Analysis of the mechanisms of cell motility using living cells and cell-free systems.

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1. Experimental systems for keeping organelle activities outside the cell. Cytoplasm at the clear zone of centrifuged xenopus eggs was isolated in paraffin oil and used it as a medium for keeping activities of organelles isolated from living cells. Cytoplasm of lens cells of fish eggs was tried to use for the same purpose.2. Micromanipulation of the mitotic apparatus and its mechanical properties. Effects of deforming and severing the mitotic apparatus on the chromosome movement in sea urchin eggs were analyzed by micromanipulation. The results suggested that the motive force for anaphase chromosome movement is located at or near the moving chromosomes. The measuement of mechanical properties of the mitotic apparatus revealed that the equatorial region of the spindle is more fragile than the remaining regicons of the spindle.3. Quantitative measurement of the spindle birefringence. A novel method was developed to measure total birefringence within a microscopic area having heterogeneous birefringence, and it was applied to measure the birefringence of the spindles in normal and Colcemid-treated sea urchin eggs.4. Measurement of mechanical properties of the cytoplasm. Movements of gold particles, oil droplets or the nucleus in the cytoplasm by centrifugal force were observed in sea urchin and starfish eggs with a centrifuge microscope. It was found that the distribution and nature of the cytoskeleton in the cytoplasm. are closely related to the movement of the particles etc. Causal relations betweenthe position of the cleavage furrow and that of the mitotic apparatus were analyzed by moving the mitotic apparatus in the cell by centrifugation at various stages of mitosis.5. Roles of calcium ions during cellular activity. Spatial and temporal changes in intracellular calcium ion concentration during fertilization in fish eggs, protoplasmic streaming of slime mold and movement of amoebae were determined by measuring the luminescence of aequorin-loaded cells.