Optical control of calcium-regulated exocytosis
Optical control of calcium-regulated exocytosis
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DOI：
10.1016/j.bbagen.2012.11.003
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发表时间：
2013-03-01
影响因子：
3
通讯作者：
Gorostiza, Pau
中科院分区：
文献类型：
--
作者：
Izquierdo-Serra, Merce;Trauner, Dirk;Gorostiza, Pau
背景：神经元通过化学突触分泌神经递质来相互传递信号，并向肌肉或腺体等非神经元细胞传递信号。为了解剖神经传递的分子机制，需要在突触前末端直接和可逆地触发神经分泌的新方法。在这里，我们利用光门控通道lilur的钙通透性，以可逆地操纵胞质钙浓度，从而控制钙调节的胞外分泌。方法：光刺激表达LiGluR的牛嗜铬细胞。通过安培法或全细胞膜片钳检测胞外事件，定量膜电容和钙内流。结果：安培法显示，光刺激持续触发嗜铬细胞胞吐。儿茶酚胺的分泌可以通过改变照明的波长在零到几赫兹之间调节。在激活LiGluR和天然电压门控钙通道（VGCCs）之间发现分泌功效的差异。我们的研究结果表明，当钙内流由lilur而不是天然vgc触发时，钙内流的位置和准备释放的囊泡之间的距离更长。结论：lilur激活可直接、可逆地增加细胞内钙浓度。光门控钙内流允许第一次控制钙调节胞外分泌，而不需要在染色质细胞中应用去极化溶液或电压夹紧。一般意义：LiGluR是研究神经传递中分泌机制及其时空模式的有用工具，并为研究肌肉收缩或细胞迁移等其他钙依赖过程打开了一扇窗。(C) 2012 Elsevier B.V.版权所有
Background: Neurons signal to each other and to non-neuronal cells as those in muscle or glands, by means of the secretion of neurotransmitters at chemical synapses. In order to dissect the molecular mechanisms of neurotransmission, new methods for directly and reversibly triggering neurosecretion at the presynaptic terminal are necessary. Here we exploit the calcium permeability of the light-gated channel LiGluR in order to reversibly manipulate cytosolic calcium concentration, thus controlling calcium-regulated exocytosis.Methods: Bovine chromaffin cells expressing LiGluR were stimulated with light. Exocytic events were detected by amperometry or by whole-cell patch-clamp to quantify membrane capacitance and calcium influx.Results: Amperometry reveals that optical stimulation consistently triggers exocytosis in chromaffin cells. Secretion of catecholamines can be adjusted between zero and several Hz by changing the wavelength of illumination. Differences in secretion efficacy are found between the activation of LiGluR and native voltage-gated calcium channels (VGCCs). Our results show that the distance between sites of calcium influx and vesicles ready to be released is longer when calcium influx is triggered by LiGluR instead of native VGCCs.Conclusion: LiGluR activation directly and reversibly increases the intracellular calcium concentration. Light-gated calcium influx allows for the first time to control calcium-regulated exocytosis without the need of applying depolarizing solutions or voltage clamping in chromaffin cells.General significance: LiGluR is a useful tool to study the secretory mechanisms and their spatiotemporal patterns in neurotransmission, and opens a window to study other calcium-dependent processes such as muscular contraction or cell migration. (C) 2012 Elsevier B.V. All rights reserved.