De novo protein synthesis in isolated axons of identified neurons

Neurons are highly polarized cells that contain a wealth of cytoplasmic and membrane proteins required for neurotransmission, synapse formation and various forms of neuronal plasticity. Typically, these proteins are differentially distributed over somatic, dendritic and axonal compartments. Until recently, it was believed that all proteins destined for various neuronal sites were synthesized exclusively in the somata and were subsequently targeted to appropriate extrasomal compartments. The discovery of various messenger RNA molecules in both dendrites and axons is suggestive of de novo protein synthesis in extrasomatic regions. The latter process has been demonstrated in few neuronal svrstems, but direct proof for the axonal transcription of a specific protein from a given messenger RNA is still lacking. This lack of fundamental knowledge in the field of cellular and molecular neurobiology is due primarily to both anatomical and experimental difficulties encountered in most animal preparations studied thus far. In this study we developed a neuronal experimental system comprising of individually identified neurons and their isolated axons from the mollusc Lymnaea stagnalis. We injected a foreign messenger RNA encoding a peptide precursor into the isolated axons of cultured neurons; and utilizing cellular, molecular and immunocytochemical techniques, we provide direct evidence for specific protein synthesis in isolated axons. The Lymnaea model provides us with an opportunity to examine the role and specificity of de novo protein synthesis in the extrasomal regions.