Nano opto-mechanical characterization of neuron membrane mechanics under cellular growth and differentiation

Gopal, Ashwini
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Neurons are among the most fundamental building blocks of modern biology and medicine. Axonal development of neurons is intimately dependent on applied mechanical tension. Local 3-D cell growing microenvironment and size-dependant mechanical stimulations can have a profound impact on the reliability of regenerated nerves, but have not been well characterized. The importance of understanding cell membrane interaction with its external mechanical environment has motivated several experiments to measure cell membrane elasticity using micro-scale strain sensors, atomic force microscopy, micropipette aspiration and optical tweezers. However, the methods employed thus far are still limited by measurement range, resolution, and probe-cell interface configuration for minimal damage to cells with small size (less than 10μm), irregular shape and fragile membranes (10-1000Pa). New tools are needed to quantify the neuron membrane properties, especially the elastic modulus, to reveal the cellular responses to mechanical stimulations. Further considerations are also needed on the compatibility of probes with materials and media in which biologically relevant studies such as nerve regeneration may be performed.Recent advancements in microfabricated multi-layer grating photonics enable non-destructive sensing and microscopy of single cells with high resolution. Here we present single-layer pitch-variable diffractive nanogratings on silicon nitride probe to measure PC12 neuron model cells during growth to investigate neuronal cell mechanics and its impact on cellular differentiation and growth. We fabricated single-layer pitch-variable diffractive nanogratings on silicon nitride probe using e-beam lithography and subsequent pattern transfer techniques. We measured the mechanical membrane characteristics of PC12 cells using the force sensors with displacement range of 10 μm and force sensitivity 8 μN/μm. Young's moduli of 425±30 Pa were measured for PC 12 cells cultured on PDMS substrate coated with collagen . We have also observed stimulation of directed neurite contraction on extended probing up to 6 μm on extended probing for a time period of 30 minutes.