Spirogyra living in running water forms a rhizoid which anchors it to the substratum. Rhizoid differentiation can be induced in the laboratory by severing algal filaments. The terminal cell changes the growth mode from diffuse growth to tip growth, and finally differentiates to be a rhizoid. We found that the intracellular osmolarity of the rhizoid was significantly lower than that of other interjacent cells which did not form rhizoids. The decrease in the intracellular osmolarity began before the start of tip growth. TEA, a K⁺ channel blocker, inhibited the decrease in the intracellular osmolarity of the terminal cells; increase in the external K⁺ also inhibited this. It was suggested that K⁺ efflux through K⁺ channel is involved in the adjustment of osmotic pressure. When the adjustment of osmotic pressure was inhibited, tip growth did not start, inevitably, no rhizoid was formed. In Spirogyra sp. which was unable to form rhizoids, the terminal cell did not show the adjustment of osmotic pressure. Thus, this adjustment seems to be intimately coupled with the rhizoid differentiation. Possible roles of the adjustment of osmotic pressure in rhizoid differentiation are discussed.