Bacteria respond to osmotic stress by a substantial increase in the intracellular osmolality, adjusting their cell turgor for altered growth conditions. Using Escherichia coli as a model organism we demonstrate here that bacterial responses to hyperosmotic stress specifically depend on the nature of osmoticum used. We show that increasing acute hyperosmotic NaCl stress above approximately 1.0 Os kg(-1) causes a dose-dependent K(+) leak from the cell, resulting in a substantial decrease in cytosolic K(+) content and a concurrent accumulation of Na(+) in the cell. At the same time, isotonic sucrose or mannitol treatment (non-ionic osmotica) results in a gradual increase of the net K(+) uptake. Ion flux data are consistent with growth experiments showing that bacterial growth is impaired by NaCl at the concentration resulting in a switch from net K(+) uptake to efflux. Microarray experiments reveal that about 40% of upregulated genes shared no similarity in their responses to NaCl and sucrose treatment, further suggesting specificity of osmotic adjustment in E. coli to ionic and non-ionic osmotica. The observed differences are explained by the specificity of the stress-induced changes in the membrane potential of bacterial cells highlighting the importance of voltage-gated K(+) transporters for bacterial adaptation to hyperosmotic stress.