Role of Nav1.9 in activity-dependent axon growth in motoneurons

Spontaneous neural activity promotes axon growth in many types of developing neurons, including motoneurons. In motoneurons from a mouse model of spinal muscular atrophy (SMA), defects in axonal growth and presynaptic function correlate with a reduced frequency of spontaneous Ca²⁺ transients in axons which are mediated by N-type Ca²⁺ channels. To characterize the mechanisms that initiate spontaneous Ca²⁺ transients, we investigated the role of voltage-gated sodium channels (VGSCs). We found that low concentrations of the VGSC inhibitors tetrodotoxin (TTX) and saxitoxin (STX) reduce the rate of axon growth in cultured embryonic mouse motoneurons without affecting their survival. STX was 5- to 10-fold more potent than TTX and Ca²⁺ imaging confirmed that low concentrations of STX strongly reduce the frequency of spontaneous Ca²⁺ transients in somatic and axonal regions. These findings suggest that the Na_V1.9, a VGSC that opens at low thresholds, could act upstream of spontaneous Ca²⁺ transients. qPCR from cultured and laser-microdissected spinal cord motoneurons revealed abundant expression of Na_V1.9. Na_V1.9 protein is preferentially localized in axons and growth cones. Suppression of Na_V1.9 expression reduced axon elongation. Motoneurons from Na_V1.9^–/– mice showed the reduced axon growth in combination with reduced spontaneous Ca²⁺ transients in the soma and axon terminals. Thus, Na_V1.9 function appears to be essential for activity-dependent axon growth, acting upstream of spontaneous Ca²⁺ elevation through voltage-gated calcium channels (VGCCs). Na_V1.9 activation could therefore serve as a target for modulating axonal regeneration in motoneuron diseases such as SMA in which presynaptic activity of VGCCs is reduced.