Fluorescent probes were used to visualize the morphology of membranes and of F‐actin in rat peritoneal mast cells, exposed to hyperosmotic medium and consequently reversed to isotonicity. Hypertonicity induced cell shrinkage followed by a regulatory volume increase, and cell alkalinization that was sensitive to amiloride, an inhibitor of the Na+/H+ exchanger (NHE), but not to Latrunculin B, an inhibitor of actin polymerization. Using Bodipy‐Sphingomyelin, we have observed formation of vacuole‐like dilations (VLDs), primarily at or close to the adhesion plane, following the reversal from hyper‐ to isotonic medium. VLD formation was not inhibited by Latrunculin B or by amiloride. Phalloidin staining has shown that actin filaments do not surround the vacuoles and latrunculin‐induced depolymerization of actin has actually promoted vacuole formation, even in isotonic conditions. The results support the idea that a decrease in membrane tension promotes the internalization of the plasma membrane.