Cytoplasmic Hydration Triggers a Transient Increase in Cytoplasmic Ca2+ Concentration in Nitella flexilis

Cells of Nitella flexilis were made inexcitable by treatmentwith 10 mM KCl for more than 24 h. A Ca²⁺-sensitive photoproteinaequorin was injected into the cytoplasm of such cells. Forvacuolar per fusion, the central part of an aequorin-loadedcell was immersed in silicone oil, and both cell ends bathedin the perfusion medium were cut off. A large light emissionfrom aequorin was observed when the vacuole was perfused witha hypotonic medium whose osmotic pressure was adjusted to halfof the osmotic pressure of the cell sap. This shows that hydrationof the cytoplasm triggers release of Ca²⁺ from internal stores,since influx of Ca²⁺ from silicone oil is excluded. Hydration of cells was induced in another way. Cells were firstdehydrated by transferring them from 10 mM KCl solution to thatwith 250 mM sorbitol added. This procedure did not affect thecytoplasmic streaming. When cells were rehydrated by transferringthem to 10 mM KCl solution, cytoplasmic streaming was eitherstopped or slowed down in a few seconds. A quick light emissionfrom aequorin was observed in the rehydration, evidence thatcytoplasmic streaming was inhibited by an increase in the cytoplasmicCa²⁺ concentration. Both streaming cessation and aequorin lightemission were observed even in KCl-treated cells which werefurther treated with 5 mM EGTA. Thus, the increase in Ca²⁺ isconcluded to be caused by the release of Ca²⁺ from internalstores. These results support our previous hypothesis Tazawa et al.(1994) Plant Cell Physiol. 35:63] that, in Nitella flexilis,the increase in the concentration of Ca²⁺ in the cytoplasm whichoccurs on the endoosmotic side of the cell during transcellularosmosis is caused by hydration of the cytoplasm. (Received June 6, 1994; Accepted December 26, 1994)