Fluxes of 45Ca2+ were studied in pancreatic islets from non-inbred ob/ob-mice. Because La3+ blocked the transmembrane fluxes of 45Ca2+ in islet cells, incubations aimed at measuring glucose-induced changes of the intracellular Ca2+ were ended by washing the islets with 2 mM-La3+ for 60 min. 2. Uptake of 45Ca2+ progressed for 2 hr; the intracellular concentration of exchangable Ca2+ was about 7 m-mole/kg dry wt., as estimated from the isotope distribution at apparent equilibrium in islets exposed to 3 mM D-glucose. Raising the D-glucose concentration to 20 mM enhanced the 45 Ca2+ uptake whether or not the islets had first been equilibrated with the isotope. The stimulatory effect of D-glucose was observed in Tris buffer containing no anions but Cl- as well as in polyanionic bicarbonate buffer. The effect could not be reproduced with equimolar L-glucose. 3. The rate of 45Ca2+ release was the same whether the islets had been pre-loaded in the presence of 3 or 20 mM D-glucose. Thus the 45Ca2+ that had been taken up in response to 20 mM D-glucose appeared to be released much more slowly than the bulk of intracellular 45Ca2+. The release of 45Ca2+ was not significantly influenced by D-glucose during the release period. Incubation for 30 min was require for half of the radioactivity to be released. 4. The rates of insulin secretion were about the same in uni-anionic Tris buffer as in polyanionic bicarbonate buffer. A marked insulin secretory response to 20 mM D-glucose was observed in either buffer. 5. It is concluded that 20 mM D-glucose causes a net uptake of Ca2+ from the extracellular fluid into the interior of the beta-cells. This uptake is probably not regulated at the level of the plasma membrane but more likely reflects an increased affinity of some intracellular phase or compartment for the ion. Because the observed uptake and release of intracellular 45Ca2+ are slow processes in comparison with the rapid effects of extracellular Ca2+ on insulin secretion, insulin secretion may also depend on a more superficial and La3+-displacable Ca2+ pool.
摘要
对非近交系ob/ob小鼠的胰岛中45Ca2+的通量进行了研究。由于La3+可阻断胰岛细胞中45Ca2+的跨膜通量，因此，旨在测量葡萄糖诱导的细胞内Ca2+变化的孵育实验，通过用2 mM-La3+洗涤胰岛60分钟来终止。2. 45Ca2+的摄取持续了2小时；根据暴露于3 mM D-葡萄糖的胰岛在表观平衡时的同位素分布估算，可交换Ca2+的细胞内浓度约为7 mmol/kg干重。无论胰岛是否首先用该同位素进行平衡，将D-葡萄糖浓度提高到20 mM均可增强45Ca2+的摄取。在不含除Cl-以外阴离子的Tris缓冲液以及聚阴离子碳酸氢盐缓冲液中均观察到D-葡萄糖的刺激作用。等摩尔的L-葡萄糖无法重现该效应。3. 无论胰岛是在3 mM还是20 mM D-葡萄糖存在下进行预加载，45Ca2+的释放速率均相同。因此，响应20 mM D-葡萄糖而摄取的45Ca2+的释放似乎比大部分细胞内45Ca2+的释放要慢得多。在释放期间，45Ca2+的释放不受D-葡萄糖的显著影响。放射性的一半释放需要孵育30分钟。4. 在单阴离子Tris缓冲液中的胰岛素分泌速率与在聚阴离子碳酸氢盐缓冲液中的大致相同。在两种缓冲液中均观察到对20 mM D-葡萄糖的明显胰岛素分泌反应。5. 得出的结论是，20 mM D-葡萄糖会导致Ca2+从细胞外液净摄取到β细胞内部。这种摄取可能不是在质膜水平上进行调节的，而更可能反映了某些细胞内相或区室对该离子的亲和力增加。由于观察到的细胞内45Ca2+的摄取和释放与细胞外Ca2+对胰岛素分泌的快速作用相比是缓慢的过程，因此胰岛素分泌可能还取决于一个更浅表且可被La3+置换的Ca2+池。
