In most neurons, efficient setting of the intracellular Cl--concentration requires the Coordinated regulation of the Cl--inward transporter NKCC1 and the Cl--outward transporter KCC2. Previously, the cation-chloride cotransporter interacting protein 1 (CIP1) was shown to inactivate NKCC1. Here, we investigated its role for KCC2 activity. After co-expression of CIP1 and KCC2 in HEK-293 cells, a physical and functional interaction was observed. CIP1 co-purified with KCC2 in pull-down experiments and significantly increased KCC2 transport activity, as determined by Rb-86(+) flux measurements. RT-PCR analysis demonstrated a ubiquitous expression during postnatal development of the rat. Real-time PCR revealed a two-fold down-regulation of CIP1 during maturation of the rat brain. Taken together, our data identify CIP1 as a potent activator of KCC2. Furthermore, the results support previous data of heteromer formation among members of the cation-chloride cotransporter gene family. (C) 2009 Elsevier Inc. All rights reserved.