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 ([Ca(2+)](i)). Extracellular application of cADPR to NG108-15 cells elevated [Ca(2+)](i) at 35 degrees C. The elevation was significantly enhanced when measured at 40 degrees C. The cADPR and heat-induced [Ca(2+)](i) increase were blocked under extracellular Ca(2+)-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 [Ca(2+)](i) in NG108-15 cells transformed to transiently express cloned human oxytocin receptors. The oxytocin-induced [Ca(2+)](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.
Amina
S
S
Department of Biophysical Genetics, Kanazawa University Graduate School of Medicine, Kanazawa 920-8640, Japan. sarwat_amina@hotmail.com
Hashii
M
M
Ma
W-J
WJ
Yokoyama
S
S
Lopatina
O
O
Liu
H-X
HX
Islam
M S
MS
Higashida
H
H
D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002460:Cell Line; D036563:Cyclic ADP-Ribose / Q000494:pharmacology*; D017931:DNA Primers; D006801:Humans; D009474:Neurons / Q000187:drug effects* / Q000378:metabolism; D010121:Oxytocin / Q000494:pharmacology*; D018045:Receptors, Oxytocin / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012377:Rodentia; D013696:Temperature
D017931:DNA Primers; D018045:Receptors, Oxytocin; D036563:Cyclic ADP-Ribose; D010121:Oxytocin; D002118:Calcium