EXTRACELLULAR ATP STIMULATES CALCIUM INFLUX IN NEUROBLASTOMA X GLIOMAHYBRID NG108-15 CELLS

ATP-induced changes in the intracellular Ca2+ concentration ([Ca2+]i) in neuroblastoma X glioma hybrid NG108-15 cells were studied. Using th e fluorescent Ca2+ indicator fura-2, we have shown that the [Ca2+]i in creased in response to ATP. ATP at 3 mM caused the greatest increase i n [Ca2+]i, whereas at higher concentrations of ATP the response became smaller. Two nonhydrolyzable ATP analogues, adenosine 5'-thiotriphosp hate and 5'-adenylyl-beta, gamma-imidodiphosphate, could not trigger s ignificant [Ca2+]i change, but they could block the ATP eff ect. Other adenine nucleotides, including ADP, AMP, alpha,beta-methylene-ATP, be ta, gamma-methylene-ATP, and 2-methylthio-ATP, as well as UTP and aden osine, all had no effect on [Ca2+]i at 3 mM. In the absence of extrace llular Ca2+, the effect of ATP was inhibited totally, but could be res tored by the addition of Ca2+ to the cells. Upon removal of Mg2+, the maximum increase in [Ca2+]i induced by ATP was enhanced by about 42%. Ca2+-channel blockers partially inhibited the ATP-induced [Ca2+]i rise . The ATP-induced [Ca2+]i rise was not affected by thapsigargin pretre atment, though such pretreatment blocked bradykinin-induced [Ca2+]i ri se completely. No heterologous desensitization of [Ca2+]i rise was obs erved between ATP and bradykinin. The magnitude of the [Ca2+]i rise in duced by ATP increased between 1.5 and 3.1 times when external Na+ was replaced with Tris, N-methyl-D-glucamine, choline, or Li+. The additi on of EGTA or verapamil to cells after their maximum response to ATP i mmediately lowered the [Ca2+]i to the basal level in Na-containing or Na-free Tris solution. Our results suggest that ATP stimulates Ca2+ in flux via at least two pathways: ion channels that are permeable to Ca2 + and Na+, and pores formed by ATp4-.