High Frequency Chemical Stimulation of Living Cells

Abstract

Fundamental and applied biomedical research increasingly relies on the quantitative analysis of the response of individual cells to controlled stimulations. The advent of microfluidics has revolutionized the methods to tailor the chemical environment of live cells, allowing stimulations to be applied with high spatiotemporal control and using minute amounts of reagents. It has been shown that Dictyostelium Discoideum cells (D.D.) can sense cAMP gradients on scales smaller than their size (a few tens of microns) and can adapt to very rapid concentration changes on sub second timescales. Classical methods to apply chemical stimulations, which are often based on the diffusion of solutes released by micropipettes, suffer from a poor spatial and temporal resolution and cannot discriminate the different timescales involved in the gradient sensing mechanism of D.D. Here we take advantage of a recently developed microfluidic toolbox based on microfluidic stickers made out of stiff polymers to stimulate adherent D.D. cells within microchannels. Microfluidic stickers allow for periodic chemical stimulation at the subcellular scale at frequencies up to 10Hz.We will present the dependence of the gradient sensing response as a function of both the temporal and spatial frequency. We will show how the microfluidic framework used in this study overcomes several experimental limitations and thus considerably extends previous investigations on the chemotaxis of D.D. by giving a new insight on previously proposed gradient sensing models.