Changes in the plasma membrane potential of two histamine-releasing cells, rat peritoneal mast cells and basophilic leukemia cells 2H3 (RBL 2H3), were recorded with the potential-sensitive dye bis-oxonol. For mast cells, the presence of ouabain or the absence of K+ increased the fluorescence intensity of bis-oxonol; gramicidin had no effect. For RBL 2H3 cells, the presence of ouabain and the absence of K+ also increased bis-oxonol fluorescence but gramicidin also increased it. These results show that the plasma membrane potential of RBL 2H3 cells is set, in part, by the activity of the Na+ pump and in part by the K+ conductance, while that of rat mast cells is set predominantly by the Na+ pump.