Fundamental Characteristics of Neuron Adhesion Revealed by Forced Peeling and Time-Dependent Healing

A current bottleneck in the advance of neurophysics is the lack of reliable methods to quantitatively measure the interactions between neural cells and their microenvironment. Here, we present an experimental technique to probe the fundamental characteristics of neuron adhesion through repeated peeling of well-developed neurite branches on a substrate with an atomic force microscopy cantilever. At the same time, a total internal reflection fluorescence microscope is also used to monitor the activities of neural cell adhesion molecules (NCAMs) during detaching. It was found that NCAMs aggregate into clusters at the neurite-substrate interface, resulting in strong local attachment with an adhesion energy of ∼0.1 mJ/m² and sudden force jumps in the recorded force-displacement curve. Furthermore, by introducing a healing period between two forced peelings, we showed that stable neurite-substrate attachment can be re-established in 2-5 min. These findings are rationalized by a stochastic model, accounting for the breakage and rebinding of NCAM-based molecular bonds along the interface, and provide new insights into the mechanics of neuron adhesion as well as many related biological processes including axon outgrowth and nerve regeneration.