Discriminating between capacitative and arachidonate-activated Ca2+ entry pathways in HEK293 cells

We have recently questioned whether the capacitative or store-operated model for receptor-activated Ca2+ entry can account for the influx of Ca2+ seen at low agonist concentrations, such a those typically producing [Ca2+](i) oscillations. Instead, we have identified an arachidonic: acid-regulated, noncapacitative Ca2+ entry mechanism that appears to be specifically responsible for the receptor-activated entry of Ca2+ under these conditions. However, it is unclear whether these two systems reflect the activity of distinct entry pathways or simply different mechanisms of regulating a common pathway. We therefore used the known selectivity of the Ca2+-stimulated type vm adenylyl cyclase for Ca2+ entry occurring via the capacitative pathway (Fagan, K. A., Mahey, R., and Cooper, D. M. F. (1996) J. Biol. Chem. 271, 12438-12444) to attempt to discriminate between these two entry mechanisms in HEK293 cells. Consistent with the earlier reports, we found that thapsigargin induced an approximate 3-fold increase in adenylyl cyclase activity that was unrelated to global changes in [Ca2+](i) or to the release of Ca2+ from internal stores but was specifically dependent on the induced capacitative entry of Ca2+. In marked contrast, the arachidonate-induced entry of Ca2+ completely failed to affect adenylyl cyclase activity despite producing a substantially greater rate of entry than that induced by thapsigargin, These data demonstrate that the arachidonate-activated entry of Ca2+ occurs via an entirely distinct influx pathway.