Studies on the mechanism of [3H]-noradrenaline release from SH-SY5Y cells: the role of Ca2+ and cyclic AMP.
1. The roles of both Ca2+ and adenosine 3':5'-cyclic monophosphate (cyclic AMP) in carbachol and K(+)-stimulated [3H]-noradrenaline release from SH-SY5Y human neuroblastoma cells were examined. 2. Both carbachol and K+ caused a time- and dose-related stimulation of [3H]-noradrenaline release. The release event in perfused cells was monophasic. Half-maximum stimulation measured in statically incubated (3 min) cells was 38 +/- 4 microM and 63 +/- 4 mM respectively. K+ (100 mM, added)-evoked release was greater than that produced by carbachol (1 mM). 3. Both carbachol and K+ caused a time- and dose (measured at 3 min)-related stimulation of cyclic AMP formation with half-maximum stimulation occurring at 5 +/- 1 microM and 49 +/- 2 mM respectively. In contrast to its effects on release, carbachol produced a greater stimulation of cyclic AMP formation than K+. 4. K(+)-stimulated [3H]-noradrenaline release was entirely dependent on Ca2+ entry as 2.5 mM Ni2+ abolished release. However, carbachol-evoked (1 mM) release appeared to be unaffected by Ni2+ pretreatment. 5. These data suggest that in SH-SY5Y cells, elevated cyclic AMP levels are not directly involved in [3H]-noradrenaline release. In addition, carbachol-stimulated release is largely independent of extracellular Ca2+ possibly implying a role for intracellular stored Ca2+ in the release process.
University Department of Anaesthesia, Leicester Royal Infirmary.
R
R
Atcheson
D G
DG
Lambert
R A
RA
Hirst
D J
DJ
Rowbotham
D002118:Calcium / Q000502:physiology*; D002121:Calcium Channel Blockers / Q000494:pharmacology; D002217:Carbachol / Q000494:pharmacology; D000242:Cyclic AMP / Q000502:physiology*; D006801:Humans; D007700:Kinetics; D008954:Models, Biological; D009447:Neuroblastoma; D009474:Neurons / Q000378:metabolism* / Q000502:physiology; D009638:Norepinephrine / Q000378:metabolism*; D011188:Potassium / Q000494:pharmacology; D013268:Stimulation, Chemical; D014316:Tritium; D014407:Tumor Cells, Cultured
D002121:Calcium Channel Blockers; D014316:Tritium; D002217:Carbachol; D000242:Cyclic AMP; D011188:Potassium; D002118:Calcium; D009638:Norepinephrine