High Resolution Mapping of Cytoskeletal Dynamics in Neurons via Combined Atomic Force Microscopy and Fluorescence Microscopy

摘要

Living neuronal cells present active mechanical structures which evolve with cellular growth and changes in the cell microenvironment. Detailed knowledge of various mechanical parameters such as cell stiffness or adhesion forces and traction stresses generated during axonal extension is essential for understanding the mechanisms that control neuronal growth, development and repair. Here we present a combined Atomic Force Microscopy (AFM)/Fluorescence Microscopy approach for obtaining systematic, high-resolution elasticity and fluorescent maps for live neuronal cells. This approach allows us to simultaneously image and apply controllable forces to neurons, and also to monitor the real time dynamics of the cell cytoskeleton. We measure how the stiffness of neurons changes both during axonal growth and upon chemical modification of the cell, and identify the cytoskeletal components most responsible for the changes in cellular elasticity. This is accomplished by identifying cellular components with unique elastic signatures, and tracking those components over time within healthy cells or within cells treated to disrupt selective components.