Osmotic shock, a 15-minute plasmolysis followed by a 15-minute rehydration in the cold, is a nondestructive technique which inhibits fusicoccin-stimulated H(+) excretion from oat mesophyll cells (Avena sativa L.). Osmotic shock also causes a loss of intracellular solutes and stimulates H(+) uptake, but osmoregulation can still occur, and enhanced H(+) uptake is observed only at low external pH. It is concluded that osmotic shock interferes directly with the excretion of H(+) rather than affecting only H(+) or counter ion uptake.Plasmolysis alone does not inhibit fusicoccin-enhanced H(+) excretion, and the rehydration step must be rapid and in the cold for maximum inhibition. This suggests that the plasma membrane is perturbed, possibly due to release or rearrangement of membrane protein. Compared to corresponding osmolarities of sorbitol or NaCl, polyethylene glycol 4000 is much less effective during the plasmolysis step; the possibility is discussed that the ineffectiveness of polyethylene glycol 4000 is due to its preservation of plasmodesmata.