Antisense oligos to neuronal nitric oxide synthase aggravate motoneuron death induced by spinal root avulsion in adult rat

Abstract

The present study used nitric oxide synthase (nNOS) antisense oligos (nNOS AS-ODN) to assess the role of nNOS in motoneuron death induced by spinal root avulsion. A right seventh cervical (C7) spinal root avulsion was performed on adult male Sprague–Dawley rats. Two weeks later, FITC-labeled random oligos (FITC-R-ODN), nNOS AS-ODN, R-ODN or TE buffer was applied to the lesioned side of the C7 spinal segment and refreshed every 3 days. FITC-R-ODN was first detected inside the injured motoneurons at 10 h, accumulated to a maximum by 24 h and faded out from 72 h. Following avulsion, nNOS AS-ODN decreased the number of nNOS-positive motoneurons in the lesioned segment compared either with buffer ( P < 0.001 at 15 days, 3 and 4 weeks post-injury) or with R-ODN control ( P = 0.002 at 15 days, P < 0.001 at 3 and 4 weeks post-injury). Interestingly, nNOS AS-ODN also decreased the number of surviving motoneurons compared either with buffer ( P = 0.005 at 15 days, P < 0.001 at 3 or 4 weeks) or with R-ODN control ( P < 0.001 at 3 or 4 weeks). Meanwhile, there were no significant differences between R-ODN and buffer control either in the number of nNOS-positive motoneurons ( P = 0.245 at 15 days, P = 0.089 at 3 weeks and P = 0.162 at 4 weeks) or in the number of surviving motoneurons ( P = 0.426 at 15 days, P = 0.321 at 3 weeks or P = 0.344 at 4 weeks). These findings indicate that nNOS AS-ODN, applied from 2 weeks after avulsion, aggravates the motoneuron death due to root avulsion by specifically down-regulating nNOS gene expression and that the expression of nNOS in adult spinal motoneurons in response to root avulsion may play a beneficial role in the survival of injured neurons.