EFFECTS OF CEREBRAL BIOMECHANICS ON THE DYNAMICS OF MEMBRANE TRAFFIC IN NEURONAL DENDRITES

Postsynaptic receptor trafficking is essential for synaptic transmission and plasticity. Receptor internalization occurs at endocytic zones (EZs) in spines near postsynaptic densities (PSDs) or along dendritic shafts. EZ localization near PSDs depends on Shank proteins. EZs, marked by clathrin, appear very static in mature neurons in culture. However, we imaged EZs in neurons in organotypic slices, and they are much more dynamic. This could be due to differences in mechanical strain experienced by neurons. Indeed, the brain is among the softest biological tissues and it was shown in non-neuronal cells that EZ dynamics is also strongly affected by substrate rigidity.
To study these effects, we developed a method to culture neurons on gels of controlled rigidity. We characterized the morphology of neurons at 17 days in vitro using transfection and immunofluorescence. Neurons cultured on softer gels have similar dendritic length and branching, but a higher density of bigger dendritic spines, as well as longer axon initial segment. Moreover, neurons on softer gels have a higher fraction of PSDs with an EZ in spines. Endocytosis is lower in neurons cultured on soft gels. However, we predict that endocytosis and recycling of post-synaptic receptors, concentrated at PSDs, could be higher in neurons growing on softer substrate. Therefore, we plan to assess the dynamics of EZs with live cell imaging, the number of post-synaptic receptors with immunofluorescence and electrophysiology (patch-clamp) and finally, synaptic plasticity, in particular long-term depression, in neurons cultured on soft and rigid substrates.