Highlights d KCC2 interacts with Task-3 potassium channels and regulates their membrane traffic d KCC2 knockdown depolarizes rat hippocampal neurons and increases their excitability d Chronic KCC2 knockdown in rat dentate gyrus is not sufficient to induce seizures d Network activity is altered by KCC2 knockdown, mostly independent of GABA signaling

Graphical AbstractHighlights d KCC2 interacts with Task-3 potassium channels and regulates their membrane traffic d KCC2 knockdown depolarizes rat hippocampal neurons and increases their excitability d Chronic KCC2 knockdown in rat dentate gyrus is not sufficient to induce seizures d Network activity is altered by KCC2 knockdown, mostly independent of GABA signaling SUMMARY KCC2 regulates neuronal transmembrane chloride gradients and thereby controls GABA signaling in the brain. KCC2 downregulation is observed in numerous neurological and psychiatric disorders. Paradoxical, excitatory GABA signaling is usually assumed to contribute to abnormal network activity underlying the pathology. We tested this hypothesis and explored the functional impact of chronic KCC2 downregulation in the rat dentate gyrus. Although the reversal potential of GABAA receptor currents is depolarized in KCC2 knockdown neurons, this shift is compensated by depolarization of the resting membrane potential. This reflects downregulation of leak potassium currents. We show KCC2 interacts with Task-3 (KCNK9) channels and is required for their membrane expression. Increased neuronal excitability upon KCC2 suppression altered dentate gyrus rhythmogenesis, which could be normalized by chemogenetic hyperpolarization. Our data reveal KCC2 downregulation engages complex synaptic and cellular alterations beyond GABA signaling that perturb network activity thus offering additional targets for therapeutic intervention.