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Spatiotemporal Characteristics of Intercellular Calcium Wave Communication in Micropatterned Assemblies of Single Cells.

Micropatterned substrates offer a unique possibility to define and control spatial organization of biological cells at the microscale, which greatly facilitates investigations of the cell-to-cell communication in vitro. Here, we developed a simple micropatterning strategy to resolve various spatiotemporal characteristics of intercellular calcium wave (ICW) communication among isolated BV-2 microglial cells. By using a single-ring assembly, we found that the direction of the initial transmitter secretion was strongly correlated with the site of the cell at which the mechanical stimulus triggering the ICWs was imposed. By using multiring assemblies, we observed that the response ratio of the same outmost cells 160 μm away from the center increased from 0% in the single-ring assembly to 9.6% in the four-ring assembly. This revealed that cells located in the interring acted as regenerative amplifiers for the ICWs generated by the central cell. By using a special oval-type micropattern, we found that calcium mobilization in lamellipodia of a fusiform BV-2 microglia cell occurred 2.9 times faster than that in the middle part of the cell, demonstrating a higher region-specific sensitivity of lamellipodia to the transmitter. Taken together, our micropatterning strategy opened up new experimental prospects to study ICWs and revealed novel spatiotemporal characteristics of ICW communication including stimulation site-dependent secretion, regenerative propagation, and region-specific cell sensitivity.