Nicotinic Receptor‐Mediated Release of Noradrenaline in the Human Neuroblastoma SH‐SY5Y

Abstract

Dimethylphenylpiperazinium iodide (a nicotinic agonist) evokes noradrenaline release from human neuroblastoma SH-SY5Y cells that have been pretreated with 12-O-tetradecanoylphorbol 13-acetate for 8 min. This effect of dimethylphenylpiperazinium iodide was inhibited by 1 microM mecamylamine but not by 1 microM atropine, which suggests that SH-SY5Y cells express nicotinic receptors coupled to the release of noradrenaline. Dimethylphenylpiperazinium iodide-evoked release was enhanced by 5 microM Bay K 8644 (an L-type calcium agonist) and inhibited by 1 microM nifedipine. Dimethylphenylpiperazinium iodide depolarised SH-SY5Y cells and enhanced the level of intracellular calcium in cells loaded with fura 2. The effects of dimethylphenylpiperazinium iodide on noradrenaline release, depolarisation, and intracellular calcium levels were all inhibited by 1 microM desmethylimipramine. The results of this study show that nicotinic receptors in SH-SY5Y cells stimulate noradrenaline release by activation of L-type calcium channels.

Bradykinin-evoked Release of [3H]Noradrenaline from the Human Neuroblastoma SH-SY5Y

“…However pretreatment of cells with IOOnM TPA for 8 min increases BK-evoked release approximately 2-fold. This suggests that BK-evoked release is enhanced by activation of protein kinase C as has previously been reported in SHSYSY for potassium-and muscarinic receptor M3-evoked [S] and for nicotinic receptorevoked release of NA [3]. The inhibition of BK-evoked release by the B2selective antagonists [D-Phe7]BK and [ThiS,8,DPhe7]BK but not by the BI selective antagonist [Des-Arg9, Leu81BK [ 1 I ] suggests that the 8 2 subtype rather than the BI subtype is coupled to NA release in SHSYSY.…”

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Our previous studies have shown that cultures of the human neuroblastoma SHSYSY express depolarisation-evoked release of NA stimulated by elevated potassium [ 1.21 or activation of nicotinic receptors [3] which is coupled to L-type calcium channels. In

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“…The human adrenergic clone SHSYSY was cultured in a 1:1 mixture of Ham's F12 and Eagle's minimal essential medium containing nonessential amino acids, supplemented with 10% foetal calf serum in a 98% air/ 2% C02 [1,3]. Release of NA ,was measured from cells subcultured in 24-well plates as described previously [ 1.31.…”

“…Release of NA ,was measured from cells subcultured in 24-well plates as described previously [ 1.31. For measurements of intracellular calcium and patch-clamp studies cells were plated onto glass cover slips at the same seeding density and changes in intracellular calcium or membrane potential recorded as described previously [3,9] or using the perforatedpatch technique [ 101. BK evokes NA release from SH-SYSY cells over the concentration range 10 to IOOOnM with maximal stimulation of 2% above basal by IOOOnM and an ECSO value of 20nM Pretreatment of cell layers with lOOnM TPA for 8min before the addition of BK enhanced NA release at all concentrations to a maximum of 4% above basal release without altering the EC50 value.…”

Calcium Flux in Neuroblastoma Cells Is a Coupling Mechanism between Non-Genomic and Genomic Modes of Estrogens

“…While L-, P- and N-type VGCCs have been reported to exist in human neuroblastoma cell lines [65,66,67], the presence of Q-type VGCCs in neurons is unclear. In this study, blockade of the L- and P-type VGCC in either pulse did not prevent E-BSA from activating later genomic transcriptional events from a consensus ERE, suggesting that the role of L- and P-type VGCC is not significant in E-BSA-induced transcription potentiation in SK-N-BE(2)C cells.…”