ADP-ribose in 3T3 cells stimulated via purinergic P2Y receptors

he role of cyclic ADP-ribose in the amplification of subcellular and global Ca 2 signaling upon stimulation of P2Y purinergic receptors was studied in 3T3 fibroblasts. Either (1) 3T3 fibroblasts (CD38 cells), (2) 3T3 fibroblasts preloaded by incubation with extracellular cyclic ADP-ribose (cADPR), (3) 3T3 fibroblasts microinjected with ryanodine, or (4) 3T3 fibroblasts transfected to express the ADP-ribosyl cyclase CD38 (CD38 cells) were used. Both preincubation with cADPR and CD38 expression resulted in comparable intracellular amounts of cyclic ADP-ribose (42.3 5.2 and 50.5 8.0 pmol/mg protein). T P2Y receptor stimulation of CD38 cells yielded a small increase of intracellular Ca 2 concentration and a much higher Ca 2 signal in CD38-transfected cells, in cADPRpreloaded cells, or in cells microinjected with ryanodine. Confocal Ca 2 imaging revealed that stimulation of ryanodine receptors by cADPR or ryanodine amplified localized pacemaker Ca 2 signals with properties resembling Ca 2