Wong S M, DeBell M C, Chase H S
Department of Medicine, Columbia University, College of Physicians and Surgeons, New York, New York 10032.
Am J Physiol. 1990 Feb;258(2 Pt 2):F292-6. doi: 10.1152/ajprenal.1990.258.2.F292.
We examined the effect of cell swelling on intracellular free calcium concentration ([Ca]i) in cultured toad bladder cells (TB-M) grown as a polarized monolayer on collagen-coated filters. [Ca]i was measured by use of fura-2, fluorescence microscopy, and simple video imaging. In preliminary experiments we determined that reducing ionic strength by 15% had no effect on the Kd of fura-2, indicating that the dye could be used to examine the effects of cell swelling on [Ca]i. Reducing the osmolality of the serosal bathing medium by 15% caused [Ca]i to increase within 10 s from 97 +/- 9 to 354 +/- 88 nM (n = 5). By 2 min [Ca]i had declined to 163 +/- 22 nM. The increase in [Ca]i was not caused by a fall in Na concentration ([Na]) because isosmotic reduction of serosal [Na] did not result in an increase in [Ca]i. The swelling-induced increase in [Ca]i could be abolished by lowering serosal [Ca] to 200 nM and by addition of lanthanum. The calcium-channel blockers nitrendipine and verapamil also inhibited the swelling-induced increase in [Ca]i, although to different degrees. These experiments demonstrate that swelling of cultured toad bladder cells results in a significant increase in [Ca]i by enhancing the rate of calcium entry across the basolateral membrane, possibly through a calcium channel.
我们研究了细胞肿胀对培养于胶原包被滤膜上的极化单层蟾蜍膀胱细胞（TB-M）细胞内游离钙浓度（[Ca]i）的影响。[Ca]i通过使用fura-2、荧光显微镜和简单的视频成像进行测量。在初步实验中，我们确定将离子强度降低15%对fura-2的解离常数（Kd）没有影响，这表明该染料可用于研究细胞肿胀对[Ca]i的影响。将浆膜侧浴液的渗透压降低15%会导致[Ca]i在10秒内从97±9 nM增加到354±88 nM（n = 5）。到2分钟时，[Ca]i已降至163±22 nM。[Ca]i的增加不是由钠浓度（[Na]）的下降引起的，因为等渗降低浆膜侧[Na]并不会导致[Ca]i增加。通过将浆膜侧[Ca]降至200 nM以及添加镧可以消除肿胀诱导的[Ca]i增加。钙通道阻滞剂尼群地平和维拉帕米也能抑制肿胀诱导的[Ca]i增加，尽管程度不同。这些实验表明，培养的蟾蜍膀胱细胞肿胀通过增强钙跨基底外侧膜的进入速率，可能是通过钙通道，导致[Ca]i显著增加。
