Selective Activation of KCa3.1 and CRAC Channels by P2Y2 Receptors Promotes Ca2+ Signaling, Store Refilling and Migration of Rat Microglial Cells

Microglial activation involves Ca2+ signaling, and numerous receptors can evoke elevation of intracellular Ca2+. ATP released from damaged brain cells can activate ionotropic and metabotropic purinergic receptors, and act as a chemoattractant for microglia. Metabotropic P2Y receptors evoke a Ca2+ rise through release from intracellular Ca2+ stores and store-operated Ca2+ entry, and some have been implicated in microglial migration. This Ca2+ rise is expected to activate small-conductance Ca2+-dependent K+ (SK) channels, if present. We previously found that SK3 (KCa2.3) and KCa3.1 (SK4/IK1) are expressed in rat microglia and contribute to LPS-mediated activation and neurotoxicity. However, neither current has been studied by elevating Ca2+ during whole-cell recordings. We hypothesized that, rather than responding only to Ca2+, each channel type might be coupled to different receptor-mediated pathways. Here, our objective was to determine whether the channels are differentially activated by P2Y receptors, and, if so, whether they play differing roles. We used primary rat microglia and a rat microglial cell line (MLS-9) in which riluzole robustly activates both SK3 and KCa3.1 currents. Using electrophysiological, Ca2+ imaging and pharmacological approaches, we show selective functional coupling of KCa3.1 to UTP-mediated P2Y2 receptor activation. KCa3.1 current is activated by Ca2+ entry through Ca2+-release-activated Ca2+ (CRAC/Orai1) channels, and both CRAC/Orai1 and KCa3.1 channels facilitate refilling of Ca2+ stores. The Ca2+ dependence of KCa3.1 channel activation was skewed to abnormally high concentrations, and we present evidence for a close physical association of the two channel types. Finally, migration of primary rat microglia was stimulated by UTP and inhibited by blocking either KCa3.1 or CRAC/Orai1 channels. This is the first report of selective coupling of one type of SK channel to purinergic stimulation of microglia, transactivation of KCa3.1 channels by CRAC/Orai1, and coordinated roles for both channels in store refilling, Ca2+ signaling and microglial migration.