Glutamatergic receptors regulate expression, phosphorylation and accumulation of neurofilaments in spinal cord...

Glutamatergic regulation of neurofilament expression, phosphorylation and accumulation in cultured spinal cord neurons was studied. At seven days in culture, 0.15% of the neurons were immunoreactive for non-phosphorylated neurofilaments, but essentially no cells immunoreactive for phosphorylated neurofilaments were seen. The number and size of the immunoreactive cells in culture corresponded well to those of rat and human spinal cord neurons in vivo. In spinal cord cultures, sublethal, long-lasting stimulation of alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionate (AMPA)/kainate or metabotrophic receptors, but not N-methyl-D-aspartate receptors, dose-dependently increased the number of non-phosphorylated neurofilament-immunoreactive cells, which was blocked by nifedipine, an antagonist of voltage-sensitive Ca2+ channels. Stimulation of kainate or all non-N-methyl-D-aspartate receptors decreased the expression of medium-molecular-weight neurofilament messenger RNA. Blockade of AMPA/kainate receptors, but not of N-methyl-D-aspartate receptors, increased the amount of phosphorylated neurofilament protein and the number of phosphorylated neurofilament-immunoreactive cell bodies. The phosphorylated neurofilament-immunoreactive cell population was different from the non-phosphorylated neurofilament-immunoreactive neurons, which lost their axonal non-phosphorylated neurofilament immunoreactivity but showed intense cytoplasmic labeling in response to the blockade of AMPA/ kainate receptors. Immunoreactivity for phosphoserine did not change upon glutamate receptor stimulation and blockade. The results show that activation of AMPA/kainate receptors decreases the expression of neurofilament messenger RNA and neurofilament phosphorylation in spinal cord neurons by a mechanism involving active voltage-sensitive Ca2+ channels. Blockade of these receptors seems to disturb axonal neurofilament transport. Because AMPA/kainate receptors mediate chronic glutamatergic death of spinal motor neurons and these receptors have been suggested to be involved in the pathogenesis of amyotrophic lateral sclerosis, the observed alteration in neurofilament phosphorylation and distribution may contribute to the pathogenesis of chronic motor neuron diseases.