Visualization of intracellular calcium transport between cells using high frequency ultrasound and FRET live-cell imaging

摘要

The live visualization with fast kinetics of the interaction between cells has been long term challenges because of the lack of efficient stimulation methods. We propose an approach to achieve single cell resolution stimulation and FRET-base calcium live cell imaging to visualize fast kinetics of calcium transport between physically connect neighboring cells. Chemical stimulation stimulates cells within a dish at the same time and is not suitable for the study of cell-cell interaction. We replaced chemical stimulation with ultrasound-based mechanical stimulation approach to provide precise spatiotemporal resolution. To achieve this, we integrated 3D translation stages and epi-fluorescence microscope and a developed 150 MHz high frequency ultrasound with fnumber of 1 and aperture size of 1 mm. The 150 MHz transducer can focus within 10 micrometers in diameter and directly stimulate cells by disturbing cell plasma membranes without microbubbles. High frequency stimulation was used to introduce calcium ions into cytoplasm of cells. Results demonstrate calcium transport between cells, visualized by FRET calcium biosensor after only one cell was stimulated by the developed high frequency ultrasonic transducer.