KCC2 drives chloride microdomain formation in dendritic blebbing.

Intracellular chloride ion concentration ([Cl]) homeostasis is critical for excitatory/inhibitory balance and volume regulation in neurons. We quantitatively map spatiotemporal dendritic [Cl] dynamics during N-methyl-d-aspartate (NMDA) excitotoxicity to determine how Cl changes contribute to localized dendritic swelling (blebbing) in stroke-like conditions. Whole-cell patch clamp electrophysiology combined with simultaneous fluorescence lifetime imaging (FLIM) of the Cl dye N-(ethoxycarbonylmethyl)-6-methoxyquinolinium bromide (MQAE; MQAE-FLIM) reliably report resting and dynamic [Cl] shifts in dendrites. NMDA application generates spatially restricted and persistent high [Cl] subdomains at dendritic blebs in a process that requires Ca influx and the subsequent opening of small-conductance Ca-activated K (SK) channels. We propose sustained and localized K efflux increased extracellular K concentrations ([K]) sufficiently at discrete regions to reverse K-Cl cotransporter (KCC2) transport and trigger synaptic swelling. Together, our data establish a mechanism for KCC2 to generate pathological [Cl] microdomains in blebbing with relevance for multiple neurological disorders.