Electric fields can have profound effects on biological systems, influencing processes such as differentiation, directed cell and neurite motility, and wound healing. Although it is widely appreciated that the fields can produce small asymmetrically distributed changes in membrane potential, it is not understood how these are amplified and translated into biochemical and biological responses at the level of the cell. This application proposes to test the hypothesis that the field induces a redistributed molecules and Ca2+ channels, clustered this way, produce foci of increased calcium activity which mediate cellular responses. To test these hypotheses, 2 cell types will be used. N1E-115 neuroblastoma cells have calcium channels which are heterogeneously distributed and appear to be involved in neurite outgrowth; they will be used as a model for galvanotropism. The relationship of the dynamics of cell surface molecules and calcium levels to the movement of fish keratocytes has been studied; they also exhibit directed motility in electric fields. Field effects on individual cells will be examined with digital imaging microscopy to study: membrane potential distribution (using potentiometric dyes), calcium channel distribution (using fluorescence and autoradiography with specific antibody or toxin probes), local changes in intracellular calcium levels (using FURA-2), cell and cell-surface particle tracking (using differential interference contrast), and actin cycling (using fluorescent actin and fluorescence recovery after photobleaching techniques).
