Astrocytes display large resting K⁺ conductances although the underlying channel(s) are ill defined. In this study, freshly isolated astrocytes of mouse hippocampus were employed to analyze K⁺ channel expression, combining patch clamp analyses in transgenic mice, immunohistochemistry, immunoblot and single cell RT-PCR. Beyond postnatal (p) day 3, the resting currents were sensitive to Ba²⁺ at submillimolar concentrations. Transcripts encoding the Kir channel subunit Kir4.1 were encountered in all cells investigated while Kir5.1, another glial Kir subunit that co- assembles with Kir4.1, was co-expressed scarcely. Kir4.1 protein was up-regulated during early postnatal development. This finding was confirmed with semi-quantitative real-time RT-PCR revealing a strong rise in Kir4.1 mRNA between p3 and p10, both in tissue samples from the CA1 region as well as in single astrocytes. Electrophysiological analysis confirmed expression of Ba²⁺-sensitive Kir currents in p3 astrocytes, but the amplitudes per cell surface area amounted only to 25% compared with astrocytes at p10. In Kir4.1-deficient mice, astrocytes were almost devoid of Ba²⁺- sensitive currents and displayed more positive resting potentials. Application of quinine led to the block of outwardly rectifying currents, suggesting the expression of K2P channels in astrocytes. Arachidonic acid enhanced the amplitude of quinine-sensitive currents. Molecular analysis suggested functional expression of TREK-1 channels in astrocytes. The increase in astroglial Kir4.1 current density observed here parallels a significant drop of the extracellular volume fraction of the developing hippocampus, suggesting an important role of Kir4.1 in extracellular K⁺ buffering. Whether K2P channels in astrocytes have also a role in spatial redistribution of K⁺ remains to be elucidated. Supported by DFG (SFB TR3; SPP 1172 SE774/3) and EC (FP7-202167 Neuroglia).