Activation of galanin receptor 2 stimulates large conductance Ca2+-dependent K+ (BK) channels through the IP3 pathway in human embryonic kidney (HEK293) cells

The large conductance Ca2+-activated K+ (BK) channels are widely distributed in the brain, and act as intracellular calcium sensors in neurons. They play an important feedback role in controlling Ca2+ flux and Ca2+-dependent processes, including neurotransmitter release and cellular excitability. In this study, the effects of the neuropeptide galanin on BK channels were examined by determining the whole-cell currents and single-channel activities in human embryonic kidney (HEK293) cells co-expressing GalR2 and the BK alpha subunit. Galanin enhanced the currents of BK channels, in a concentration-dependent and PTX-independent manner, with an ED50 value of 71.8 ± 16.9 nM. This activation was mediated by GalR2, since its agonist AR-M1896 mimicked the effect of galanin, and since galanin did not facilitate BK currents in cells co-expressing cDNAs of BK and GalR1 or GalR3. The galanin-induced BK current persisted after replacement with Ca2+-free solution, suggesting that extracellular Ca2+ is not essential. Chelating intracellular Ca2+ by either the slow Ca2+ buffer EGTA or the fast Ca2+ buffer BAPTA abolished galanin-mediated activation of BK channels, indicating the important role of intracellular Ca2+. The role of Ca2+ efflux from the sarcoplasmic reticulum/endoplasmic reticulum (SR/ER) was confirmed by application of thapsigargin, an irreversible inhibitor that depletes Ca2+ from SR/ER. Moreover, the inositol-1,4,5-triphosphate receptor (IP3R) was identified as the mediator responsible for increased intracellular Ca2+ activating BK channels. Taken together, activation of GalR2 leads to elevation of intracellular Ca2+ is due to Ca2+ efflux from ER through IP3R sequentially opening BK channels.