Determination of mitochondrial calcium content in hepatocytes by a rapid cellular-fractionation technique. Alpha-adrenergic agonists do not mobilize mitochondrial Ca2+.
Determination of mitochondrial calcium content in hepatocytes by a rapid cellular-fractionation technique. Alpha-adrenergic agonists do not mobilize mitochondrial Ca2+.
复制标题
通过快速细胞分级技术测定肝细胞中线粒体钙含量。
作者：
S. Shears;C. Kirk
一种快速的细胞分离技术[前文，Shears&Kirk(1984)Biochem.，219,375-382]被用来在几秒钟内从肝细胞中分离出富含线粒体的部分。线粒体钙估计不超过细胞总钙的41%。至少一半的线粒体钙存在于一个能量依赖的池中；总细胞钙的20%在10s内可被EGTA获得。α-肾上腺素能激动剂苯肾上腺素在0.5min内刺激糖原磷酸化酶活性100%，10min后导致EGTA不能进入的池中总细胞钙的20%损失。然而，在加入苯肾上腺素后的0.5到10分钟内，线粒体富钙部分的钙含量没有显著变化。在0-4℃下孵育90分钟的肝细胞在37℃孵育20分钟时排出这种阳离子，在此之后，苯肾上腺素不能改变线粒体富含钙的部分的钙含量。结论：α-肾上腺素能激动剂不能动员肝细胞线粒体中的钙离子。
A rapid cellular fractionation technique [the preceding paper, Shears & Kirk (1984) Biochem. J., 219, 375-382] was employed to separate a mitochondria-rich fraction from hepatocytes within seconds. Mitochondrial Ca was estimated to be no more than 41% of total cell Ca. At least half of the mitochondrial Ca was present in an energy-dependent pool; 20% of total cell Ca was accessible to EGTA within 10s. The alpha-adrenergic agonist phenylephrine stimulated glycogen phosphorylase activity by 100% within 0.5 min and induced a loss of 20% of total cell Ca after 10 min from the EGTA-inaccessible pool. However, between 0.5 and 10 min after the addition of phenylephrine to hepatocytes there was no significant change in the Ca content of the mitochondria-rich fraction. Hepatocytes that were preloaded with Ca2+ during 90 min incubation at 0-4 degrees C expelled this cation during 20 min incubation at 37 degrees C. After this time, phenylephrine failed to alter the Ca content of a mitochondria-rich fraction. It is concluded that alpha-adrenergic agonists do not mobilize Ca2+ from hepatocyte mitochondria.