Anterior Horn Neurons Research Articles

F-waves in neonates: increased spinal anterior horn motor neuron excitability

We analyzed characteristics of F-waves in neonates to establish normal parameters for this age. F-waves recorded from the median nerve in 26 normal neonates were analyzed and compared to those in 10 adults. F-wave parameters measured included occurrence rate (Fpersistence), latency (Flatency), latency corrected for subject height (normalized Flatency), duration (Fduration), amplitude (Famplitude) and its ratio to M-wave amplitude (F/M ratio), F-wave conduction velocity (FCV), and Fchronodispersion. Mean Fpersistence in neonates was 100%. Famplitude and the F/M ratio were higher in neonates than in adults. Similar F-wave waveforms were observed repeatedly in neonates, while waveforms varied in adults. These results in neonates suggest increased excitability of motor neurons in the spinal anterior horn reflecting immaturity of inhibition by the central nervous system.

Early decrease of survival signal protein and DNA repair enzyme in spinal motor neuron of presymptomatic transgenic mice with a mutant SOD1 gene

We examined expressions of phosphatidylinositol 3-kinase (PI3-K), Akt and redox factor-1 protein (Ref-1) in the spinal cord of transgenic mice with G93A mutant superoxide dismutase (SOD1) (Tg), a valuable model of human amyotrophic lateral sclerosis (ALS), at early and presymptomatic stage, 18 and 25 weeks of age. Significant decrease of PI3-K, and Akt in Tg/18 mice, and of PI3-K, Akt and Ref-1 in Tg/25 was observed on immunoblotting. Immunohistochemistry revealed the significant decrease of PI3-K, Akt and Ref-1 in spinal motor neurons of Tg/25 mice. No significant loss of anterior horn neurons was detected. Therefore, the decrease of survival signal or impairment of DNA repair in the spinal motor neurons precedes a subsequent death of anterior horn neurons, and may account for the mutant SOD1-mediated motor neuronal death in this model.

Classical infantile spinal muscular atrophy with SMN deficiency causes sensory neuronopathy

Classic infantile spinal muscular atrophy (SMA) is believed to be a purely motor disorder, affecting neurons of the spinal anterior horn and nuclei of the lower cranial nerves. Other organ malformations or peripheral nerve involvement have been regarded as exclusion criteria for infantile SMA. Whether SMN protein deficiency can also lead to loss of sensory neurons has not been systematically addressed. The authors evaluated the sural nerve biopsies of 19 patients with infantile SMA of varying severity. The diagnosis of SMA was confirmed by the presence of a homozygous deletion of the SMN1 gene in all patients. In seven unrelated infants with SMA type I, axonal degeneration of the sural nerve was noted. Five patients showed abnormal sensory conduction, thus prompting sural nerve biopsy. Sural nerves showed different degrees of axonal loss: fiber density ranged from 3.482 to 22.076/mm2 and was markedly reduced in four patients. There was no evidence of primary demyelination: the ratio of total myelinated fiber thickness to axon diameter (g-ratio) was normal in the patients examined. In seven patients with SMA II and five patients with SMA III, no sural nerve alterations were seen, and conduction velocity was normal. In addition to SMN1 gene deletions, homozygous NAIP gene deletions were detected in six out of seven infants with peripheral neuropathy, whereas there was no evidence of a large deletion including the multicopy markers C212 and Ag1-CA in two out of three families tested. In this series of patients with SMA I through III who underwent sural nerve biopsy, there was significant sensory nerve pathology in severely affected patients with SMA type I, whereas there were no sensory nerve alterations clinically or morphologically in patients with milder SMA type II or III.

Abstracts of the 8th Meeting of the Italian Peripheral Nerve Study Group: 8

A process of motor units (MU) rearrangement may compensate the loss of motor neurons in spinal cord anterior horns in Amyotrophic Lateral Sclerosis (ALS), at least in the early stages of the disease. Standard electromyography is unable to precisely quantify these pathological changes. Recently, Motor Unit Number Estimation (MUNE) has been introduced in clinical practice in order to provide a quantitative measure of the process as well as short‐ and long‐term follow‐up data. We investigated 8 male patients (mean age ± SD: 64 ± 10.9 yrs, range 45–78) affected by definite or probable ALS according to El Escorial Criteria. In order to quantify the total number of motor units, MUNE was performed during the diagnostic work‐up (basal, T0), after three (T1) and six months (T2), in abductor digiti minimi (ADM) and tibialis anterior (TA) muscles. We found a significant decrease in the mean number of motor units (16.6% at T1 [p < 0.002] and 37.9% at T2 [p < 0.04]), recording from ADM, while the decrease in mean and maximal area did not reach the statistical significance at the same time intervals. Similarly, we found a significant decrease in the mean number of motor units (20.6% at T1 [p < 0.008] and 40.3% at T2 [p < 0.0006]), but not in the mean and maximal area, recording from TA. According to previous data, we found about a loss of about 50% motor units in both the muscles six months after diagnosis. We conclude that MUNE is a useful tool to quantify MU loss in ALS and may result helpful in following the course of ALS and in predicting the prognosis.

Interleukin-6 Protects Anterior Horn Neurons from Lethal Virus-Induced Injury

We evaluated the role of interleukin-6 (IL-6) in neuronal injury after CNS infection. IL-6-/- and IL-6+/+ mice of resistant major histocompatibility complex (MHC) H-2b haplotype intracerebrally infected with Theiler's virus cleared the infection normally without development of viral persistence, lethal neuronal infection, or late phase demyelination. In contrast, infection of IL-6-/- mice on a susceptible H-2q haplotype resulted in frequent deaths and severe neurologic deficits within 2 weeks of infection as compared with infected IL-6+/+ H-2q littermate controls. Morphologic analysis demonstrated dramatic injury to anterior horn neurons of IL-6-/- H-2q mice at 12 d after infection. Infectious viral titers in the CNS (brain and spinal cord combined) were equivalent between IL-6-/- H-2q and IL-6+/+ H-2q mice. In contrast, more viral RNA was detected in the spinal cord of IL-6-/- mice compared with IL-6+/+ H-2q mice. Virus antigen was localized predominantly to anterior horn cells in infected IL-6-/- H-2q mice. IL-6 deletion did not affect the humoral response directed against virus, nor did it affect the expression of CD4, CD8, MHC class I, or MHC class II in the CNS. Importantly, IL-6 was expressed by astrocytes of infected IL-6+/+ mice but not in astrocytes of IL-6-/- mice or uninfected IL-6+/+ mice. Furthermore, expression of various chemokines was robust at 12 d after infection in both H-2b and H-2q IL-6-/- mice, indicating that intrinsic CNS inflammatory responses did not depend on the presence of IL-6. Finally, in vitro analysis of virus-induced death in neuroblastoma-spinal cord-34 motor neurons and primary anterior horn cell neurons showed that IL-6 exerted a neuroprotective effect. These data support the hypothesis that IL-6 plays a critical role in protecting specific populations of neurons from irreversible injury.

Sleep disorder in alien hand syndrome

A 63-year-old right-handed woman developed an alien hand syndrome (AHS) after an acute infarction in the territory of the left anterior cerebral artery. The uncontrolled hand movements were present during the daytime and eventually disturbed sleep. Polysomnography revealed that these motor actions only appeared when the patient was awake. These awakenings emerged mostly from NREM sleep stage 2 during the first half of the night. There was no evidence of any epileptiform activity, dyssomnia or parasomnia. These movements were controlled making her wear an oven mitt during sleep. The temporal distribution of this motor activity seems to follow the progressive hyperpolarization of anterior horn neurons that occurs when sleep deepens. The accommodation of the grasp reflex in AHS probably helps control this unwanted motor activity.

Neuroprotective effects of glial cell line-derived neurotrophic factor mediated by an adeno-associated virus vector in a transgenic animal model of amyotrophic lateral sclerosis.

Amyotrophic lateral sclerosis (ALS) is a relentlessly progressive lethal disease that involves selective annihilation of motoneurons. Glial cell line-derived neurotrophic factor (GDNF) is proposed to be a promising therapeutic agent for ALS and other motor neuron diseases. Because adeno-associated virus (AAV) has been developed as an attractive gene delivery system with proven safety, we explored the therapeutic efficacy of intramuscular delivery of the GDNF gene mediated by an AAV vector (AAV-GDNF) in the G93A mouse model of ALS. We show here that AAV-GDNF leads to substantial and long-lasting expression of transgenic GDNF in a large number of myofibers with its accumulation at the sites of neuromuscular junctions. Detection of GDNF labeled with FLAG in the anterior horn neurons, but not beta-galactosidase expressed as a control, indicates that most of the transgenic GDNF observed there is retrogradely transported GDNF protein from the transduced muscles. This transgenic GDNF prevents motoneurons from their degeneration, preserves their axons innervating the muscle, and inhibits the treated-muscle atrophy. Furthermore, four-limb injection of AAV-GDNF postpones the disease onset, delays the progression of the motor dysfunction, and prolongs the life span in the treated ALS mice. Our finding thus indicates that AAV-mediated GDNF delivery to the muscle is a promising means of gene therapy for ALS.

Neuroprotective Effects of Glial Cell Line-Derived Neurotrophic Factor Mediated by an Adeno-Associated Virus Vector in a Transgenic Animal Model of Amyotrophic Lateral Sclerosis

Amyotrophic lateral sclerosis (ALS) is a relentlessly progressive lethal disease that involves selective annihilation of motoneurons. Glial cell line-derived neurotrophic factor (GDNF) is proposed to be a promising therapeutic agent for ALS and other motor neuron diseases. Because adeno-associated virus (AAV) has been developed as an attractive gene delivery system with proven safety, we explored the therapeutic efficacy of intramuscular delivery of the GDNF gene mediated by an AAV vector (AAV-GDNF) in the G93A mouse model of ALS. We show here that AAV-GDNF leads to substantial and long-lasting expression of transgenic GDNF in a large number of myofibers with its accumulation at the sites of neuromuscular junctions. Detection of GDNF labeled with FLAG in the anterior horn neurons, but not beta-galactosidase expressed as a control, indicates that most of the transgenic GDNF observed there is retrogradely transported GDNF protein from the transduced muscles. This transgenic GDNF prevents motoneurons from their degeneration, preserves their axons innervating the muscle, and inhibits the treated-muscle atrophy. Furthermore, four-limb injection of AAV-GDNF postpones the disease onset, delays the progression of the motor dysfunction, and prolongs the life span in the treated ALS mice. Our finding thus indicates that AAV-mediated GDNF delivery to the muscle is a promising means of gene therapy for ALS.

The prion protein in human neurodegenerative disorders

We evaluate cellular prion protein (PrP C) immunoreactivity (IR) in Alzheimer's, Parkinson's, diffuse Lewy body, and motor neuron diseases (MND), progressive supranuclear palsy, and multiple system atrophy. We use immunohistochemistry for PrP, including five monoclonal antibodies against different epitopes and three different pretreatments, alpha-synuclein, phosphorylated tau, beta-amyloid, and ubiquitin. Disease-specific inclusions are devoid of PrP C IR. Using double immunofluorescence and confocal laser microscopy we observe focal overlapping of PrP C with tau and with alpha-synuclein in early, but not in fully developed inclusions. However, PrP C IR neurons may contain abnormal tau or alpha-synuclein aggregates. Additionally, we observe a loss of PrP C IR in anterior horn neurons in MND. Our results suggest that expression of PrP C reflects a general response to cellular stress rather than specific co-operation in aggregation of other proteins.

Masses of phosphorylated neurofilaments are associated with abnormal golgi apparatus of anterior horn neurons of beta, beta'-iminodipropionitrile-intoxicated rats.

The Golgi apparatus (GA) of anterior horn neurons of rats chronically intoxicated with beta,beta'-iminodipropionitrile (IDPN) in drinking water was examined with an organelle-specific antibody. The neuropile of the anterior horns contained the typical axonal spheroids associated with IDPN toxicity while the perikarya of approximately one-third of the neurons contained phosphorylated neurofilaments, which are not found in the perikarya of control rat neurons. By serial or double immunostaining with the SMI-31 and anti-MG 160 antibodies, there were no morphological changes of the GA in the majority of neurons including neurons with a mild to moderate degree of neurofilamentous accumulation. However, a few neurons with a massive accumulation of phosphorylated neurofilaments contained abnormal profiles of the GA which consisted of focal clustering, reduction in size and fragmentation. The results suggest that masses of phosphorylated neurofilaments are associated with structural abnormalities of the GA.

Early decrease of the immunophilin FKBP 52 in the spinal cord of a transgenic model for amyotrophic lateral sclerosis

Expressions of immunophilin FKBP-12 and FKBP-52 were examined in the spinal cord of transgenic mice with an ALS-linked mutant Cu/Zn superoxide dismutase (SOD1) gene. The immunoreactivity of FKBP-12 was present predominantly in the cytoplasm, but did not show a difference between age-matched wild type and transgenic (Tg) mice at 25 and 35 weeks. In contrast, the immunoreactivity of FKBP-52 was predominantly present in the nucleus, which progressively declined only in the Tg mice as early as an early presymptomatic stage at 25 weeks of age in the anterior horn neurons. The present result suggests that the downregulation of FKBP-52 may be involved in the pathogenesis in the early stages of amyotrophic lateral sclerosis (ALS).

Clinical features and neuropathological findings of familial amyotrophic lateral sclerosis with a His46Arg mutation in Cu/Zn superoxide dismutase

We examined the characteristic clinical features of one family of familial amyotrophic lateral sclerosis (FALS) with a His46Arg mutation in the enzyme Cu/Zn superoxide dismutase-1 (SOD1). The disease duration for this family was 18.1±13.2 (mean±S.D.) years, with the age at onset being 39.7±10.5 years old (mean±S.D.). The initial sign was distal weakness of the unilateral lower limb, extending to the lower limb of the other side. A wheel chair became necessary at 9.8±3.2 years after the onset. Upper limb weakness started at 15.5±8.9 years following from the onset. An autopsy was performed on a 71-year-old woman of the family with the mutation. Her disease duration was 47 years, and she died of pneumonia. She had no clear upper motor neuron involvement. Bulbar sign and respiratory muscle weakness had developed 2 years before her death. Neuropathological findings showed degeneration of corticospinal tracts, anterior/posterior spinocerebellar tracts, posterior columns, and Clarke's columns. There were few anterior horn cells in the lumbar spinal cord and no Lewy body-like hyaline inclusion bodies in these remaining anterior horn neurons. This is the first autopsy report of FALS with a His46Arg mutation in the SOD1 enzyme.

Neuronal nitric oxide synthase (nNOS) immunoreactivity in the spinal cord of transgenic mice with G93A mutant SOD1 gene.

Immunohistochemical and quantitative analyses were used to examine the evolution of neuronal nitric oxide synthase (nNOS) with time in spinal motor neurons of transgenic mice with a G93A mutant Cu/Zn superoxide dismutase (SOD1) gene. Specimens from age-matched non-transgenic wild-type mice served as controls. In the controls, the anterior horn including the anterior horn neurons was not immunostained for nNOS. In the transgenic mice, at the age of 24 weeks (early presymptomatic), when no pathological change was observed in the spinal cord, anterior horn neurons were only occasionally immunostained for nNOS (0.3%). At the age of 28 weeks (late presymptomatic), nNOS-positive anterior horn neurons and their neuronal processes were occasionally observed (7.6%), and at the age of 32 weeks (early symptomatic), nNOS-positive anterior horn cells, including degenerated ones showing central chromatolysis, were frequently demonstrated (27.6%) and nNOS-positive cord-like swollen proximal axons were also observed in the anterior horns. nNOS expression in the anterior horn neurons was almost always observed in the somata. At the age of 35 weeks (end stage), neuronal loss of the anterior horn cells was severe, and nNOS-positive anterior horn neurons and cord-like swollen axons in the anterior horns were less prominent compared to those at the age of 32 weeks (33.8%), but many reactive astrocytes were immunostained for nNOS. Thus, nNOS immunoreactivity in the anterior horn neurons is observed as early as the presymptomatic stage and varies with the progression of the disease. The selective localization of positive nNOS immunoreactivity in the anterior horn neurons and degenerated ones in particular, and swollen proximal axons suggests that nNOS immunoreactivity may be involved in the degeneration of anterior horn neurons in this SOD1 transgenic mouse model.

Erratum to: Immunohistochemical and ultrastructural study of basophilic inclusions in adult-onset motor neuron disease

We immunohistochemically and ultrastructurally studied basophilic inclusions (BI) in a patient with adult-onset sporadic motor neuron disease (MND). BI were frequently observed not only in degenerated anterior horn cells, such as central chromatolytic neurons, but also in normal-appearing large anterior horn neurons. They had various shapes, round, elliptical or irregular, and occasionally they had distinct basophilic rims. They also varied in size. There were no halos around them nor core in their centers. Immunohistochemically, some BI were immunostained for ubiquitin or SOD1, but BI were not immunoreactive with anti-phosphorylated neurofilament (SMI 31), phosphorylated tau, cystatin C or Golgi (MG-160) antibodies. Ubiquitin-positive skein-like inclusions (SI) were occasionally observed in the somata of anterior horn neurons. Ultrastructurally, BI consisted of filamentous structures associated with granules, which were attached to thick filaments. The thick filaments were straight without constriction or side arms and their diameter was twice that of the neurofilaments. BI occasionally contained tubular structures among the granule-associated filaments. The granulo-filamentous profiles varied from being compactly arranged to being more loosely packed. The structure of BI resembles that of the Lewy body-like hyaline inclusions (LBHI) observed in sporadic MND patients. Bundles of filaments resembling SI, which were composed of compactly packed filaments without fine granules running parallel to the longitudinal axis, were frequently observed inside or at the periphery of BI, and occasionally clustered in the perikarya. Each filament measured approximately 15-25 nm in diameter, and a bundle of these grouped filaments was sometimes surrounded by a unit membrane. We also occasionally observed in-between structures of BI and bundles of filaments resembling SI. These findings suggest a certain relationship between BI, SI and LBHI in the pathomechanism of BI development. Further studies are needed to elucidate whether sporadic adult-onset MND characterized by BI forms a different subtype of MND.

Inducible nitric oxide synthase (iNOS) and nitrotyrosine immunoreactivity in the spinal cords of transgenic mice with a G93A mutant SOD1 gene.

We performed a prospective, longitudinal immunohistochemical study of the spinal cords of transgenic mice with a G93A mutant SOD1 gene at 4 fixed points in time, using antibodies to inducible nitric oxide synthase (iNOS) and nitrotyrosine. The purpose of this study was to characterize the temporal and topographic distribution of iNOS and nitrotyrosine immunoreactivity in the spinal cord over a certain period, thus illuminating the possible role of increased oxidative damage to the motor system in the neurodegenerative process in this animal model. Specimens from age-matched non-transgenic wild-type mice served as controls. The control mice showed no positive iNOS or nitrotyrosine immuunoreactivity in the somata of anterior horn neurons or their neuronal processes at any age. On the other hand, the transgenic mice demonstrated a common immunostaining pattern of iNOS and nitrotyrosine in the anterior horn neurons. When the mice reached the age of 24 wk (early presymptomatic stage), the anterior horn neurons and their neuronal processes were occasionally immunostained for iNOS and nitrotyrosine; at 28 wk (late presymptomatic stage), the anterior horn neurons were not uncommonly immunostained; at 32 wk (early symptomatic stage) and 35 wk (end-stage), positive iNOS and nitrotyrosine immunoreactivity was frequently observed in proliferated reactive astrocytes as well as in the somata of the anterior horn cells. The selective localization of positive iNOS and nitrotyrosine immunoreactivity in the anterior horn neurons suggests that oxidative stress may be involved in the pathomechanism of degeneration of motor neurons in this transgenic animal model.

Neuronal nitric oxide synthase immunoreactivity in the spinal cord in amyotrophic lateral sclerosis

We investigated the spinal cords of 15 patients with sporadic amyotrophic lateral sclerosis (ALS) immunohistochemically using an anti-human neuronal nitric oxide synthase (nNOS) antibody to examine whether there is increased nNOS immunoreactivity in anterior horn neurons. Specimens from 16 patients without any neurological disease served as controls. In the controls, nNOS immunoreactivity of large anterior horn neurons was detected in 10 out of 16 cases. However, there were few nNOS-positive neurons, and most of large anterior horn neurons were spared. In the ALS patients, the mean number of nNOS-positive anterior horn neurons per transverse section of L4 and L5 was significantly larger (16.2 +/- 10.9) than that in the controls (7.0 +/- 9.2) (P < 0.0001). Moreover, 41.4% of large anterior horn neurons in ALS showed nNOS immunoreactivity in remarkable contrast to 7.6% in the controls. All ALS patients, whether showing mild, moderate or severe depletion of anterior horn neurons, displayed a higher percentage of nNOS-positive anterior horn neurons than the control patients showing nNOS immunoreactivity (P < 0.01). Most of the remaining anterior horn neurons in ALS showed more intense nNOS immunoreactivity on the surface of the neurons and their neuronal processes compared with the controls. Degenerated anterior horn neurons frequently demonstrated more intense nNOS immunoreactivity on the surface of the neurons than normal-appearing neurons. Some anterior horn cells displayed nNOS immunoreactivity in the somata. Dot-like nNOS deposits on anterior horn neurons were also positively immunoreactive with anti-synaptophysin antibody. Thus, increased nNOS expression is located mainly at the synaptic sites on the anterior horn neurons in sporadic ALS, which may be related to the degeneration of anterior horn neurons in this disease. Further studies are needed to determine whether the increased nNOS immunoreactivity plays a neuroprotective or neurotoxic role in the anterior horn neurons, and to show nitric oxide production in ALS.

Therapeutic efficacy of EGb761 (Gingko biloba extract) in a transgenic mouse model of amyotrophic lateral sclerosis.

EGb761 is a standardized extract of green Gingko biloba, which exerts protective effects against mitochondrial damage and oxidative stress. We examined whether oral administration of 0.022% or 0.045% EGb761 in the diet could impart neuroprotective effects in a transgenic mouse model (G93A) of amyotrophic lateral sclerosis (ALS). EGb761 significantly improved motor performance and survival, and protected against a loss of spinal-cord anterior motor horn neurons in male G93A mutant transgenic ALS mice, but not in littermate female mutant transgene mice. While EGb761 extended survival in littermate female G93A mice, significance was not reached. EGb761, however, significantly improved weight loss in both male and female transgenic ALS mice. These findings provide evidence for a gender-specific neuroprotective effect of EGb761 in a transgenic model of ALS and suggest that EGb761 may be a potential effective treatment in patients with ALS.

Hereditary motor and sensory neuropathy with absence of large myelinated fibers due to absence of large neurons in dorsal root ganglia and anterior horns, clinically associated with deafness, mental retardation, and epilepsy (HMSN‐ADM)

Hereditary motor and sensory neuropathy (HMSN) with autosomal recessive inheritance represents a genetically heterogeneous group of disorders with variable clinical, pathologic and electrophysiologic manifestations. A new variant of autosomal recessive HMSN, clinically defined by sensorimotor polyneuropathy associated with deafness and mental retardation, has recently been described. We report on the first autopsy case with this type of HMSN: a girl of non-consanguineous parents with a presumably autosomal recessive type of motor and sensory neuropathy clinically associated with deafness, mental retardation, and epilepsy. The autopsy showed complete absence of large myelinated fibers in peripheral motor and sensory nerves corresponding to a lack of large neurons in dorsal root ganglia and anterior horns of the spinal cord, moderate neurogenic muscle atrophy, and nearly complete absence of neurons in the dentate nucleus of the cerebellum. Molecular genetic analyses in our case revealed neither genetic alterations in the survival motor neuron gene nor in the PMP-22 gene.

INOS and nitrotyrosine immunoreactivity in amyotrophic lateral sclerosis

We carried out an immunohistochemical investigation of the spinal cords of 15 patients with sporadic amyotrophic lateral sclerosis (ALS), using antibodies to inducible nitric oxide synthase (iNOS) and nitrotyrosine; our purpose was to search for a possible role of increased oxidative damage in the motor system that may contribute to the neurodegenerative process in this disease. Specimens from 16 patients without any neurological disease served as controls. In the controls, normal-appearing neurons and their dendrites were negatively immunostained for iNOS. In the ALS patients, most of normal-appearing anterior horn neurons did not show iNOS immunoreactivity either in the perikarya or in their dendrites. However, many of the degenerated neurons showing central chromatolysis or simple atrophy demonstrated focally or diffusely positive iNOS immunoreactivity within the perikarya and their neuronal processes. In the neuropil of the anterior horns, the reactive astrocytes were more intensely immunostained for iNOS as compared with the controls. Some of the swollen proximal axons (spheroids) were focally or diffusely immunostained by the antibody. The corticospinal tracts demonstrated positive iNOS immunoreactivity of proliferated reactive astrocytes. The immunostaining pattern of nitrotyrosine in the anterior horn neurons of the spinal cord was similar to that of iNOS. These findings suggest that selective nitric oxide-mediated oxidative damage in the motor system plays a part in the pathomechanism of the neuronal degeneration in the spinal cord of sporadic ALS.

Excitatory amino acid transporter 1 and 2 immunoreactivity in the spinal cord in amyotrophic lateral sclerosis.

The spinal cord of 20 patients with amyotrophic lateral sclerosis (ALS) and 5 patients with lower motor neuron disease (LMND) were investigated immunohistochemically using anti-human excitatory amino acid transporter 1 (EAAT1) and EAAT2 antibodies which are the astrocytic transporters. The purpose of the study was to examine relationships between EAAT1 and EAAT2 immunoreactivity and degeneration of anterior horn neurons. Specimens from 20 patients without any neurological disease served as controls. In controls, spinal cord gray matter was densely immunostained by antibodies, whereas the white matter was generally not immunostained. In motor neuron disease (MND) patients, EAAT1 immunoreactivity was relatively well preserved in the gray matter despite neuronal loss of anterior horn cells. On the other hand, EAAT2 immunoreactivity in anterior horns correlated with the degree of neuronal loss of anterior horn cells: in the patients with mild neuronal depletion, anterior horns were densely immunostained by the antibody, whereas in the patients with severe neuronal loss, EAAT2 expression was markedly reduced. Degenerated anterior horn cells frequently showed a much denser EAAT1 and EAAT2 immunoreactivity around the surface of the neurons and their neuronal processes than that observed in normal-appearing neurons. There was no difference in the expression of EAAT1 and EAAT2 immunoreactivity between LMND and ALS patients. These findings suggest that in the early stage of degeneration of anterior horn cells, EAAT1 and EAAT2 immunoreactivity is preserved in the astrocytic foot directly attached to normal-appearing neurons, whereas levels of EAAT1 and EAAT2 protein rather increase in the astrocytic foot directly attached to degenerated anterior horn neurons; the latter effect most probably reduces the elevated glutamate level, compensates for the reduced function of astroglial glutamate transporters, or represents a condensation of EAAT1 and EAAT2 immunoreactivity secondary to loss of neurites and greater condensation of astrocytic processes. Thus, we demonstrate a difference in EAAT1 and EAAT2 immunoreactivity in different stages of progression in ALS, as a feature of the pathomechanism of this disease.