Hyperosmotic stress results in the abolishment of endo- and exocytosis partially due to cytosolic acidification (A) Cells which were resuspended in SPB buffer avidly endocytose the fluorescent fluid-phase marker FITC-Dextran, whereas no fluid uptake was observed with cells exposed to 400 mM sorbitol in SPB buffer. (B) Correspondingly, cells in SPB buffer rapidly exocytosed ingested FITC-Dextran, whereas no loss of fluorescence was measured with cell suspended in 400 mM sorbitol, indicating that exocytosis is blocked. (C) Cells, which were artificially acidified by suspending the cells in SPB buffer/5 mM propionic acid (pH 6.0)/FITC-Dextran did not accumulate any FITC-Dextran, indicating that endocytosis was blocked. Also suspending the cells in SPB buffer/30 μM DES/FITC-Dextran resulted in the inhibition of endocytosis. However, a resumption of endocytic activity was observed after 1 h of hyperosmotic shock, presumably due to adaptation. As control, cells were suspended in SPB buffer/FITC-Dextran. (D) Artificial acidification of cells partially inhibits exocytosis. Control cells labeled with FITC-Dextran exocytosed about 90% of the dye within 2 h, whereas a loss of 55%-60% of cell-associated fluorescence was measured with cells exposed to 5 mM propionic acid or 30 μM DES.