ISOLATED SECRETION GRANULES FROM PAROTID-GLANDS OF CHRONICALLY STIMULATED RATS POSSESS AN ALKALINE INTERNAL PH AND INWARD-DIRECTED H+ PUMP ACTIVITY
ISOLATED SECRETION GRANULES FROM PAROTID-GLANDS OF CHRONICALLY STIMULATED RATS POSSESS AN ALKALINE INTERNAL PH AND INWARD-DIRECTED H+ PUMP ACTIVITY
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DOI：
10.1083/jcb.103.4.1257
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发表时间：
1986-10-01
影响因子：
7.8
通讯作者：
CASTLE, JD
中科院分区：
文献类型：
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作者：
ARVAN, P;CASTLE, JD
已从长期用β-肾上腺素能激动剂异丙肾上腺素刺激的大鼠腮腺中分离出分泌颗粒。这些颗粒之所以令人感兴趣，是因为与正常腺体的颗粒相比，它们包装了一组数量不同的分泌蛋白。富含脯氨酸、甘氨酸和谷氨酰胺的多肽，已知其pI'''s>10，取代α-淀粉酶(pI''s=6.8)作为主要内容物种类。接受治疗的大鼠颗粒的内部pH值范围为硫酸钾介质中的7.8至氯化胆碱介质中的6.9。与正常腮腺颗粒相比（约 6.8）增加的 pH 值似乎反映了分泌内容物组成的变化。虽然正常成熟腮腺颗粒的 H+ 泵 ATP 酶活性水平几乎可以忽略不计（Arvan, P., G. Rudnick, 和 J.D. Castle, 1985, J. Biol. Chem., 260, 14945-14952），但从异丙肾上腺素处理的大鼠中分离的颗粒经历了时间依赖性内部酸化（约 0.2 pH 单位），需要 ATP 的存在并被 H+ 离子载体消除。此外，添加 ATP 后，会产生内部阳性颗粒跨膜电位，这取决于 ATP 水解。使用了两种独立的方法来排除污染细胞器是 H+-ATPase 活性来源的可能性。这些数据共同提供了明确的证据，证明长期受刺激的大鼠腮腺颗粒膜中存在 H+ 泵。然而，尽管存在 H+ 泵活性，但使用弱碱吖啶橙的荧光显微镜显示，活细胞的颗粒内 pH 值高于细胞质的 pH 值。
Secretion granules have been isolated from the parotid glands of rats that have been chronically stimulated with the .beta.-adrenergic agonist, isoproterenol. These granules are of interest because they package a quantitatively different set of secretory proteins in comparison with granules from the normal gland. Polypeptides enriched in proline, glycine, and glutamine, which are known to have pI''s > 10, replace .alpha.-amylase (pI''s = 6.8) as the principal content species. The internal pH of granules from the treated rats ranges from 7.8 in a potassium sulfate medium to 6.9 in a choline chloride medium. The increased pH over that of normal parotid granules (.apprx. 6.8) appears to reflect the change in composition of the secretory content. Whereas normal mature parotid granules have practically negligible levels of H+ pumping ATPase activity (Arvan, P., G. Rudnick, and J.D. Castle, 1985, J. Biol. Chem., 260, 14945-14952) the isolated granules from isoproterenol-treated rats undergo a time-dependent internal acidification (.apprx. 0.2 pH unit) that requires the presence of ATP and is abolished by an H+ ionophore. Additionally, an inside-positive granule transmembrane potential develops after ATP addition that depends upon ATP hydrolysis. Two independent methods have been used that exclude the possibility that contaminating organelles are the source of the H+-ATPase activity. Together these data provide clear evidence for the presence of an H+ pump in the membranes of parotid granules from chronically stimulated rats. However, despite the presence of H+-pump activity, fluorescence microscopy with the weak base, acridine orange, reveals that the intragranular pH in live cells is greater than that of the cytoplasm.