Immunocytochemical study of the distribution of the free radical scavenging enzymes Cu/Zn superoxide dismutase (SOD1); MN superoxide dismutase (MN SOD) and catalase in the normal human spinal cord and in motor neuron disease.

Interest in free radical-mediated mechanisms of motor neuron injury has arisen following the discovery of point mutations in the Cu/Zn superoxide dismutase (SOD1) gene in a proportion of cases of familial motor neuron disease (MND). Evidence is emerging which indicates that oxidative stress may contribute to the pathophysiology of sporadic MND. Free radical scavenging enzymes form a major component of the anti-oxidant defense system. The aim of this study was to compare the distribution and density of immunoreactivity to Cu/Zn SOD. Mn SOD and catalase in the spinal cord of sporadic MND cases (n = 10) compared to normal controls (n = 8). There was abundant expression of Cu/Zn SOD, Mn SOD and catalase in spinal motor neurons, suggesting important roles for these enzymes in neuroprotective pathways in this cell group. In MND cases, there was no evidence in surviving motor neurons of a consistent alteration in the protein expression of any of these enzymes. There was evidence of increased expression of these enzymes in glial cells present in the ventral and intermediate grey matter and degenerating descending motor pathways of the spinal cord in MND cases. The changes observed were more marked in the cervical compared to lumbar spinal segments. Further investigation is required to determine whether these findings represent a compensatory response to a pathophysiological process involving oxidative stress.

Quantitative study of synaptophysin immunoreactivity of cerebral cortex and spinal cord in motor neuron disease.

Synaptophysin immunoreactivity can be quantified by image analysis to evaluate loss of presynaptic terminals in human neurodegenerative diseases. The extent and regional distribution of such loss is reported in motor neuron disease (MND). Autopsy samples of spinal cord and cerebral cortex were examined from 28 cases of MND and 28 age and sex matched controls. The MND group included individuals with amyotrophic lateral sclerosis (17[ALS]), and progressive muscular atrophy (11[PMA]). In the spinal cord, there was significant reduction of presynaptic terminals in the lateral ventral horn (15%) in both the ALS (p < 0.01) and PMA (p < 0.05) groups. Perisomatic synaptophysin profiles on lower motor neurons are preserved late in the disease and are not related to corticospinal innervation. Less marked presynaptic loss was demonstrable more widely in the medial ventral, intermediate and dorsal spinal grey matter (10%) in both ALS (p = 0.03) and PMA (p = 0.05). In the cerebral cortex no synaptic loss was demonstrated in motor or anterior cingulate regions in any of the MND cases. Spinal degeneration in MND is associated with loss of presynaptic terminals in all grey matter regions. It is most marked in the limb motor neuron area and is independent of corticospinal tract degeneration. The cerebral pathology of ALS is not associated with significant loss of presynaptic terminals in the cortical areas studied.

Non-NMDA receptors in motor neuron disease (MND): a quantitative autoradiographic study in spinal cord and motor cortex using [3H]CNQX and [3H]kainate.

The distribution and density of non-NMDA receptors in spinal cord and motor cortex was compared in 10 cases of motor neuron disease (MND) and 8 neurologically normal controls by quantitative autoradiography using [3H]CNQX and [3H]kainate. In the motor cortex of MND cases, an increased density of [3H]kainate binding sites was observed which was most marked in the deep layers. No significant differences were observed in [3H]CNQX binding in the motor cortex between MND and control cases. In the spinal cord significantly increased densities of both [3H]CNQX and [3]kainate binding sites were found in the substantia gelatinosa and the intermediate grey matter in the MND group. The changes in [3H]kainate binding were observed only in the amyotrophic lateral sclerosis (ALS) subgroup of MND, while the changes in [3H]CNQX binding in the spinal cord were more marked in ALS compared to progressive muscular atrophy (PMA) cases. These findings provide evidence in support of a disturbance of glutamatergic neurotransmission in MND and suggest that there may be an increased excitatory drive to motor neurons via non-NMDA receptors. It is unclear at present whether the changes observed represent a compensatory response to loss of motor neurons in MND or a pathophysiological phenomenon contributing to motor neuron degeneration. Modulation of non-NMDA receptor activity may represent a possible target for therapeutic intervention in this disease.

[3H]D-aspartate binding sites in the normal human spinal cord and changes in motor neuron disease: a quantitative autoradiographic study.

The distribution and density of glutamate transporter sites was determined in human cervical and lumbar spinal cord, by quantitative autoradiography using [3H]D-aspartate. In the normal human spinal cord (n = 8) there was specific binding of [3H]D-aspartate throughout the spinal grey matter, with the highest levels observed in the substantia gelatinosa and central grey matter. In the ventral horns, particularly at the L5 level, focal hot spots of binding were observed in a distribution corresponding to that of lower motor neuron somata. Comparison of motor neuron disease (MND) cases (n = 12) with normal controls showed a reduction in the density of [3H]D-aspartate binding in the intermediate grey matter and the substantia gelatinosa of the lumbar cord. These changes were more marked in the amyotrophic lateral sclerosis (ALS) compared to the progressive muscular atrophy (PMA) subgroup, and may be due to loss of glutamatergic terminals of the corticospinal tract. The changes observed in the cervical cord were milder and did not reach statistical significance. No differences were found between [3H]D-aspartate binding in the spinal cords of the normal controls and a neurological disease control group (n = 6), suggesting that the changes observed in MND are disease specific. These findings provide further evidence in support of a disturbance of glutamatergic neurotransmission in MND.

Autoradiographic distribution of binding sites for the non-NMDA receptor antagonist [3H]CNQX in human motor cortex, brainstem and spinal cord.

The distribution of non-NMDA receptors in the normal human motor cortex, brainstem and spinal cord has been investigated using [3H]CNQX. In the motor and premotor cortex, specific [3H]CNQX binding was present in all cortical laminae with the highest density of binding sites in laminae I, II and the upper part of III. In the normal brainstem, non-NMDA receptors labelled by [3H]CNQX had a heterogeneous distribution. Brainstem motor nuclei subserving eye movements, which tend to be spared in motor neuron disease (MND), had a higher density of [3H]CNQX binding sites compared to other cranial nerve motor nuclei (VII, X, XII) which tend to be affected. Specific [3H]CNQX binding was present throughout the spinal grey matter, the greatest density of binding being found in the substantia gelatinosa. Excitotoxicity at non-NMDA receptors has been implicated in chronic neurodegenerative diseases such as motor neuron disease. This study suggests that the density of non-NMDA receptors, labelled by [3H]CNQX, does not account for selective vulnerability of motor neurons in this disorder.