4D confocal microscopy of time dependent cell and nuclear osmotic response reveals differential osmoregulation and transport kinetics

We investigated the equilibrium and dynamic cell and nuclear volumetric responses in adherent human hepatoblastoma (HepG2) cell monolayers using live three-dimensional imaging. We have further demonstrated that both cells and nuclei regulate their volume in response to osmotic stress. For instance, during hyposmotic stress, cells swell and lose important osmolytes through active volume regulation or bleb formation, however, nuclei undergo more expansion than cells. Whereas during hyperosmotic stress, both cells and nuclei shrink rapidly into irregular shapes and undergo passive volume regulation. Furthermore, we explored the mechanism of osmoregulation in these cells by restoring isosmotic conditions after osmotic stress and found a higher return-to-equilibrium volume for nuclei compared to their initial isosmotic volume, while cytosol returned to a lower volume than its isosmotic volume after swelling. Moreover, water and solute permeability estimates for both cells and nuclei were also inferred from exposure to different anisosmotic media by fitting coupled transport models. This accurate estimation of cell and nuclear volumetric responses and water and solute permeability parameters could have important implications for understanding cell physiology and the mechanism of osmoregulation.