. L1 overexpression led to improved stepping abilities and muscle coordination during ground locomotion over a 5-week observation period. Superior functional improvement was associated with enhanced reinnervation of the lumbar spinal cord by 5-HT axons. Corticospinal tract axons did not regrow beyond the lesion scar but extended distally into closer proximity to the injury site in AAV-L1-treated compared with control mice. The expression of the neurite outgrowth-inhibitory chondroitin sulphate proteoglycan NG2 was decreased in AAV-L1-treated spinal cords, along with reduction of the reactive astroglial marker GFAP. In vitro experiments confirmed that L1 inhibits astrocyte proliferation, migration, process extension and GFAP expression. Analyses of intracellular signalling indicated that exogenous L1 activates diverse cascades in neurons and glia. Thus, AAV-mediated L1 overexpression appears to be a potent means to favourably modify the local environment in the injured spinal cord and promote regeneration. Our study demonstrates a clinically feasible approach of promising potential.

Cell Adhesion Molecule Close Homolog of L1 (CHL1) Guides the Regrowth of Regenerating Motor Axons and Regulates Synaptic Coverage of Motor Neurons.

The close homolog of L1 (CHL1) is a cell adhesion molecule involved in regulation of neuronal differentiation and survival, neurite outgrowth and axon guidance during development. In the mature nervous system, CHL1 regulates synaptic activity and plasticity. The aim of the present study was to evaluate the influence of CHL1 on peripheral nerve regeneration after trauma. Using the established model of mouse femoral nerve regeneration, CHL1 knock-out mice were investigated in comparison to the wild type littermates. First, non-injured mice of both genotypes were compared regarding the synaptic phenotypes in the corresponding spinal cord segment. While no differences in phenotypes were detectable in the femoral nerve, corresponding segments in the spinal cord were observed to differ in that inhibitory perisomatic innervation of motor neurons was increased in CHL1-deficient mice, and numbers of perisomatic cholinergic synapses on motor neuronal somata were reduced. Regarding the femoral nerve after injury, CHL1-deficient mice demonstrated preferential motor axon regrowth into the saphenous vs. quadriceps branch after nerve transection upstream of the nerve bifurcation by 8 weeks after transection, indicating decreased preferential motor re-innervation. Furthermore, in injured wild-type mice, enhanced CHL1 expression was observed in regenerating axons in the proximal nerve stump upstream of the bifurcation at days 1, 3, 5, 7 and 14, and in the distal stump at days 7 and 14 after injury, when compared to non-injured mice. Injury-related upregulation of CHL1 expression was more pronounced in axons than in Schwann cells. Despite a more pronounced capacity for preferential motor axon regrowth in wild-type vs. mutant mice, only a tendency for difference in recovery of motor functions was observed between genotypes, without statistical significance Taken together, these results indicate that CHL1 is involved in peripheral nerve regeneration, because it guides regrowing axons into the appropriate nerve branch and regulates synaptic coverage in the spinal cord.

Tenascin-R restricts posttraumatic remodeling of motoneuron innervation and functional recovery after spinal cord injury in adult mice.

Tenascin-R (TNR) is an extracellular glycoprotein in the CNS implicated in neural development and plasticity. Its repellent properties for growing axons in a choice situation with a conducive substrate in vitro have indicated that TNR may impede regeneration in the adult mammalian CNS. Here we tested whether constitutive lack of TNR has beneficial impacts on recovery from spinal cord injury in adult mice. Using the Basso, Beattie, Bresnahan (BBB) locomotor rating scale, we found that open-field locomotion in TNR-deficient (TNR-/-) mice recovered better that in wild-type (TNR+/+) littermates after compression of the thoracic spinal cord. We also designed, validated, and applied a motion analysis approach allowing numerical assessment of motor functions. We found, in agreement with the BBB score, that functions requiring low levels of supraspinal control such as plantar stepping improved more in TNR-/- mice. This was not the case for motor tasks demanding precision such as ladder climbing. Morphological analyses revealed no evidence that improved recovery of some functions in the mutant mice were attributable to enhanced tissue sparing or axonal regrowth. Estimates of perisomatic puncta revealed reduced innervation by cholinergic and GABAergic terminals around motoneurons in intact TNR-/- compared with TNR+/+ mice. Relative to nonlesioned animals, spinal cord repair was associated with increase in GABAergic and decrease of glutamatergic puncta in TNR-/- but not in TNR+/+ mice. Our results suggest that TNR restricts functional recovery by limiting posttraumatic remodeling of synapses around motoneuronal cell bodies where TNR is normally expressed in perineuronal nets.