In response to damage of the brain, astrocytes are activated and increase expression of GFAP. In this study, I found that activated astrocytes increased expression of Kir4.1, a major potassium channel in astrocytes, with markers of neural stem cells including Sox2 and nestin. Kir4.1 expression was also increased with that of nestin and Sox2 when spheres were formed from dissociated P7 mouse brain. In experiments to find out correlation between Kir4.1 and acquisition of stemness, BaCl2, an inhibitor of Kir4.1, dose-dependently increased size of spheres and Sox2 levels although nestin levels were little changed. In further experiments, expression and functions of Kir4.1 in differentiation of neural stem cells were examined. For this, stem cells were cultured from E13 mouse brain were differentiated. At 1 d after differentiation of stem cells was induced by withdrawing EGF and FGF from the culture media, Kir4.1 expression sharply increased, and then decreased thereafter to 5 d while Sox 2 and nestin levels continuously decreased to 5 d. BaCl2, however, did not change expression levels of Sox2 and nestin as well as differentiation markers, GFAP and TUJ-1. Taken together, these results suggest that Kir4.1 may control gain of stemness but not differentiation of stem cells.