Microtubule dynamics in adult retinal ganglion cells and dorsal root ganglion neurons

Introduction: While axon regeneration is very limited in the adult central nervous system (CNS) in vivo, this is not the case in the peripheral nervous system (PNS). Indeed, both CNS and PNS neurons can regenerate in vitro although to varying degrees. Given the role of microtubule stabilization in promoting regeneration, we have examined microtubule polymerization during the regeneration of two types of adult neurons in vitro, retinal ganglion cells (RGCs) from the CNS and dorsal root ganglion (DRG) neurons from the PNS. Methods: In order to compare microtubule dynamics between these cell types, the density, polymerization rate and orientation of microtubules have been analysed during neurite regeneration in both cell types by analysing GFP-tagged Microtubule End Binding 3 (EB3) protein transfected into the neurons. Results: The density of EB3 comets and the speed of EB3 movement was similar in both cell types, although only one subtype of RGC regenerated sufficiently long neurites for analysis. In the absence of extracellular substances that could inhibit neurite regeneration, the dynamics of the microtubules of the RGC subtype that extend long neurites are very similar to those in DRG neurons. However, some RGCs with very short neurites exhibited EB3 comets that progressed retrogradely. Additionally, live imaging of mitochondria was performed in both neuronal cultures. Discussion: Regenerating neurites assessed in our study exhibited similar microtubule extension dynamics in both CNS- and PNS-originated neurons. Importantly, the observation that robust neurite outgrowth is restricted to RGC subtypes highlights the need to integrate molecular heterogeinity among RGCs in future studies.