| Amyotrophic lateral sclerosis (ALS) is a progressive neurodegenerative disease, selectively involving the upper and lower motor neurons that control voluntary movement. The disease is characterized by progressive muscle weakness with death, usually resulting from respiratory failure, occurring 3-5 years from the first appearance of symptoms. The disease can be either sporadic (SALS; more than 90%) or familial (FALS; less than 10%) in nature, with an average age onset of 59 years. Its prevalence is about five to seven in 100,000 individuals, making it the most frequent paralytic disease in adults, with an incidence of 1-3 per 100 000. Pathologically, ALS is characterized by a loss of upper motor neurons in the cerebral cortex and of the lower motor neurons in the spinal cord and brainstem. Often, there is also a profound degeneration of the pyramidal tracts, which is most evident at the level of the spinal cord. The few remaining motor neurons are generally atrophic, and many demonstrate abnormal accumulation of neurofilament, in both their cell bodies and axons. To date, only a few approved treatments (e.g., mechanical ventilation and riluzole) prolong survival in ALS patients to some extent. However, the development of more effective neuroprotective therapies remains impeded by our limited knowledge of the actual mechanisms by which neurons die in ALS, and of how the disease progresses and propagates. Establishing experimental models will be useful to investigate the mechanism of ALS and to explore new neuroprotective treatment.Etiological study have found that a total of 10 to 20% of FALS cases, which make up 5 to 10% of all ALS cases, are associated with a mutation in the copper/zinc superoxide dismutase (Cu/Zn SOD) gene. This suggests that FALS should be a kind of genetic disease. The etiology of SALS, which accounts for the majority of all ALS cases, remains to be resolved, and several pathogenic mechanisms have been suggested, including excitotoxicity, autoimmunity, oxidative stress, neuron apoptosis, mitochondria dysfunction, et al. Among these, a considerable amount of evidence lends support to the excitotoxic hypothesis.Glutamate is the principal excitatory neurotransmitter in the central nervous system. Over activity of the glutamatergic neurons and excessive extracellular accumulation of Glu surrounding the synaptic cleft may cause exicitotoxic damage in the postsynaptic neurons and other surrounding tissues through over activation of glutamate receptors. Glutamate reuptake is mediated by glial and neuronal transporters. Removal of glutamate through the action of glial and neuronal glutamate transporters, especially the high affinity glutamate transport in astrocyte, is important in preventing excitotoxicity in vivo, which is also the main mechanism of avoiding excitotoxicity. Inhibition significantly the function of glutamate transporters will result in elevation of excellular glutamate and if sustained might produce extracellular glutamate accumulation and exctotoxicity.Organotypic spinal cord culture can be reliably maintained for long time and have well preserved organotypic morphology, which provides a more fitful way to study spinal cord diseases than dissociated cell culture. We used organotypic spinal cord cultures to develop an in vitro model of ALS by chronically blocking the glutamate carriers, further to study the mechanism of selective motor neurons injury. Our study includes three parts. In the first part we established the spinal cord organotypic culture method and compared the cultures with age-matched spinal cord in vivo. In the second part, based on the hypothesis that glutamate can produce excitotoxicity, we used proper concentration of threohydroxyaspartate (THA) to inhibit glutamate transport that results in elevation of extracellular glutamate and motor neurons loss with sensory neurons not affected. Thus the in vitro culture model of ALS can be established. In the third part, we used spinal cord cultures to observe if the serum from a patient with ALS affected survival...
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