Yanagita T, Wada A, Yamamoto R, Kobayashi H, Yuhi T, Urabe M, Niina H
Department of Pharmacology, Miyazaki Medical College, Japan.
J Neurochem. 1996 Mar;66(3):1249-53. doi: 10.1046/j.1471-4159.1996.66031249.x.
Treatment of cultured bovine adrenal chromaffin cells with 12-O-tetradecanoylphorbol 13-acetate (TPA), an activator of protein kinase C (PKC), decreased [3H]saxitoxin ([3H]STX) binding in a concentration (IC50 = 19 nM)- and time (t1/2 = 4.5 h)-dependent manner. TPA (100 nM for 15 h) lowered the Bmax of [3H]STX binding by 53% without altering the KD value. Phorbol 12,13-dibutyrate (PDBu) also reduced [3H]STX binding, whereas 4 alpha-TPA, an inactive analogue, had no effect. The inhibitory effect of TPA was abolished when H-7 (an inhibitor of PKC), but not H-89 (an inhibitor of cyclic AMP-dependent protein kinase), was included in the culture medium for 1 h before and during TPA treatment. Simultaneous treatment with TPA in combination with either actinomycin D or cycloheximide, an inhibitor of protein synthesis, nullified the effect of TPA. TPA treatment also attenuated veratridine-induced 22Na+ influx but did not alter the affinity of veratridine for Na channels as well as an allosteric potentiation of veratridine-induced 22Na+ influx by brevetoxin. These results suggest that an activation of PKC down-regulates the density of Na channels without altering their pharmacological features; this down-regulation is mediated via the de novo synthesis of an as yet unidentified protein(s), rather than an immediate effect of Na channel phosphorylation.
用蛋白激酶C（PKC）激活剂12 - O - 十四烷酰佛波醇13 - 乙酸酯（TPA）处理培养的牛肾上腺嗜铬细胞，[3H]石房蛤毒素（[3H]STX）结合量以浓度（IC50 = 19 nM）和时间（t1/2 = 4.5小时）依赖性方式降低。TPA（100 nM，处理15小时）使[3H]STX结合的Bmax降低53%，而KD值不变。佛波醇12,13 - 二丁酸酯（PDBu）也降低[3H]STX结合，而无活性类似物4α - TPA则无作用。在TPA处理前及处理期间，将H - 7（PKC抑制剂）而非H - 89（环磷酸腺苷依赖性蛋白激酶抑制剂）加入培养基1小时，可消除TPA的抑制作用。TPA与放线菌素D或蛋白质合成抑制剂环己酰亚胺同时处理，可使TPA的作用无效。TPA处理还减弱了藜芦碱诱导的22Na+内流，但不改变藜芦碱对钠通道的亲和力以及短裸甲藻毒素对藜芦碱诱导的22Na+内流的变构增强作用。这些结果表明，PKC的激活下调钠通道密度而不改变其药理学特性；这种下调是通过一种尚未鉴定的蛋白质的从头合成介导的，而非钠通道磷酸化的直接作用。
