Sphingolipid-gated Ca signaling is mediated through Ca-permeable channels. In this report, we characterize the properties of the channel in a human endothelial cell line (EA.hy926). Ca release from intracellular stores is not antagonized by nifedipine, conotoxin G-VIa, or heparin. To further characterize the molecular properties of the channel, we developed a novel assay to directly measure efflux of Ca from intracellular stores of permeabilized Xenopus oocytes. Following size fractionation by sucrose gradient, poly(A) RNA from EA.hy926 cells is microinjected into oocytes of Xenopuslaevis. We find that the mRNA encoding Ca release activity is 1.5-2.0 kilobases in length. The sphingolipid-gated Ca-permeable channel is thus likely to be a novel Ca-permeable channel distinct from other characterized intracellular Ca channels such as the ryanodyne receptor and the inositol 1,4,5-trisphosphate receptor. The method described here provides a new approach to further characterizing this channel and other intracellular Ca channels.

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