Modulation of mitochondrial Ca2+ in ECV304 endothelial cells by agents which elevate cAMP.

Abstract In the human umbilical vein endothelial cell-derived cell line, ECV304, we have previously shown that the elevation of [Ca 2+ ] m in response to agonist stimulation is dependent on Ca 2+ influx, i.e. an ATP-induced sustained increase in [Ca 2+ ] c results in a slow-onset, sustained elevation in [Ca 2+ ] m [Lawrie A.M., Rizzuto R., Pozzan T., Simpson A.W.M. A role for calcium influx in the regulation of mitochondrial calcium in endothelial cells. J Biol Chem 1996; 271 : 10753–10759]. In this study, we have investigated the effect of raising CAMP on ATP-evoked elevations in both [Ca 2+ ] m and [Ca 2+ ] c by: (i) activating adenylate cyclase with the forskolin analogue — forskolin 6-[3′-(N,N-dimethylamino-propionyl)]-HCI (1 μM) (FA); (ii) addition of membrane permeable dibutyryl-CAMP (100 μM) (dbcAMP); and (iii) a combination of FA plus inhibition of CAMP phosphodiesterase using RO 20–1724 (17.5 μM) (RO);. We have found that protocols aimed at elevating CAMP significantly reduce the ATP-evoked (1–10 μM) rise in [Ca 2+ ] m ( n = 14); however, the [Ca 2+ ] c response to ATP was not affected ( n = 33). This new evidence shows that a second messenger system, other than Ca 2+ itself, may influence [Ca 2+ ]m changes in response to agonist stimulation.