Ishihata A, Endoh M
Department of Pharmacology, Yamagata University School of Medicine, Japan.
Life Sci. 1991;48(6):583-91. doi: 10.1016/0024-3205(91)90474-p.
Confluent monolayers of human umbilical vein endothelial cells subcultured on glass coverslips were loaded with the fluorescent Ca2+ indicator, fura-2. Changes in fura-2 fluorescence were detected by means of a fluorescence spectrophotometer. Both ATP and ADP (0.3-100 microM) caused a concentration-dependent transient peak response of the intracellular free calcium concentration ([Ca2+]i), followed by a lower sustained response. AMP and adenosine did not induce detectable changes in [Ca2+]i. The sustained response to ATP was abolished by superfusion with the Ca2(+)-free solution (with 1 mM EGTA), while the transient peak response was uninfluenced. The transient peak response to ATP (30 microM) was inhibited by pre-exposure to ATP in a graded manner depending on the concentration of ATP. The response to ATP recovered after washout for 20 min with the solution containing Ca2+, but not with the Ca2(+)-free solution. The transient peak response to ATP was markedly reduced by preceding exposure to histamine, while the response to histamine was not influenced by pre-exposure to ATP. These findings indicate that depletion and refilling of the ATP-sensitive intracellular Ca2+ store may be responsible for the desensitization and recovery of the ATP-induced [Ca2+]i response. The pharmacological characteristics of the ATP-sensitive intracellular Ca2+ store seem different from those of the histamine-sensitive store.
接种于玻璃盖玻片上并进行传代培养的人脐静脉内皮细胞融合单层，用荧光钙指示剂fura - 2进行负载。通过荧光分光光度计检测fura - 2荧光的变化。ATP和ADP（0.3 - 100微摩尔）均可引起细胞内游离钙浓度（[Ca2 +]i）呈浓度依赖性的瞬时峰值反应，随后是较低的持续反应。AMP和腺苷未诱导[Ca2 +]i出现可检测到的变化。用无钙溶液（含1毫摩尔EGTA）灌流可消除对ATP的持续反应，而瞬时峰值反应不受影响。对ATP（30微摩尔）的瞬时峰值反应会根据ATP浓度以分级方式受到预先暴露于ATP的抑制。用含钙溶液洗脱20分钟后，对ATP的反应恢复，但用无钙溶液则不能恢复。预先暴露于组胺会显著降低对ATP的瞬时峰值反应，而对组胺的反应不受预先暴露于ATP的影响。这些发现表明，ATP敏感的细胞内钙库的耗竭和再充盈可能是ATP诱导的[Ca2 +]i反应脱敏和恢复的原因。ATP敏感的细胞内钙库的药理学特性似乎与组胺敏感的钙库不同。
