Intracellular Calcium Elevation Induced by Extracellular Application of Cyclic-ADP-Ribose or Oxytocin is Temperature-Sensitive in Rodent NG108-15 Neuronal Cells with or without Exogenous Expression of Human Oxytocin Receptors

ADP-ribosyl cyclase and/or CD38 are activated after oxytocin receptor stimulation in the hypothalamus and pituitary in adult mice, leading to facilitation of oxytocin secretion. Although cyclic adenosine 5'-diphosphoribose (cADPR) primarily acts as an intracellular second messenger, it has been suggested that extracellular cADPR stimulates intracellular ryanodine receptors after internalisation via the nucleotide-transporting capacity of CD38 in fibroblasts and astrocytes. However, little is known about whether extracellular cADPR activates neurones. To address this question, we used a model neuronal cell line, NG108-15 mouse neuroblastoma x rat glioma hybrid cells possessing CD38 but not oxytocin receptors, and measured cytosolic free calcium concentrations ([Ca2+](i)). Extracellular application of cADPR to NG108-15 cells elevated [Ca2+](i) at 35 degrees C. The elevation was significantly enhanced when measured at 40 degrees C. The cADPR and heat-induced [Ca2+](i) increase were blocked under extracellular Ca2+-free conditions and by 2-aminoethoxydiphenyl borate, an antagonist of melastatin-related transient receptor potential channel 2 (TRPM2) cation channels. Reverse transcriptation-polymerase chain reaction analyses indicated that TRPM2 channels were expressed in NG108-15 cells. Application of oxytocin elevated [Ca2+](i) in NG108-15 cells transformed to transiently express cloned human oxytocin receptors. The oxytocin-induced [Ca2+](i) response was also enhanced by heat. These results indicate that the extracellular application of cADPR, together with heat, activates cation influx downstream of oxytocin receptor signalling in NG108-15 neuronal cells, and suggest the possible involvement of TRPM2 channels in oxytocin release in the mammalian brain.