Metabolic pools of ATP in cultured bovine adrenal medullary chromaffin cells.
Metabolic pools of ATP in cultured bovine adrenal medullary chromaffin cells.
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培养的牛肾上腺髓质嗜铬细胞中 ATP 的代谢池。
DOI：
10.1111/j.1471-4159.1986.tb00702.x
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发表时间：
1986
影响因子：
4.7
通讯作者：
Kirshner,N
中科院分区：
文献类型：
--
作者：
Corcoran,JJ;Korner,M;Caughey,B;Kirshner,N
培养的牛肾上腺嗜铬细胞含有一个池的ATP螯合内的嗜铬囊泡和囊外池的ATP。在以前的研究中，它表明，在囊外池ATP的营业额是双相的。一个阶段的t1/2为3.5-4.5 h，而第二个阶段的t1/2为数天。本文所述的研究通过检测代谢抑制剂腺苷和毛地黄皂苷对ATP利用率和亚细胞定位的影响，进一步表征囊泡和囊外ATP池，这些影响发生在[3 H]腺苷和32 Pi标记后即刻和48 h。在标记氰化物、2-脱氧-D-葡萄糖、β-胆甾-1，5-内酯的组合后，30分钟内标记的ATP立即消耗90-95%，但内源性ATP仅消耗25%。标记后48小时，添加抑制剂导致[3 H]ATP消耗70%，但[32 P]ATP和内源性ATP仅消耗25%。在标记后即刻或48 h，向培养基中加入10 μMadenosine，在检测的细胞中导致类似的[3 H] ATP损失。标记后立即加入腺苷可增加[32 P]ATP的量，但标记后48 h加入腺苷对[32 P]ATP含量无影响。通过用毛地黄皂苷处理质膜的透化导致标记后立即检查的细胞中标记的ATP损失90-95%，但仅20%的内源性ATP损失，并且在标记后48小时检查的细胞中[3 H]ATP损失79%，[32 P]ATP损失32%，内源性ATP损失23%。这些研究表明，在嗜铬囊泡内所含的ATP是代谢惰性的，不容易与胞质ATP交换，但它们不区分标记ATP的囊泡外池。ATP产生突然停止后ATP消失的计算表明，培养的肾上腺嗜铬细胞利用ATP的速率为0.24 nmol/min/106个细胞，相当于囊外池的周转时间为12.5 min。
Cultured bovine adrenal chromaffin cells contain a pool of ATP sequestered within the chromaffin vesicles and an extravesicular pool of ATP. In a previous study it was shown that the turnover of ATP in the extravesicular pool was biphasic. One phase occurred with a t1/2of 3.5–4.5 h whereas the second phase occurred with a t1/2of several days. The studies described here were undertaken to characterize further the vesicular and extravesicular pools of ATP by examining the effects of metabolic inhibitors, adenosine, and digitonin on ATP utilization and subcellular localization immediately after and 48 h after labeling with [3H]adenosine and32Pi. Immediately after labeling a combination of cyanide, 2‐deoxy‐D‐glucose, the β‐glucono‐l,5‐lactone resulted in a 90–95% depletion of the labeled ATP but only a 25% depletion of the endogenous ATP within 30 min. Forty‐eight hours after labeling, addition of the inhibitors resulted in a 70% depletion of the [3H]ATP but only a 25% depletion of the [32P]ATP and endogenous ATP. Addition of 10 μMadenosine to the media resulted in a similar loss of [3HJATP in cells examined immediately after or 48 h after labeling. Adenosine increased the amounts of [32P]ATP when added immediately after labeling but had no effect on the [32P]ATP content when added 48 h after labeling. Permeabilization of the plasma membrane by treatment with digitonin resulted in a loss of 90–95% of the labeled ATP but only 20% of the endogenous ATP in cells examined immediately after labeling and in a 79% loss of [3H]ATP, a 32% loss of [32P]ATP, and a 23% loss of endogenous ATP in cells examined 48 h after labeling. These studies show that ATP contained within chromaffin vesicles is metabolically inert and does not exchange readily with cytosolic ATP, but they do not differentiate extravesicular pools of labeled ATP. Calculation of ATP disappearance after abrupt cessation of ATP production indicates that cultured adrenal chromaffin cells utilize ATP at a rate of 0.24 nmol/min/106cells, corresponding to a turnover time of 12.5 min for the extravesicular pool.