Lalik P H, Krafte D S, Volberg W A, Ciccarelli R B
Department of Molecular Biology, Sterling Winthrop Pharmaceuticals Research Division, Rensselaer, New York 12144.
Am J Physiol. 1993 Apr;264(4 Pt 1):C803-9. doi: 10.1152/ajpcell.1993.264.4.C803.
Chinese hamster ovary (CHO-K1) cells were observed to display transient inward Na+ currents of average amplitude (-92 +/- 20 pA), which activated at voltages more than -40 mV, and peak inward currents were observed at potentials equal to or more than +10 mV. Inward Na+ currents in these cells were eliminated after treatment with 500 or 50 nM tetrodotoxin (TTX), whereas 5 nM TTX resulted in 64 +/- 10% inhibition of Na+ current. Using DNA primers designed to bind to the rat brain IIA Na+ channel subtype, we amplified specific polymerase chain reaction (PCR) fragments from CHO-K1 poly-(A)+RNA. The cloning and sequencing of two of these fragments confirmed the presence of an endogenously expressed Na+ channel gene in these cells, which we have termed cho 1. Comparison of the DNA sequence of cho 1 PCR fragments with other known Na+ channel genes indicated a high degree of homology with rat brain Na+ channel subtypes. Northern blots using riboprobes generated from the cho 1 PCR fragments revealed the presence of a specific 9-kb mRNA in these cells. The molecular and electrophysiological data suggest that the cho 1 Na+ channel gene from CHO-K1 cells is closely related to brain-type Na+ channels.
观察到中国仓鼠卵巢（CHO-K1）细胞表现出平均幅度为（-92±20 pA）的瞬时内向Na⁺电流，该电流在电压高于-40 mV时激活，在等于或高于+10 mV的电位时观察到内向电流峰值。用500或50 nM河豚毒素（TTX）处理后，这些细胞中的内向Na⁺电流被消除，而5 nM TTX导致Na⁺电流抑制64±10%。使用设计用于与大鼠脑IIA Na⁺通道亚型结合的DNA引物，我们从CHO-K1聚腺苷酸（poly-(A)⁺）RNA中扩增出特异性聚合酶链反应（PCR）片段。对其中两个片段的克隆和测序证实了这些细胞中存在内源性表达的Na⁺通道基因，我们将其命名为cho 1。将cho 1 PCR片段的DNA序列与其他已知的Na⁺通道基因进行比较，表明与大鼠脑Na⁺通道亚型具有高度同源性。使用从cho 1 PCR片段产生的核糖探针进行的Northern印迹显示这些细胞中存在一种特异性的9 kb mRNA。分子和电生理数据表明，来自CHO-K1细胞的cho 1 Na⁺通道基因与脑型Na⁺通道密切相关。
