Lithium causes differential effects on postsynaptic stability in normal and denervated neuromuscular synapses.

Lithium chloride has been widely used as a therapeutic mood stabilizer. Although cumulative evidence suggests that lithium plays modulatory effects on postsynaptic receptors, the underlying mechanism by which lithium regulates synaptic transmission has not been fully elucidated. In this work, by using the advantageous neuromuscular synapse, we evaluated the effect of lithium on the stability of postsynaptic nicotinic acetylcholine receptors (nAChRs) in vivo. We found that in normally innervated neuromuscular synapses, lithium chloride significantly decreased the turnover of nAChRs by reducing their internalization. A similar response was observed in CHO-K1/A5 cells expressing the adult muscle-type nAChRs. Strikingly, in denervated neuromuscular synapses, lithium led to enhanced nAChR turnover and density by increasing the incorporation of new nAChRs. Lithium also potentiated the formation of unstable nAChR clusters in non-synaptic regions of denervated muscle fibres. We found that denervation-dependent re-expression of the foetal nAChR γ-subunit was not altered by lithium. However, while denervation inhibits the distribution of ß-catenin within endplates, lithium-treated fibres retain ß-catenin staining in specific foci of the synaptic region. Collectively, our data reveal that lithium treatment differentially affects the stability of postsynaptic receptors in normal and denervated neuromuscular synapses in vivo, thus providing novel insights into the regulatory effects of lithium on synaptic organization and extending its potential therapeutic use in conditions affecting the peripheral nervous system.

Evaluación externa de los cambios funcionales y la marcha tras una sesión de miofibrotomía múltiple en escolares con diplejía espástica / External evaluation of gait and functional changes after a single-session multiple myofibrotenotomy in school-aged children with spastic diplegia

Introducción. Unos 23.000 pacientes con espasticidad han optado voluntariamente por la miofibrotomía múltiple (MFM), una técnica alternativa consistente en seccionar tejidos blandos para liberar restricciones articulares. Iniciada por Ulzibat (Rusia), se realiza fuera de controles clínicos ortodoxos. Objetivo. Evaluar externamente los efectos de la MFM sobre la funcionalidad motora y la marcha. Pacientes y métodos. Estudio observacional autocontrolado que evalúa cambios en variables funcionales -Gross Motor Function Classification System, dimensión E de la Gross Motor Function Measure (GMFM) y Functional Mobility Scale- y 32 parámetros de marcha (medidos mediante análisis instrumental) en 22 escolares con diplejía espástica (edad mediana: 9 años y 6 meses; Q1-Q3: 7 años y 11 meses a 11 años y 6 meses) cuyos padres optaron por una MFM (mediana de observación: 4 meses; rango: 3-7 meses). Resultados. El análisis de conglomerados jerárquicos utilizado para determinar patrones topográficos de cortes en los pacien s detectó que los cirujanos aplicaban a cada paciente uno de tres conjuntos de cortes. Analizados los tres grupos de pacientes, se observó que un grupo empeoró globalmente y una mejora significativa general en la dimensión E del GMFM (diferencia de mediana: 4,86%; intervalo de confianza al 95% = 0-6,94%) tras la MFM. Algunos parámetros de la marcha se normalizaron significativamente (izquierda: rango flexión-cadera, máxima dorsiflexión en apoyo; derecha: velocidad, rotación pélvica media en apoyo y máxima dorsiflexión en apoyo). Conclusiones. Estos resultados no apoyan ni justifican el uso de la MFM como opción para el tratamiento de la espasticidad. Insistimos en que esta técnica debe evitarse fuera de protocolos de estudio. Sin embargo, los resultados asientan una base objetiva para justificar la realización de un ensayo clínico y estudios observacionales a largo plazo (AU)

Introduction. About 23,000 patients with spasticity voluntarily chose to undergo a multiple myofiberotomy (MMF), which is an alternative technique consisting in sectioning soft tissues in order to relieve restrictions in joint movements. This technique, first employed by Ulzibat (Russia), is performed outside orthodox clinical controls. Aims. To perform an external evaluation of the effects of MMF on motor functionality and gait. Patients and methods. The study was self-controlled and observation-based and was designed to evaluate the changes in functional variables -Gross Motor Function Classification System, E-dimension of the Gross Motor Function Measure (GMFM) and the Functional Mobility Scale- and 32 gait parameters (measured using analytical instruments) in 22 schoolchildren with spastic diplegia (mean age: 9 years and 6 months; Q1-Q3: 7 years and 11 months to 11 years and 6 months) whose parents opted for an MMF (median of observation: 4 months; range: 3-7 months). Results. The analysis of hierarchi l conglomerates used to determine topographic cut patterns in patients revealed that the surgeons applied one of three sets of cuts to each patient. On analysing the three groups of patients, it was observed that one group worsened overall and another underwent a general significant improvement in the E-dimension of the GMFM (difference of median: 4.86%; 95% confidence interval = 0-6.94%) after the MMF. Some of the gait parameters became significantly normalised (left: hip-flexion range, maximum dorsiflexion with support; right: speed, mean pelvic rotation with support and maximum dorsiflexion with support). Conclusions. These findings do not back up or justify the use of MMF as a therapeutic option to treat spasticity. It has to be stressed that this technique must be avoided outside study protocols. Nevertheless, results do lay an objective base that may justify a clinical trial and long-term observation-based studies (AU)

Severe neuromuscular denervation of clinically relevant muscles in a mouse model of spinal muscular atrophy.

Spinal muscular atrophy (SMA), a motoneuron disease caused by a deficiency of the survival of motor neuron (SMN) protein, is characterized by motoneuron loss and muscle weakness. It remains unclear whether widespread loss of neuromuscular junctions (NMJs) is involved in SMA pathogenesis. We undertook a systematic examination of NMJ innervation patterns in >20 muscles in the SMNΔ7 SMA mouse model. We found that severe denervation (<50% fully innervated endplates) occurs selectively in many vulnerable axial muscles and several appendicular muscles at the disease end stage. Since these vulnerable muscles were located throughout the body and were comprised of varying muscle fiber types, it is unlikely that muscle location or fiber type determines susceptibility to denervation. Furthermore, we found a similar extent of neurofilament accumulation at NMJs in both vulnerable and resistant muscles before the onset of denervation, suggesting that neurofilament accumulation does not predict subsequent NMJ denervation. Since vulnerable muscles were initially innervated, but later denervated, loss of innervation in SMA may be attributed to defects in synapse maintenance. Finally, we found that denervation was amendable by trichostatin A (TSA) treatment, which increased innervation in clinically relevant muscles in TSA-treated SMNΔ7 mice. Our findings suggest that neuromuscular denervation in vulnerable muscles is a widespread pathology in SMA, and can serve as a preparation for elucidating the biological basis of synapse loss, and for evaluating therapeutic efficacy.

Electrophysiological characterization of neuromuscular synaptic dysfunction in mice.

Emerging evidence suggests that synaptic dysfunction occurs prior to neuronal loss in neurodegenerative diseases, such as amyotrophic lateral sclerosis (ALS). Therefore, monitoring synaptic activity during early stages of neurodegeneration may provide valuable information for the development of diagnostic and/or therapeutic strategies. Here, we describe an electrophysiological method routinely applied in our laboratory for investigating synaptic activity of the neuromuscular junction (NMJ), the synaptic connection between motoneurons and skeletal muscles. Using conventional intracellular sharp electrodes, both spontaneous synaptic activity (miniature end-plate potentials) and evoked synaptic activity (end-plate potentials) can be readily recorded in acutely isolated nerve-muscle preparations. This method can also be adapted to various simulation protocols for studying short-term plasticity of neuromuscular synapses.

Nerve-muscle-endplate band grafting: a new technique for muscle reinnervation..

BACKGROUND: Because currently existing reinnervation methods result in poor functional recovery, there is a great need to develop new treatment strategies. OBJECTIVE: To investigate the efficacy of our recently developed nerve-muscle-endplate band grafting (NMEG) technique for muscle reinnervation. METHODS: Twenty-five adult rats were used. Sternohyoid (SH) and sternomastoid (SM) muscles served as donor and recipient muscle, respectively. Neural organization of the SH and SM muscles and surgical feasibility of the NMEG technique were determined. An NMEG contained a muscle block, a nerve branch with nerve terminals, and a motor endplate band with numerous neuromuscular junctions. After a 3-month recovery period, the degree of functional recovery was evaluated with a maximal tetanic force measurement. Retrograde horseradish peroxidase tracing was used to track the origin of the motor innervation of the reinnervated muscles. The reinnervated muscles were examined morphohistologically and immunohistochemically to assess the extent of axonal regeneration. RESULTS: Nerve supply patterns and locations of the motor endplate bands in the SH and SM muscles were documented. The results demonstrated that the reinnervated SM muscles gained motor control from the SH motoneurons. The NMEG technique yielded extensive axonal regeneration and significant recovery of SM muscle force-generating capacity (67% of control). The mean wet weight of the NMEG-reinnervated muscles (87% of control) was greater than that of the denervated SM muscles (36% of control). CONCLUSION: The NMEG technique resulted in successful muscle reinnervation and functional recovery. This technique holds promise in the treatment of muscle paralysis.

Dissection and imaging of active zones in the Drosophila neuromuscular junction.

The Drosophila larvae neuromuscular junction (NMJ) is an excellent model for the study of synaptic structure and function. Drosophila is well known for the ease of powerful genetic manipulations and the larval nervous system has proven particularly useful in studying not only normal function but also perturbations that accompany some neurological disease (Lloyd and Taylor, 2010). Many key synaptic molecules found in Drosophila are also found in mammals and like most CNS excitatory synapses in mammals, the Drosophila NMJ is glutamatergic and demonstrates activity-dependent remodeling (Koh et al. , 2000). Additionally, Drosophila neurons can be individually identified because their innervation patterns are stereotyped and repetitive making it possible to study identified synaptic terminals, such as those between motor neurons and the body-wall muscle fibers that they innervate (Keshishian and Kim, 2004). The existence of evolutionarily conserved synapse components along with the ease of genetic and physical manipulation make the Drosophila model ideal for investigating the mechanisms underlying synaptic function (Budnik, 1996). The active zones at synaptic terminals are of particular interest because these are the sites of neurotransmitter release. NC82 is a monoclonal antibody that recognizes the Drosophila protein Bruchpilot (Brp), a CAST1/ERC family member that is an important component of the active zone (Wagh et al. , 2006). Brp was shown to directly shape the active zone T-bar and is responsible for effectively clustering Ca(2+) channels beneath the T-bar density (Fouquet et al. , 2009). Mutants of Brp have reduced Ca(2+) channel density, depressed evoked vesicle release, and altered short-term plasticity (Kittel et al., 2006). Alterations to active zones have been observed in Drosophila disease models. For example, immunofluorescence using the NC82 antibody showed that the active zone density was decreased in models of amyotrophic lateral sclerosis and Pitt-Hopkins syndrome (Ratnaparkhi et al. , 2008; Zweier et al. , 2009). Thus, evaluation of active zones, or other synaptic proteins, in Drosophila larvae models of disease may provide a valuable initial clue to the presence of a synaptic defect. Preparing whole-mount dissected Drosophila larvae for immunofluorescence analysis of the NMJ requires some skill, but can be accomplished by most scientists with a little practice. Presented is a method that provides for multiple larvae to be dissected and immunostained in the same dissection dish, limiting environmental differences between each genotype and providing sufficient animals for confidence in reproducibility and statistical analysis.

[Reconstruction of defects at the neuromuscular junction]. / Rekonstruktion bei Defekten im neuromuskulären Ubergang.

Loss of muscle tissue at the area of the neuromuscular junction after tumor resection or after trauma precludes the reconstruction with conventional nerve grafts, because the distal nerve stump is absent. For these cases, we recommend direct insertion of the nerve grafts into the muscle. We describe a standardized technique, which has been performed in 19 patients and led to a mean motor recovery of grade M4 after Highet. The key procedure of this technique is the interfascicular dissection of the nerve grafts, which allows a wide distribution of the grafts into the muscle tissue.

Promoting nerve regeneration through long gaps using a small nerve tissue graft.

BACKGROUND: If nerve tissue is capable of inducing regeneration, as suggested by the neurotropism theory, then even small pieces of nerve tissue should have the potential to induce nerve regeneration. Therefore, long gaps might presumably be bridged via the neurotrophic potential of small pieces of nerve tissue grafted into the middle of the nerve gap. It is necessary to confirm the validity of the neurotropism theory and to also explore the potential usefulness of small nerve grafting through long gaps. METHODS: A small piece of nerve tissue was grafted into a silicone tube bridging a relatively long nerve gap in an attempt to promote nerve regeneration. A 15-mm gap was created in the left sciatic nerve of 31 Wistar rats (8 weeks of age). The experimental groups included one with nonvascularized nerve tissue grafted into a silicone tube with no distal nerve suturing (NV-A), another with vascularized nerve tissue grafted into a silicone tube with no distal nerve suturing (V-A), a third group with nonvascularized nerve tissue grafted into a silicone tube with distal nerve suturing (NV-P), a fourth group with vascularized nerve tissue grafted into a silicone tube with distal nerve suturing (V-P), and a group with no nerve segment grafted into the silicone tube (control). Electrophysiologic and histologic examinations were performed 10 weeks after the operation. RESULTS: No regeneration was obtained in the control group. Nerve regeneration was evident at the proximal end of the tube in the NV-A, V-A, NV-P, and V-P groups, and at the distal end in the NV-P and V-P groups. The degree of distal regeneration was extremely slight in the NV-A and V-A groups. An electrophysiologic examination performed in the NV-P and V-P groups revealed better results in the latter group. CONCLUSION: Small nerve grafts are capable of inducing nerve regeneration even over a long nerve gap, by grafting nerve tissue into the middle of the lesion using a silicone tube.

Refinements in nerve to muscle neurotization.

A modified surgical technique is introduced, enabling restoration of muscle function with direct muscular neurotization. Reliable clinical outcomes result from this technique. We report on a series of 10 patients in whom the supplying motor nerve had been lost at the level of the neuromuscular junction as the result of trauma or tumor resection. Our modification of the operative technique ensures a wide distribution of nerve fibers throughout the remaining muscle tissue and produces a mean motor recovery of M4 after a period of 1 to 2 years.

Identification and catheter ablation of extracardiac and intracardiac components of ligament of Marshall tissue for treatment of paroxysmal atrial fibrillation.

INTRODUCTION: The ligament of Marshall is a left atrial neuromuscular bundle with sympathetic innervation that may be a source of atrial fibrillation (AF)-inducing automatic activity. METHODS AND RESULTS: Twenty-four patients with paroxysmal AF (including 18 with adrenergic AF) and 25 with other arrhythmias underwent catheter mapping. In cases of adrenergic AF, radiofrequency ablation was attempted when Marshall potentials were recorded. Patients were followed for 2 months before and 11.2 +/- 4.2 months after the procedure. Catheterization of the distal superoposterior coronary sinus was feasible in 14 patients with AF (10 with adrenergic AF) and 12 patients without AF. A discrete Marshall potential was recorded in 12 patients with AF versus 3 patients without AF (P = 0.004). In 10 patients with adrenergic AF, this potential followed the atrial electrogram during sinus rhythm by 26 +/- 5 msec on left atrial recordings and 24 +/- 4 msec on coronary sinus recordings, and preceded it during atrial ectopy by 29 +/- 5 msec and 26 +/- 5 msec, respectively. It was abolished by epicardial (n = 1), endocardial (n = 4), or combined epicardial and endocardial ablation (n = 5). Seven patients with ablation showed significant reductions in adrenergic AF, whereas no significant change was seen in 8 adrenergic AF patients not undergoing ablation (P = 0.004). No improvement was seen in 3 of 4 patients with only endocardial ablation, whereas all 6 patients with epicardial ablation improved (P = 0.033). CONCLUSION: Recording of Marshall potential is feasible in patients with paroxysmal AF. Combined epicardial and endocardial catheter ablation of ligament of Marshall tissue may reduce the paroxysms of adrenergic AF.

Morphologic correlates for laryngeal reinnervation.

OBJECTIVE: To describe morphologic correlates for laryngeal reinnervation. STUDY DESIGN: Review of anatomic experiments dealing with laryngeal innervation performed over a 25-year period. METHODS: Description of results from experimental studies on the cat and human laryngeal muscles and nerve supply. RESULTS: Despite separation of abductor and adductor laryngeal motor neurons in the central nervous system, the mixture of abductor and adductor axons in the recurrent laryngeal nerve indicates that selective re-innervation of an individual laryngeal muscle must be accomplished at the neuromuscular junction (NMJ) of the muscle. The optimal time for a reinnervating neural source to re-occupy vacated NMJ is at the time of denervation. If the reinnervation procedure is attempted long (>1 mo) after denervation, extraneous end plates of other neural systems must be eliminated to provide vacant NMJ. The nerve muscle pedicle (NMP) concept is an effective model for reinnervation of a laryngeal muscle provided its activity pattern is similar to that of the denervated muscle and its insertion into vacated NMJ is timely. CONCLUSION: NMP offers a logical method for selective laryngeal muscle reinnervation. Critical to the success of NMP are the physiological input to the NMP and timing of NMP implantation.

Schwann cells induce and guide sprouting and reinnervation of neuromuscular junctions.

The "terminal' Schwann cells that sit atop the neuromuscular junction sense neuromuscular transmission and respond to perturbations of this transmission by extending long processes. These processes have the ability to induce nerve growth and serve as substrates to guide this growth. These processes thus play major roles in muscle reinnervation and in sprouting. An absence of nerve sprouting is correlated with the apoptotic death of terminal Schwann cells at denervated endplates in neonatal muscles. Thus, Schwann cells appear to participate actively in the maintenance and repair of neuromuscular synapses.

[Neuro-vascular decompression in hemifacial spasm: anatomical, electrophysiological and therapeutic results apropos of 100 cases]. / Décompression neuro-vasculaire dans le spasme de l'hémiface: résultats anatomiques, électrophysiologiques et thérapeutiques à propos de 100 cas.

Hemifacial spasm is a neurological disorder due to abnormal hyperactivity of the facial nerve. The most common cause of hemifacial spasm is a neuro-vascular conflict in the cerebellopontine angle between a vascular loop and the root of the facial nerve (96% of cases). Tumors are the cause of hemifacial spasm in only 1% of cases). The authors present their results in 100 patients who underwent microvascular decompression for essential hemifacial spasm between 1990 and 1995. They used microsurgical and endoscopic procedures by a minimal retrosigmoid approach in all cases. The most common offending vessels were the posterior inferior cerebellar artery (70%), the vertebral artery (41%) and the anterior inferior cerebellar artery (28%). An aberrant vein was found in 2 cases. There were 38% of multiple artery-nerve conflicts. Physiopathology of hemifacial spasm is explained by two principal theories: in the ephaptic theory, hyperactivity and an abnormal nervous impulse pathway are due to a short demyelinated area on the nerve trunk caused by the offending vessel, inducing short circuiting between adjacent nerve fibers. In the nuclear theory, hyperactivity of the facial nerve is due to an abnormal and automatic activity of the facial nerve nucleus itself, induced by the vessel. The authors used pre and postoperative electromyographic tests and intraoperative electromyographic tests. Their results tend to prove the nuclear theory. Ninety per cent of the patients had a good result, with a mean follow-up time of 30 months in 60 cases. In 82% of the cases, there was a total recovery after a single procedure. There was no mortality and no facial palsy. Hearing loss occurred in less than 5%.

Neuromuscular pedicle graft for restoration of arytenoid abductor function in dogs with experimentally induced laryngeal hemiplegia.

Left laryngeal hemiplegia was induced by resection of the left recurrent laryngeal nerve in 12 dogs. A neuromuscular pedicle graft formed from the first cervical nerve and sternothyroideus muscle was transplanted after 1 week to the denervated cricoarytenoideus dorsalis muscle in 8 dogs. The remaining 4 dogs served as controls. Left arytenoid abduction was blindly evaluated by laryngoscopy with video photography at time 0, at 1 week, and at 19 weeks in all dogs. At 19 weeks, biopsy specimens of the left cricoarytenoideus dorsalis muscle and the neuromuscular pedicle were taken from 4 of the treatment dogs, and biopsy specimens of the left cricoarytenoideus dorsalis muscle were taken from the 4 control dogs. All biopsy specimens were blindly evaluated by histologic and histochemical examination. At 36 to 44 weeks, the remaining 4 treatment dogs, from which biopsy specimens had not been taken, were reevaluated by use of laryngoscopy with video photography. Complications and difficulties encountered during surgery included hemorrhage in the area of the cricoarytenoideus dorsalis muscle, location of a branch of the first cervical nerve that was long enough to prevent tension at the graft site, orientation of the muscle pedicle in the cricoarytenoideus dorsalis muscle without the use of an operating microscope, and preservation of the terminal portion of the first cervical nerve while forming the neuromuscular pedicle. Results of the arytenoid movement evaluations revealed improvement in arytenoid abductor function in the treatment group, compared with that in the control group at 19 weeks. Arytenoid abduction in the treatment group at this time, however, was still significantly decreased (P less than 0.05), compared with presurgical movement evaluations.(ABSTRACT TRUNCATED AT 250 WORDS)

Preliminary observations after facial rehabilitation with the ansa hypoglossi pedicle transfer.

Facial paralysis is a very disabling condition, both functionally and cosmetically. Despite the different methods of facial reanimation that have been described, there is no single method that will restore normal facial tone and motion. Of the methods available, primary neurorrhaphy is probably the most effective. The recovery period, however, is prolonged and, as a result, muscle tone and bulk may be lost. The hypoglossal-facial anastomosis is also a very reliable and effective technique but requires necessary interruption of both major cranial nerve trunks. Transfer of a neuromuscular pedicle (based on the ansa hypoglossi) has been offered as a method of facial reanimation that involves neither prolonged recovery nor interruption of the major cranial nerve functions. The application of this technique for reinnervation of a paralyzed larynx was first described by Tucker in 1970, and the technique was applied to facial muscle (in animal models) in 1977. The effectiveness of this technique--and its application in the management of facial paralysis in the human patient--remains controversial. We report our experience with a series of six patients who underwent neuromuscular pedicle transfer in conjunction with other more conventional methods of facial reanimation. The function of the pedicle and its contribution to the overall facial rehabilitation were assessed clinically and electromyographically. Factors influencing the success of the procedure and clinical and experimental evidence to support its application are discussed. While our experience with this technique is limited, it would appear that the neuromuscular pedicle transfer may play a useful adjunctive role in reanimation of the face in selective cases of facial paralysis.

Dual simultaneous systems for facial reanimation.

Successful reanimation of the paralyzed face requires a specific yet adaptable procedural armamentarium. Usually, in the treatment of regional paralysis, one distinct technique is deemed most appropriate and dependable. In cases of total hemiparesis secondary to surgical ablation or trauma, however, the simultaneous use of two separate but complementary rehabilitative systems has proved valuable in 15 patients. The reconstructive concept described divides the face into two functional spheres, an upper periorbital area and a lower perioral region. The integral system includes a direct facial nerve-to-cable graft reanastomosis for the upper division combined with a masseter muscle transfer for the lower facial region. The immediate supportive effects of the masseter transposition integrated with its long-term ability to rehabilitate via myoneurotization complement the more physiologically exacting effects of the nerve anastomosis.

Horseradish peroxidase studies in animals with neuromuscular transpositions.

Horseradish peroxidase (HRP) is used to trace axonal connections from the motor end-plate to the driving neuron. This technique has confirmed that the neurons activating the sternothyroid muscle are located in the cervical spinal cord, while those controlling the posterior cricoarytenoid (PCA) are found in the nucleus ambiguus ipsilaterally. Eight rabbits underwent a sternothyroid ansa pedicle implantation to the PCA at the time of sectioning the recurrent laryngeal nerve ipsilaterally. After two months, four of these animals received HRP injections into the previously implanted PCA. Brainstem staining HRP did not reveal any retrograde transport to the motor neurons that were known to control the sternothyroid. Possible for the failure of retrograde transport are discussed.