Valencia L, Bidet M, Martial S, Sanchez E, Melendez E, Tauc M, Poujeol C, Martin D, Namorado M D, Reyes J L, Poujeol P
Departamento de Fisiología, Centro de Investigación y de Estudios Avanzados del Institúto Politécnico Nacional, Mexico City DF 07000, Mexico.
Am J Physiol Cell Physiol. 2001 May;280(5):C1193-203. doi: 10.1152/ajpcell.2001.280.5.C1193.
To characterize Ca(2+) transport in newborn rat cortical collecting duct (CCD) cells, we used nifedipine, which in adult rat distal tubules inhibits the intracellular Ca(2+) concentration (Ca(2+)) increase in response to hormonal activation. We found that the dihydropyridine (DHP) nifedipine (20 microM) produced an increase in Ca(2+) from 87.6 +/- 3.3 nM to 389.9 +/- 29.0 nM in 65% of the cells. Similar effects of other DHP (BAY K 8644, isradipine) were also observed. Conversely, DHPs did not induce any increase in Ca(2+) in cells obtained from proximal convoluted tubule. In CCD cells, neither verapamil nor diltiazem induced any rise in Ca(2+). Experiments in the presence of EGTA showed that external Ca(2+) was required for the nifedipine effect, while lanthanum (20 microM), gadolinium (100 microM), and diltiazem (20 microM) inhibited the effect. Experiments done in the presence of valinomycin resulted in the same nifedipine effect, showing that K(+) channels were not involved in the nifedipine-induced Ca(2+) rise. H(2)O(2) also triggered Ca(2+) rise. However, nifedipine-induced Ca(2+) increase was not affected by protamine. In conclusion, the present results indicate that 1) primary cultures of cells from terminal nephron of newborn rats are a useful tool for investigating Ca(2+) transport mechanisms during growth, and 2) newborn rat CCD cells in primary culture exhibit a new apical nifedipine-activated Ca(2+) channel of capacitive type (either transient receptor potential or leak channel).
为了表征新生大鼠皮质集合管（CCD）细胞中的钙（Ca²⁺）转运，我们使用了硝苯地平，在成年大鼠远曲小管中，它可抑制因激素激活而导致的细胞内钙浓度（[Ca²⁺]i）升高。我们发现，二氢吡啶（DHP）硝苯地平（20 μM）可使65%的细胞中[Ca²⁺]i从87.6±3.3 nM升高至389.9±29.0 nM。还观察到其他DHP（BAY K 8644、伊拉地平）有类似作用。相反，DHP对来自近端曲管的细胞中的[Ca²⁺]i没有诱导任何升高。在CCD细胞中，维拉帕米和地尔硫䓬均未诱导[Ca²⁺]i升高。在存在乙二醇双（2-氨基乙基醚）四乙酸（EGTA）的情况下进行的实验表明，硝苯地平发挥作用需要细胞外钙，而镧（20 μM）、钆（100 μM）和地尔硫䓬（20 μM）可抑制该作用。在存在缬氨霉素的情况下进行的实验产生了相同的硝苯地平效应，表明钾（K⁺）通道不参与硝苯地平诱导的[Ca²⁺]i升高。过氧化氢（H₂O₂）也可引发[Ca²⁺]i升高。然而，硝苯地平诱导的[Ca²⁺]i升高不受鱼精蛋白影响。总之，目前的结果表明：1）新生大鼠终末肾单位细胞的原代培养是研究生长过程中Ca²⁺转运机制的有用工具；2）原代培养的新生大鼠CCD细胞表现出一种新的顶端硝苯地平激活的电容性钙通道（瞬时受体电位通道或渗漏通道）。
