Antisense oligodeoxynucleotide (ODN) against neuronal NOS down-regulated the de novo expression of NOS and decreased the survival of adult spinal motoneurons following root avulsion

Abstract

We have previously shown that nNOS is de novo expressed in adult spinal motoneurons after root avulsion. Two hypotheses have been proposed to interpret such expression. One is that injury-induced nNOS is neurotoxic and directly responsible for the death of injured neurons. Alternative hypothesis is that induction of nNOS represents an attempt of injured motoneurons to resist an unknown substance that causes the death of motoneurons. In the present study antisense ODN against nNOS was applied in root avulsion model and its effect on nNOS expression and motoneuron survival were observed. Two weeks after C7 root avulsion in adult rats, antisense nNOS ODN was applied intrathecally (4 nmol/each time, repeat in every 3 days). Randomized ODNs set was used as sham control. The nNOS immunocytochemistry, NADPH-d histochemistry and neutral red counter stain were used to assess the effect of treatments. Our result showed that treatments with randomized ODNs set did not alter the expression of nNOS or the survival rate of motoneurons as compared with the control (avulsion only). In contrast, treatment with antisense nNOS ODN significantly down-regulated the expression of nNOS and the number of nNOS positive motoneurons significantly decreased as compared with the control (19.22% vs 55.92% by 3 weeks and 11.48% vs 32.27% by 4 weeks post-injury respectively). At the same time, the treatment significantly reduced the number of survival motoneurons as compared with the control (57.45% vs 76.78% by 3 weeks and 35.44% vs 61.59% by 4 weeks post-injury respectively). Results of the present study support the hypothesis that injury-induced nNOS in injured spinal motoneurons plays a beneficial role on the survival of spinal motoneurons. Supported by The Hong Kong Research Grants Council