Diverse inflammatory threats modulate astrocytes Ca2+ signaling via connexin43 hemichannels in organotypic spinal slices.

Neuroinflammation is an escalation factor shared by a vast range of central nervous system (CNS) pathologies, from neurodegenerative diseases to neuropsychiatric disorders. CNS immune status emerges by the integration of the responses of resident and not resident cells, leading to alterations in neural circuits functions. To explore spinal cord astrocyte reactivity to inflammatory threats we focused our study on the effects of local inflammation in a controlled micro-environment, the organotypic spinal slices, developed from the spinal cord of mouse embryos. These organ cultures represent a complex in vitro model where sensory-motor cytoarchitecture, synaptic properties and spinal cord resident cells, are retained in a 3D fashion and we recently exploit these cultures to model two diverse immune conditions in the CNS, involving different inflammatory networks and products. Here, we specifically focus on the tuning of calcium signaling in astrocytes by these diverse types of inflammation and we investigate the mechanisms which modulate intracellular calcium release and its spreading among astrocytes in the inflamed environment. Organotypic spinal cord slices are cultured for two or three weeks in vitro (WIV) and exposed for 6 h to a cocktail of cytokines (CKs), composed by tumor necrosis factor alpha (TNF-α), interleukin-1 beta (IL-1 ß) and granulocyte macrophage-colony stimulating factor (GM-CSF), or to lipopolysaccharide (LPS). By live calcium imaging of the ventral horn, we document an increase in active astrocytes and in the occurrence of spontaneous calcium oscillations displayed by these cells when exposed to each inflammatory threat. Through several pharmacological treatments, we demonstrate that intracellular calcium sources and the activation of connexin 43 (Cx43) hemichannels have a pivotal role in increasing calcium intercellular communication in both CKs and LPS conditions, while the Cx43 gap junction communication is apparently reduced by the inflammatory treatments.

In vivo two-photon imaging of motoneurons and adjacent glia in the ventral spinal cord.

BACKGROUND: Interactions between motoneurons and glial cells are pivotal to regulate and maintain functional states and synaptic connectivity in the spinal cord. In vivo two-photon imaging of the nervous system provided novel and unexpected knowledge about structural and physiological changes in the grey matter of the forebrain and in the dorsal white matter of the spinal cord. NEW METHOD: Here, we describe a novel experimental strategy to investigate the spinal grey matter, i.e. the ventral horn motoneurons and their adjacent glial cells by employing in vivo two-photon laser-scanning microscopy (2P-LSM) in anesthetized transgenic mice. RESULTS: After retrograde tracer labelling in transgenic mice with cell-specific expression of fluorescent proteins and surgical exposure of the lumbar intumescence groups of motoneurons could be visualized deeply localized in the ventral horn. In this region, morphological responses of microglial cells to ATP could be recorded for an hour. In addition, using in mice with expression of GCaMP3 in astrocytes, physiological Ca2+ signals could be recorded after local noradrenalin application. COMPARISON WITH EXISTING METHODS: Previous in vivo imaging protocols were restricted to the superficial dorsal white matter or upper layers of the dorsal horn. Here, we modified a multi-step procedure originally established for a root-crush injury. We adapted it to simultaneously visualize motoneurons and adjacent glial cells in living animals. CONCLUSION: A modified surgery approach is presented to visualize fluorescently labelled motoneurons and glial cells at a depth of more than 200 µm in the grey matter ventral horn of the mouse spinal cord.

Intraspinal bone-marrow cell therapy at pre- and symptomatic phases in a mouse model of amyotrophic lateral sclerosis.

BACKGROUND: Amyotrophic lateral sclerosis (ALS) is a progressive neurological disease that selectively affects the motor neurons. The details of the mechanisms of selective motor-neuron death remain unknown and no effective therapy has been developed. We investigated the therapy with bone-marrow mononuclear cells (BMMC) in a mouse model of ALS (SOD1(G93A) mice). METHODS: We injected 10(6) BMMC into the lumbar portion of the spinal cord of SOD1(G93A) mice in presymptomatic (9 weeks old) and symptomatic (14 weeks old) phases. In each condition, we analyzed the progression of disease and the lifespan of the animals. RESULTS: We observed a mild transitory delay in the disease progression in the animals injected with BMMC in the presymptomatic phase. However, we observed no increase in the lifespan. When we injected BMMC in the symptomatic phase, we observed no difference in the animals' lifespan or in the disease progression. Immunohistochemistry for NeuN showed a decrease in the number of motor neurons during the course of the disease, and this decrease was not affected by either treatment. Using different strategies to track the BMMC, we noted that few cells remained in the spinal cord after transplantation. This observation could explain why the BMMC therapy had only a transitory effect. CONCLUSION: This is the first report of intraspinal BMMC therapy in a mouse model of ALS. We conclude this cellular therapy has only a mild transitory effect when performed in the presymptomatic phase of the disease.

Improved neurological outcome by intramuscular injection of human amniotic fluid derived stem cells in a muscle denervation model.

PURPOSE: The skeletal muscle develops various degrees of atrophy and metabolic dysfunction following nerve injury. Neurotrophic factors are essential for muscle regeneration. Human amniotic fluid derived stem cells (AFS) have the potential to secrete various neurotrophic factors necessary for nerve regeneration. In the present study, we assess the outcome of neurological function by intramuscular injection of AFS in a muscle denervation and nerve anastomosis model. MATERIALS AND METHODS: Seventy two Sprague-Dawley rats weighing 200-250 gm were enrolled in this study. Muscle denervation model was conducted by transverse resection of a sciatic nerve with the proximal end sutured into the gluteal muscle. The nerve anastomosis model was performed by transverse resection of the sciatic nerve followed by four stitches reconnection. These animals were allocated to three groups: control, electrical muscle stimulation, and AFS groups. RESULTS: NT-3 (Neurotrophin 3), BDNF (Brain derived neurotrophic factor), CNTF (Ciliary neurotrophic factor), and GDNF (Glia cell line derived neurotrophic factor) were highly expressed in AFS cells and supernatant of culture medium. Intra-muscular injection of AFS exerted significant expression of several neurotrophic factors over the distal end of nerve and denervated muscle. AFS caused high expression of Bcl-2 in denervated muscle with a reciprocal decrease of Bad and Bax. AFS preserved the muscle morphology with high expression of desmin and acetylcholine receptors. Up to two months, AFS produced significant improvement in electrophysiological study and neurological functions such as SFI (sciatic nerve function index) and Catwalk gait analysis. There was also significant preservation of the number of anterior horn cells and increased nerve myelination as well as muscle morphology. CONCLUSION: Intramuscular injection of AFS can protect muscle apoptosis and likely does so through the secretion of various neurotrophic factors. This protection furthermore improves the nerve regeneration in a long term nerve anastomosis model.

Restoration of prehensile function for motor paralysis in Hopkins syndrome: case report.

Hopkins syndrome is a rare cause of poliomyelitis-like paralysis affecting 1 or more extremities after an acute attack of asthma. The exact etiology of Hopkins syndrome is not known. A 4-year-old girl developed acute asthma followed by complete flaccid paralysis of the left upper extremity. She underwent staged reconstruction using the double free muscle transfer technique. Rigorous postoperative physiotherapy was carried out to achieve a good functional outcome. At recent follow-up, 27 months after the first procedure, the patient was able to effectively use the reconstructed hand for most daily activities. She had good control and could perform 2-handed activities. The selection of a suitable operative treatment and suitable donor nerves is critical, and there are no clear guidelines in the literature. The double free muscle transfer can be effectively employed in similar cases to restore grasping function.

Cervical spinal cord therapeutics delivery: preclinical safety validation of a stabilized microinjection platform.

OBJECTIVE: The current series represents a preclinical safety validation study for direct parenchymal microinjection of cellular grafts into the ventral horn of the porcine cervical spinal cord. METHODS: Twenty-four 30- to 40-kg female Yorkshire farm pigs immunosuppressed with cyclosporine underwent a cervical laminectomy and ventral horn human neural progenitor cell injection. Cell transplantation in groups 1 to 3 (n = 6 pigs each) was undertaken with the intent of assessing the safety of varied injection volumes: 10, 25, and 50 microL injected at 1, 2.5, and 5 microL/min, respectively. Groups 4 and 5 (n = 3 pigs each) received prolonged immunosuppressant pretreatment in an attempt to demonstrate graft viability. The latter was undertaken in an alternate species (mini-pig versus Yorkshire pig). RESULTS: Neurological morbidity was observed in 1 animal and was attributable to the presence of a resolving epidural hematoma noted at necropsy. Although instances of ventral horn targeting were achieved in all injection groups with a coordinate-based approach, opportunities exist for improvement in accuracy and precision. A relationship between injection volume and graft site cross-sectional area suggested limited reflux. Only animals from group 5 achieved graft survival at a survival end point (t = 1 week). CONCLUSION: This series demonstrated the functional safety of targeted ventral horn microinjection despite evidence for graft site immune rejection. Improvements in graft delivery may be augmented with an adapter to improve control of the cannula entry angle, intraoperative imaging, or larger graft volumes. Finally, demonstration of long-term graft viability in future preclinical toxicity studies may require tailored immunosuppressive therapies, an allograft construct, or tailored choice of host species.

Roles of platelet-derived growth factor-B expression in the ventral horn and motor cortex in the spinal cord-hemisected rhesus monkey.

It is well known that platelet-derived growth factor-B (PDGF-B), a member of the neurotrophic factor family, is involved in normal physiological conditions, pathological changes, and neuroregulation following lesions. But the roles of endogenous PDGF-B in neuroregulation following spinal cord injury are far from being well known, especially in primates. This study explored the role of PDGF-B in the spinal cord and motor cortex in rhesus monkeys subjected to cord hemisection. Evaluation of the hindlimb motor function and the cortical somatosensory evoked potentials (CSEP) demonstrated a significant partial recovery from 30 days post-operation (dpo) to 90 dpo. Immunostaining revealed PDGF-B expression in neurons and scattered macrophages in the spinal cord. The number of PDGF-B immunoreactive neurons in the ventral horn of the spinal cord decreased significantly at the injury site at 14 dpo, followed by a rapid increase that surpassed the numbers in the control group at 30 dpo, and remained at these levels until 90 dpo. The protein levels of PDGF-B and platelet-derived growth factor receptor-beta (PDGFR-beta) as assessed by Western blot, as well as the mRNA levels of PDGF-B as assessed by RT-PCR demonstrated a tendency similar to that seen with immunohistochemistry. PDGF-B antibody administration effectively decreased locomotor function in the hindlimbs, especially on the injured side. No PDGF-B immunoreactive cells were detected in the motor cortex. Taken together, the present findings indicate that intrinsic PDGF-B expressed in the spinal cord may play an essential role in neuroregulation in primates following cord hemisection.

Adenovirus-mediated retrograde transfer of neurotrophin-3 gene enhances survival of anterior horn neurons of twy/twy mice with chronic mechanical compression of the spinal cord.

Chronic mechanical compression of the spinal cord causes neural tissue damage, including loss of anterior horn cells around the level of injury. Exogenous delivery of neurotrophins to neuronal cells could provide neuroprotection to a spinal cord subjected to mechanical injury. We investigated the efficacy of retrograde gene delivery of adenoviral vector (AdV) carrying neurotrophin-3 (NT-3) gene into twy (twy/twy) mouse spinal cord anterior horn neurons with chronic and progressive mechanical compression at C1-C2 level. AdV-NT-3 was used for retrograde delivery via the sternomastoid muscle to the cervical spinal accessory motoneurons in 16-week-old adult twy mice with relatively mild spinal cord compression. Four weeks after the AdV-NT-3 or AdV-beta-galactosidase cDNA (LacZ) as a marker gene injection, the compressed cervical spinal cord was examined histologically, immunohistologically, and by immunoblot analysis. Immunoreactivity to NT-3 was significantly enhanced in the AdV-NT-3-injected twy mice compared with the AdV-LacZ-injected mice. The numbers of anterior horn neurons of Nissl-, choline acetyltransferase (ChAT)-, and trkC-stained and wheat germ agglutinin-horseradish peroxidase (WGA-HRP)-labeled neurons at the spinal cord level with maximum compression were significantly higher in AdV-NT-3-transfected than in AdV-LacZ-transfected twy mice. Retrograde NT-3 gene transfer to twy mouse anterior horn neurons increased neurite axonal length and arborization of WGA-HRP-labeled neurons. Our results suggest that targeted retrograde NT-3 gene delivery is feasible in the intact animal and that it enhances neuronal survival even under chronic mechanical compression of the spinal cord.

Targeted spinal cord therapeutics delivery: stabilized platform and microelectrode recording guidance validation.

BACKGROUND/AIMS: No validated delivery technique exists for accurate, reproducible delivery of biological therapies to discrete spinal cord targets. To address this unmet need, we have constructed a stabilized platform capable of supporting physiologic mapping, through microelectrode recording, and cellular or viral payload delivery to the ventral horn. METHODS: A porcine animal model (n = 7) has been chosen based upon the inherent morphologic similarities between the human and porcine spine. Animals underwent physiologic mapping and subsequent microinjection of a green-fluorescent-protein-labeled cell suspension. Sacrifice (t = 3 h) was performed immediately following behavioral assessment. RESULTS: Histologic analysis has supported our ability to achieve localization to the ipsilateral ventral horn in the spinal cord. Complications included death due to malignant hyperthermia (n = 1), hindlimb dysfunction attributable to epidural hematoma (n = 1), and hindlimb dysfunction attributable to cord penetration (n = 2). CONCLUSIONS: These results indicate an ability to achieve accurate targeting, but the elevated incidence of neurologic morbidity will require further studies with longer follow-ups that incorporate procedural and equipment modifications that will allow for a reduced number of cord penetrations and will account for observed cardiorespiratory-associated cord movement. These initial results reinforce the challenges of translating biological restorative therapies from small to large animal models and ultimately to humans.

Rescue of rat anterior horn neurons after spinal cord injury by retrograde transfection of adenovirus vector carrying brain-derived neurotrophic factor gene.

We investigated the efficacy of retrograde gene delivery via the sternomastoid muscle of recombinant adenovirus vector (AdV) carrying brain-derived neurotrophic factor (BDNF) gene for the rescue of injured rat spinal cord. One hundred-thirty five adult Sprague-Dawley rats were used in the study with a standard weight-compression technique to produce spinal cord injury. AdV-BDNF gene or AdV-beta-galactosidase (AdV-LacZ) gene was injected into the sternomastoid muscle immediately after traumatic C4 segment spinal cord injury. AdV-BDNF was successfully appeared in the injured cervical spinal cord following injection into the sternomastoid muscle. BDNF expression in the anterior horn neurons of the cervical spinal cord reached peak levels at 1-2 weeks; and the expression persisted at significant levels for approximately 4 weeks after injury. AdV-BDNF transfection was associated with increased numbers of intact neurons as confirmed by Nissl, cholineacetyltransferase (ChAT), and acetylcholine esterase (AChE) staining especially from 2 weeks after injury, compared with the AdV-LacZ injected rats. Our results suggest that in vivo targeted retrograde AdV-BDNF-gene delivery may enhance neuronal survival following traumatic injury of the spinal cord.

ERG conductance expression modulates the excitability of ventral horn GABAergic interneurons that control rhythmic oscillations in the developing mouse spinal cord.

During antenatal development, the operation and maturation of mammalian spinal networks strongly depend on the activity of ventral horn GABAergic interneurons that mediate excitation first and inhibition later. Although the functional consequence of GABA actions may depend on maturational processes in target neurons, it is also likely that evolving changes in GABAergic transmission require fine-tuning in GABA release, probably via certain intrinsic mechanisms regulating GABAergic neuron excitability at different embryonic stages. Nevertheless, it has not been possible, to date, to identify certain ionic conductances upregulated or downregulated before birth in such cells. By using an experimental model with either mouse organotypic spinal cultures or isolated spinal cord preparations, the present study examined the role of the ERG current (I(K(ERG))), a potassium conductance expressed by developing, GABA-immunoreactive spinal neurons. In organotypic cultures, only ventral interneurons with fast adaptation and GABA immunoreactivity, and only after 1 week in culture, were transformed into high-frequency bursters by E4031, a selective inhibitor of I(K(ERG)) that also prolonged and made more regular spontaneous bursts. In the isolated spinal cord in which GABA immunoreactivity and m-erg mRNA were colocalized in interneurons, ventral root rhythms evoked by NMDA plus 5-hydroxytryptamine were stabilized and synchronized by E4031. All of these effects were lost after 2 weeks in culture or before birth in coincidence with decreased m-erg expression. These data suggest that, during an early stage of spinal cord development, the excitability of GABAergic ventral interneurons important for circuit maturation depended, at least in part, on the function of I(K(ERG)).

Rapid pH and PO2 changes in the tissue recording chamber during stoppage of a gas-equilibrated perfusate: effects on calcium currents in ventral horn neurons.

In vitro studies often use bicarbonate-buffered saline solutions to mimic the normal extracellular environment of tissues. These solutions are typically equilibrated with gaseous O2 and CO2, the latter interacting with bicarbonate ions to maintain a physiological pH. In vitro tissue chambers, like those used for electrophysiology, are usually continually perfused with the gassed buffer, but stopping the perfusion to add expensive chemicals or acquire imaging data is a common practice. The present study demonstrates that this procedure leads to rapid (< 30 s) increases in pH and decreases in PO2 of the detained solution in the tissue chamber. During the first 200 s, pH increased by 0.4 units and resulted in a 25% PO2 reduction of the detained solution. The rates of these changes were dependent on the volume of solution in the chamber. In experiments using acute transverse slices from the lumbar spinal cord of neonatal (postnatal day 0-10) mice, perfusion stoppage of the same duration was accompanied by a 34.7% enhancement of the peak voltage-gated calcium current recorded from ventral horn neurons. In these cells both low voltage-activated and high voltage-activated currents were affected. These currents were unaffected by decreasing PO2 when a CO2-independent buffer was used, suggesting that changes in pH were responsible for the observed effects. It is concluded that the procedure of stopping a bicarbonate/CO2-buffered perfusate results in rapid changes in pH and PO2 of the solution detained in the tissue chamber, and that these changes have the potential to covertly influence experimental results.

Polyglutamine tract-binding protein-1 dysfunction induces cell death of neurons through mitochondrial stress.

Polyglutamine tract-binding protein-1 (PQBP-1) is a nuclear protein that interacts and colocalizes with mutant polyglutamine proteins. We previously reported that PQBP-1 transgenic mice show a late-onset motor neuron disease-like phenotype and cell death of motor neurons analogous to human neurodegeneration. To investigate the molecular mechanisms underlying the motor neuron death, we performed microarray analyses using the anterior horn tissues of the spinal cord and compared gene expression profiles between pre-symptomatic transgenic and age-matched control mice. Surprisingly, half of the spots changed more than 1.5-fold turned out to be genes transcribed from the mitochondrial genome. Northern and western analyses confirmed up-regulation of representative mitochondrial genes, cytochrome c oxidase (COX) subunit 1 and 2. Immunohistochemistry revealed that COX1 and COX2 proteins are increased in spinal motor neurons. Electron microscopic analyses revealed morphological abnormalities of mitochondria in the motor neurons. PQBP-1 overexpression in primary neurons by adenovirus vector induced abnormalities of mitochondrial membrane potential from day 5, while cytochrome c release and caspase 3 activation were observed on day 9. An increase of cell death by PQBP-1 was also confirmed on day 9. Collectively, these results indicate that dysfunction of PQBP-1 induces mitochondrial stress, a key molecular pathomechanism that is shared among human neurodegenerative disorders.

[The primary observation of congenital clubfoot].

OBJECTIVE: To explore the possible correlations between clinical and experimental pathological changes of congenital clubfoot and the pathodynamic developmental procedure. METHODS: Eighty-three female Wistar rats were administered with retinoic acid on the 10th day after pregnancy. And from February 2001 to February 2004, 48 patients were analyzed with electropysiological examination. RESULTS: There was clubfoot-like deformity in 53.7% of the experimental fetuses. Persistence of the embryonic position of the talus and tibia in fetuses was observed. Poor overlapping between talus and calcaneus was seen. Cell apoptosis at the anterior corner of spinal cord were seen. Of all the patients, 68.3% were abnormal with electropysiological examination. The pathological sites were frequently localized in lumbarsacral region. CONCLUSION: Congenital clubfoot is correlated closely with defects of neural tube and spinal cord.

Enhanced functional recovery after proximal nerve root injury by vector-mediated gene transfer.

In order to test the functional implication of herpes simplex virus (HSV) vector-mediated gene transfer after axonal injury, we injected replication-incompetent HSV vectors coding for the anti-apoptotic peptide Bcl-2 and the glial cell-derived neurotrophic factor (GDNF), separately or in combination into ventral spinal cord 30 min after a crush injury to the proximal spinal root that was combined with moderate mechanical traction. HSV-mediated expression of Bcl-2 or GDNF enhanced functional recovery assessed by histologic, electrophysiologic, and behavioral parameters up to 5 months after injury. The most sensitive measure of distal motor function, the sciatic function index, was significantly improved in animals injected with the two vectors together. These results suggest an approach to root trauma that might be used to enhance functional recovery after injury.

Early decrease of survival factors and DNA repair enzyme in spinal motor neurons of presymptomatic transgenic mice that express a mutant SOD1 gene.

The primary pathogenetic mechanisms of amyotrophic lateral sclerosis (ALS) have been elusive. Some of the mechanisms would be implicated in an imbalance between death and survival factors, and impairment of DNA repair possibly caused by oxidative stress. Phosphatidylinositol 3-kinase (PI3-K) and its downstream effector, Akt/protein kinase B (PKB), have been shown to play a pivotal role in neuronal survival against apoptosis supported by neurotrophic factors. To elucidate the mechanisms of motor neuron death in ALS, we examined the expression of PI3-K, Akt, and the DNA repair enzyme redox factor-1 (Ref-1) protein in the spinal cord of transgenic mice with an ALS-linked mutant Cu/Zn superoxide dismutase (SOD1) gene, a valuable model for human ALS. Immunoblotting and immunocytochemical analyses showed that most spinal motor neurons lost immunoreactivity for PI3-K, Akt, and Ref-1 in the presymptomatic stage that preceded a significant loss of neurons. These results suggest that an early decrease of survival signal proteins and a DNA repair enzyme in the spinal motor neurons may account for the mutant SOD1-mediated motor neuron death in this animal model of ALS.

Giant repeater F-wave in patients with anterior horn cell disorders. Role of motor unit size.

Conventional F-wave responses as well as single motor unit F-wave responses together with the volitionally recruited motor unit action potentials (MUAP) were studied in hand and feet muscles of 10 healthy subjects and 32 patients with anterior horn cell disorders. The amplitude of the largest F-wave (Fl) was significantly greater in the affected patients compared with healthy subjects. Giant repeater F-wave responses "up to 4 mV" were recorded in muscles having volitionally recruited giant MUAPs. Although, the group mean percentage of motor unit F-wave responses per stimulation in all tested orthodromic MUAPs was significantly decreased in amyotrophic lateral sclerosis patients, the group mean percentage of motor unit F-wave responses per stimulation in all tested orthodromic MUAPs that gave motor unit F-wave response was significantly increased compared with healthy subjects. The responding orthodromic MUAP gave identical motor unit F-wave response, even for complex polyphasic units. Enhanced monosynaptic (H-) reflex, proximal axon reflex (A-wave), and repetitive muscle response as possible explanations for the giant F-wave responses could be discounted. The electrophysiologic behavior of the giant late responses described here fits well with the criteria of F-waves modulated by newly formed distal (and or proximal) axonal branching.

Paired pulse facilitation of GABAergic IPSCs in ventral horn neurons in neonatal rat spinal cord.

Whole-cell patch-clamp recording of GABAergic inhibitory postsynaptic currents (IPSCs) were made in ventral horn neurons of neonatal rat lumbar spinal cord in slice. In contrast to the hippocampus where paired pulse depression is reported to be observed for GABAergic IPSCs, double pulse stimulation of GABAergic inputs resulted in enhancement in the amplitude of the second IPSC in the spinal ventral horn. The facilitation ratio was decreased during enhanced synaptic transmission by increasing Ca2+ concentration in the external recording solution. Baclofen and adenosine. which are reported to depress synaptic transmission by presynaptic mechanisms, depressed IPSCs and increased the facilitation ratio. A postsynaptic manipulation such as application of bicuculline or changing the driving force did not affect the facilitation ratio. These results suggest that paired pulse facilitation of GABAergic IPSCs observed in neonatal rat spinal ventral horn appears to be based upon a mechanism similar to that underlying frequency-dependent facilitation of excitatory synaptic transmission, and is sensitive to presynaptic changes in synaptic strength.

Reinnervation of avulsed and reimplanted ventral rootlets in the cervical spinal cord of the cat.

Spinal nerve root avulsions frequently occur in brachial plexus injuries caused by traction. Such lesions are considered to afflict the central nervous system (CNS) and are, therefore, believed to be beyond surgical repair. The present experimental study was initiated to challenge this hypothesis. The ventral rootlets of C-7 were avulsed from the spinal cord in 28 cats via an anterior approach and subsequently reimplanted into the cord at the site of origin. In nonoperated control cats and cats undergoing reimplantation, electrophysiological experiments were performed and horseradish peroxidase was administered to the spinal nerve on the reimplanted side after survival times ranging from 6 to 293 days. Spinal cord sections in all cats were stained for neurofilament, acetylcholinesterase (AChE), Nissl, and glial fibrillary acidic protein. Horseradish peroxidase-labeled ventral horn motoneurons were found as early as 14 days after reimplantation and their number increased with time. On Days 209 and 293, the number of labeled neurons equaled the number of labeled ventral horn neurons in the two control cats that did not undergo surgery. Starting on Day 6 after reimplantation, the appearance of the ventral horn and the white matter in the neurofilament, AChE, and Nissl-stained sections changed as a result of the CNS response to the injury. A return to their normal appearance could be observed in these stainings from Day 209 onward. Glial fibrillary acidic protein-positive astrocytic tissue was consistently found in the ventral horn and in the white matter reimplantation area. From Day 69 onward, electrophysiological stimulation of the spinal nerve C-7 on the reimplanted side elicited an electromyogram response in the spinodeltoid muscle. The latency and threshold intensity of the C-7 responses were initially increased but equalized to match the nonoperated controls between 98 and 122 days after reimplantation. The results of this study show that functional regeneration of ventral horn neurons after root avulsion and subsequent reimplantation in the cat is possible.