Chase H S, Wong S M
Department of Medicine, College of Physicians and Surgeons, Columbia University, New York, New York 10032.
Am J Physiol. 1988 Mar;254(3 Pt 2):F374-84. doi: 10.1152/ajprenal.1988.254.3.F374.
We examined the relationship between cyclic AMP and the intracellular free calcium concentration ([Ca]i) in MDCK cells, a hormonally responsive and chloride-secreting cell line. We measured [Ca]i using the calcium-sensitive dye fura-2, fluorescence microscopy, and a silicon intensifier target camera to amplify the signal. Isoproterenol, known to stimulate chloride transport via cyclic AMP, increased [Ca]i from 112 +/- 17 to 373 +/- 53 nM. The rise appeared due to an increase in cyclic AMP: isobutylmethylxanthine enhanced the effect of a submaximal dose of isoproterenol on [Ca]i, the cyclic AMP analogue 8-bromo-cyclic AMP caused [Ca]i to increase from 100 +/- 8 to 278 +/- 40 nM, and direct activation of adenylate cyclase with forskolin increased [Ca]i from 192 +/- 29 to 279 +/- 35 nM. The rise in [Ca]i after cyclic AMP may be due to an influx of calcium from the outside: the cyclic AMP-induced increase in [Ca]i was prevented either by lowering extracellular [Ca] or by addition of 1 mM lanthanum. The mechanism by which calcium enters the cell may not be a calcium channel because neither verapamil nor nitrendipine prevented cyclic AMP from increasing [Ca]i. Cyclic AMP also does not appear to act directly on sodium-calcium exchange. [Ca]i increased as well in low [Na] as in high [Na] following addition of the nucleotide. Thus, isoproterenol, acting through an increase in [cyclic AMP] causes an increase in [Ca]i in MDCK cells. The source and route of entry of calcium into the cytoplasm remain uncertain.
我们研究了环磷酸腺苷（cAMP）与MDCK细胞（一种对激素有反应且能分泌氯离子的细胞系）细胞内游离钙浓度（[Ca]i）之间的关系。我们使用钙敏染料fura-2、荧光显微镜和硅增强靶相机来放大信号，从而测量[Ca]i。已知异丙肾上腺素通过cAMP刺激氯离子转运，它使[Ca]i从112±17 nM增加到373±53 nM。这种升高似乎是由于cAMP增加所致：异丁基甲基黄嘌呤增强了亚最大剂量异丙肾上腺素对[Ca]i的作用，环磷酸腺苷类似物8-溴环磷酸腺苷使[Ca]i从100±8 nM增加到278±40 nM，用福斯可林直接激活腺苷酸环化酶使[Ca]i从192±29 nM增加到279±35 nM。cAMP升高后[Ca]i的增加可能是由于钙从细胞外流入：通过降低细胞外[Ca]或添加1 mM镧可阻止cAMP诱导的[Ca]i增加。钙进入细胞的机制可能不是通过钙通道，因为维拉帕米和尼群地平都不能阻止cAMP增加[Ca]i。cAMP似乎也不直接作用于钠钙交换。添加核苷酸后，在低[Na]和高[Na]条件下[Ca]i均增加。因此，异丙肾上腺素通过增加[cAMP]导致MDCK细胞中[Ca]i增加。钙进入细胞质的来源和途径仍不确定。
