Osmotic swelling of human and rat erythrocytes does not induce regulatory volume decrease. Regulatory volume increase was observed in shrunken erythrocytes of rats only. This reaction was blocked by the inhibitors of Na+/H+ exchange. Cytoplasmic acidification in erythrocytes of both species increases the amiloride-inhibited component of22Na influx by five- to eight-fold. Both the osmotic and isosmotic shrinkage of rat erythrocytes results in the 10- to 30-fold increase of amiloride-inhibited22Na influx and a two-fold increase of furosemide-inhibited86Rb influx. We failed to indicate any significant changes of these ion transport systems in shrunken human erythrocytes. The shrinking of quin 2-loaded human and rat erythrocytes results in the two- to threefold increase of the rate of45Ca influx, which is completely blocked by amiloride. The dependence of volume-induced22Na influx in rat erythrocytes and45Ca influx in human erythrocytes on amiloride concentration does not differ. The rate of45Ca influx in resealed ghosts was reduced by one order of magnitude when intravesicular potassium and sodium were replaced by choline. It is assumed that the erythrocyte shrinkage increases the rate of a nonselective Cao2+(Nai+, Ki+) exchange. Erythrocyte shrinking does not induce significant phosphorylation of membrane protein but increases the32P incorporation in diphosphoinositides. The effect of shrinkage on the32P labeling of phosphoinositides is diminished after addition of amiloride. It is assumed that volume-induced phosphoinositide response plays an essential role in the mechanism of the activation of transmembrane ion movements.