Lester D S, Asher C, Garty H
Department of Membrane Research, Weizmann Institute of Science, Rehovot, Israel.
Am J Physiol. 1988 Jun;254(6 Pt 1):C802-8. doi: 10.1152/ajpcell.1988.254.6.C802.
Incubating toad bladder with 10 mU/ml vasopressin increases the amiloride-blockable Na+ flux in membrane vesicles derived from the epithelial cells by about twofold. This stimulation is further enhanced by 3-isobutyl-1-methylxanthine and can be mimicked by 8-bromoadenosine 3', 5'-cyclic monophosphate. Thus the natriferic action of cAMP involves a sustained change of the apical membrane preserved by the isolated vesicles. The possibility that transport is modulated by direct phosphorylation/dephosphorylation of the Na+ channel was tested. Trapping purified cAMP-dependent protein kinase, cAMP, and ATP in apical vesicles failed to alter Na+ transport even though the enzyme proved active and could phosphorylate intravesicular proteins. Trapping several phosphatases partially purified from toad bladder in vesicles was ineffective as well. These data suggest that the cAMP-induced increase in Na+ conductance involves processes other than phosphorylation of the channel protein or direct channel-cAMP interaction.
用10 mU/ml抗利尿激素孵育蟾蜍膀胱，可使来源于上皮细胞的膜囊泡中氨氯吡咪可阻断的Na⁺通量增加约两倍。3-异丁基-1-甲基黄嘌呤可进一步增强这种刺激作用，且8-溴腺苷3',5'-环磷酸单酯可模拟该作用。因此，环磷酸腺苷（cAMP）的排钠作用涉及分离出的囊泡所保留的顶端膜的持续变化。对Na⁺通道通过直接磷酸化/去磷酸化调节转运的可能性进行了测试。将纯化的依赖cAMP的蛋白激酶、cAMP和ATP捕获到顶端囊泡中，尽管该酶被证明有活性且能使囊泡内蛋白质磷酸化，但未能改变Na⁺转运。将从蟾蜍膀胱中部分纯化的几种磷酸酶捕获到囊泡中也无效。这些数据表明，cAMP诱导的Na⁺电导增加涉及通道蛋白磷酸化或通道与cAMP直接相互作用以外的过程。
