Xie Li, Zhang Ming, Zhou Wei, Wu Zhengxing, Ding Jiuping, Chen Liangyi, Xu Tao
Joint Laboratory of Institute of Biophysics and Huazhong University of Science and Technology, National Key Laboratory of Biomacromolecules, Institute of Biophysics, Chinese Academy of Science, Beijing 100101, China.
Traffic. 2006 Apr;7(4):429-39. doi: 10.1111/j.1600-0854.2006.00401.x.
Three different methods, membrane capacitance (C(m)) measurement, amperometry and FM dye labeling were used to investigate the role of extracellular ATP in insulin secretion from rat pancreatic beta cells. We found that extracellular application of ATP mobilized intracellular Ca(2+) stores and synchronously triggered vigorous exocytosis. No influence of ATP on the readily releasable pool of vesicles was observed, which argues against a direct modulation of the secretory machinery at a level downstream of Ca(2+) elevation. The stimulatory effects of ATP were greatly reduced by intracellular perfusion of BAPTA but not EGTA, suggesting a close spatial association of fusion sites with intracellular Ca(2+) releasing sites. ATP-induced Ca(2+) transients and exocytosis were not blocked by thapsigargin (TG), by a ryanodine receptor antagonist or by dissipation of pH in acidic stores by monensin alone, but they were greatly attenuated by IP(3) receptor inhibition as well as ionomycin plus monensin, suggesting involvement of IP(3)-sensitive acidic Ca(2+) stores. Taken together, our data suggest that extracellular ATP triggers exocytosis by mobilizing spatially limited acidic Ca(2+) stores through IP(3) receptors. This mechanism may explain how insulin secretion from the pancreas is coordinated through diffusible ATP that is co-released with insulin.
采用三种不同方法，即膜电容（C(m)）测量、安培测量法和FM染料标记法，研究细胞外ATP在大鼠胰腺β细胞胰岛素分泌中的作用。我们发现，细胞外施加ATP可动员细胞内Ca(2+)储存并同步触发强烈的胞吐作用。未观察到ATP对易释放囊泡池有影响，这表明在Ca(2+)升高下游水平，ATP对分泌机制无直接调节作用。通过细胞内灌注BAPTA而非EGTA，ATP的刺激作用大幅降低，这表明融合位点与细胞内Ca(2+)释放位点在空间上紧密相关。毒胡萝卜素（TG）、ryanodine受体拮抗剂或仅用莫能菌素耗散酸性储存中的pH值，均未阻断ATP诱导的Ca(2+)瞬变和胞吐作用，但IP(3)受体抑制以及离子霉素加莫能菌素可使其大幅减弱，这表明IP(3)敏感的酸性Ca(2+)储存参与其中。综上所述，我们的数据表明，细胞外ATP通过IP(3)受体动员空间有限的酸性Ca(2+)储存来触发胞吐作用。这一机制可能解释了胰腺胰岛素分泌如何通过与胰岛素共同释放的可扩散ATP进行协调。
