AbstractChronic loss of intracellular K+ can induce neuronal apoptosis in pathological conditions. However, the mechanism by which the K+ channels are regulated in this process remains largely unknown. Here, we report that the increased membrane expression of Kv2.1 proteins in cortical neurons deprived of serum, a condition known to induce K+ loss, promotes neuronal apoptosis. The increase in IK current density and apoptosis in the neurons deprived of serum were inhibited by a dominant negative form of Kv2.1 and MK801, an antagonist to NMDA receptors. The membrane level of Kv2.1 and its interaction with SNAP25 were increased, whereas the Kv2.1 phosphorylation was inhibited in the neurons deprived of serum. Botulinum neurotoxin, an agent known to prevent formation of soluble N‐ethylmaleimide‐sensitive factor attachment protein receptor complex, suppressed the increase in IK current density. Together, these results suggest that NMDA receptor‐dependent Kv2.1 membrane translocation is regulated by a soluble N‐ethylmaleimide‐sensitive factor attachment protein receptor‐dependent vesicular trafficking mechanism and is responsible for neuronal cell death induced by chronic loss of K+.