SH-SY5Y细胞中[3H]-去甲肾上腺素释放机制的研究：钙离子和环磷酸腺苷的作用

Studies on the mechanism of [3H]-noradrenaline release from SH-SY5Y cells: the role of Ca2+ and cyclic AMP.

Atcheson R, Lambert D G, Hirst R A, Rowbotham D J

University Department of Anaesthesia, Leicester Royal Infirmary.

Br J Pharmacol. 1994 Mar;111(3):787-92. doi: 10.1111/j.1476-5381.1994.tb14806.x.

Abstract

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.

摘要

研究了钙离子（Ca2+）和3':5'-环磷酸腺苷（环磷酸腺苷，cyclic AMP）在卡巴胆碱和钾离子（K+）刺激人神经母细胞瘤SH-SY5Y细胞释放[3H]-去甲肾上腺素过程中的作用。2. 卡巴胆碱和K+均引起[3H]-去甲肾上腺素释放的时间和剂量依赖性刺激。灌注细胞中的释放事件是单相的。在静态孵育（3分钟）的细胞中测得的半最大刺激浓度分别为38±4微摩尔/升和63±4毫摩尔/升。添加的100毫摩尔/升K+诱发的释放大于1毫摩尔/升卡巴胆碱产生的释放。3. 卡巴胆碱和K+均引起环磷酸腺苷形成的时间和剂量（在3分钟时测量）依赖性刺激，半最大刺激分别出现在5±1微摩尔/升和49±2毫摩尔/升。与对释放的影响相反，卡巴胆碱对环磷酸腺苷形成的刺激大于K+。4. K+刺激的[3H]-去甲肾上腺素释放完全依赖于Ca2+内流，因为2.5毫摩尔/升的镍离子（Ni2+）消除了释放。然而，卡巴胆碱诱发的（1毫摩尔/升）释放似乎不受Ni2+预处理的影响。5. 这些数据表明，在SH-SY5Y细胞中，环磷酸腺苷水平升高并不直接参与[3H]-去甲肾上腺素的释放。此外，卡巴胆碱刺激的释放很大程度上独立于细胞外Ca2+，这可能意味着细胞内储存的Ca2+在释放过程中起作用。