Jeremic A, Jeftinija K, Stevanovic J, Glavaski A, Jeftinija S
Department of Biomedical Sciences, Neuroscience Program, Iowa State University, Ames, USA.
J Neurochem. 2001 Apr;77(2):664-75. doi: 10.1046/j.1471-4159.2001.00272.x.
ATP caused a dose-dependent, receptor-mediated increase in the release of glutamate and aspartate from cultured astrocytes. Using calcium imaging in combination HPLC we found that the increase in intracellular calcium coincided with an increase in glutamate and aspartate release. Competitive antagonists of P(2) receptors blocked the response to ATP. The increase in intracellular calcium and release of glutamate evoked by ATP were not abolished in low Ca(2+)-EGTA saline, suggesting the involvement of intracellular calcium stores. Pre-treatment of glial cultures with an intracellular Ca(2+) chelator abolished the stimulatory effects of ATP. Thapsigargin (1 microM), an inhibitor of Ca(2+)-ATPase from the Ca(2+) pump of internal stores, significantly reduced the calcium transients and the release of aspartate and glutamate evoked by ATP. U73122 (10 microM, a phospholipase C inhibitor, attenuated the ATP-stimulatory effect on calcium transients and blocked ATP-evoked glutamate release in astrocytes. Replacement of extracellular sodium with choline failed to influence ATP-induced glutamate release. Furthermore, inhibition of the glutamate transporters p-chloromercuri-phenylsulfonic acid and Ltrans-pyrolidine-2,4-dicarboxylate failed to impair the ability of ATP to stimulate glutamate release from astrocytes. However, an anion transport inhibitor, furosemide, and a potent Cl(-) channel blocker, 5-nitro-2(3-phenylpropylamino)-benzoate, reduced ATP-induced glutamate release. These results suggest that ATP stimulates excitatory amino acid release from astrocytes via a calcium-dependent anion-transport sensitive mechanism.
ATP 可引起培养的星形胶质细胞中谷氨酸和天冬氨酸释放呈剂量依赖性、受体介导的增加。通过结合钙成像和高效液相色谱法，我们发现细胞内钙的增加与谷氨酸和天冬氨酸释放的增加同时发生。P(2) 受体的竞争性拮抗剂可阻断对 ATP 的反应。在低钙 - 乙二醇双四乙酸盐盐溶液中，ATP 引起的细胞内钙增加和谷氨酸释放并未被消除，这表明细胞内钙库参与其中。用细胞内钙螯合剂预处理胶质细胞培养物可消除 ATP 的刺激作用。毒胡萝卜素（1 microM），一种来自内质网 Ca(2+) 泵的 Ca(2+)-ATP 酶抑制剂，可显著降低由 ATP 引起的钙瞬变以及天冬氨酸和谷氨酸的释放。U73122（10 microM，一种磷脂酶 C 抑制剂）可减弱 ATP 对钙瞬变的刺激作用，并阻断 ATP 诱发的星形胶质细胞谷氨酸释放。用胆碱替代细胞外钠未能影响 ATP 诱导的谷氨酸释放。此外，抑制谷氨酸转运体对氯汞苯磺酸和 L-反式-吡咯烷-2,4-二羧酸未能损害 ATP 刺激星形胶质细胞释放谷氨酸的能力。然而，一种阴离子转运抑制剂呋塞米和一种强效 Cl(-) 通道阻滞剂 5-硝基-2(3-苯丙基氨基)-苯甲酸可降低 ATP 诱导的谷氨酸释放。这些结果表明，ATP 通过一种钙依赖性阴离子转运敏感机制刺激星形胶质细胞释放兴奋性氨基酸。
