Li Chunyan ,Wang Xiaojuan , Song Xueqin , Wang Liqin

Abstract objective: To investigate the effects of serum from a patient with amyotrophic lateral sclerosis (ALS) on organotypic spinal cord cultures and to study some mechanisms of selective motor neuron damage in ALS.

Methods: 240 organotypic spinal cord cultures were prepared using lumbar spinal cord slices from 120 82 day2 old rats1 Serum from healthy individual or ALS patient with high level of glutamate (Glu) was continuously added into the culture medium for 4 weeks. 1 Ventral a 2 motor neurons survival was evaluated by monoclonal antibody S M I2 32, a nonphosphorylated neurofilament marker, immune 2 histochemistry staining, and interneurons in the dorsal horn were identified bymonoclonal anticalretinin staining compared with controls Mitochondrial function was observed by succinate dehydrogenase (SDH) enzyme chemical staining Lactate dehydrogenase (LDH), (Glu), super oxide dismutase (SOD ) and malondialdehyde (MDA) levels in culture medium were also measured.

Results: The spinal cord explants in the control group could maintain an excellent organotypic cellular organization and a stable population of ventral motor neurons (about 15a2 mot or per explant). However, the treatment for 4 weeks with the patient' s serum produced a significant loss of motor neurons (about 7 per explant) as compared with controls. The number of interneurons in the dorsal horn showed no significant difference and mitochondrial SDH activity in the ventral horn was also inhibited after being cultured for 3 weeks. LDH and MDA were increased by 22.4% and 48.2% in culture medium of the patient serum treated group, while SOD activity was slightly decreased by 12.6%. Glutamate levels in culture medium of the ALS patients treated group were continuously higher than the controls (P <0101).

Conclusion: Excitatory amino acid toxicity and free radicals injury might play an important role in pathological changes of ALS Organotypic spinal cord culture and should provide an effective way to study the mechanisms of motor neuron selective damage in ALS.

Keywords: Amyotrophic lateral sclerosis; Spinal cord; Glutamic acid; Free radicals

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