Strange K, Morrison R, Shrode L, Putnam R
Department of Medicine, Children's Hospital, Harvard Medical School, Boston, Massachusetts 02115.
Am J Physiol. 1993 Jul;265(1 Pt 1):C244-56. doi: 10.1152/ajpcell.1993.265.1.C244.
Rat C6 glioma cells chronically acclimated to hypertonic media accumulate large quantities of inositol. When returned to isotonic conditions, the cells swell and lose inositol slowly via a four- to fivefold increase in the rate of passive inositol efflux. The inositol efflux pathway is a Na(+)-independent transport mechanism with low affinity for inositol and is inhibited by quinidine, quinine, various anion transport blockers, and cis-unsaturated fatty acids. Ionomycin-induced elevation of intracellular Ca2+ (Ca2+i) had no effect on basal or swelling-induced inositol efflux. Inositol efflux was not inhibited by chelation of Ca2+i with 1,2-bis(2-aminophenoxy)-ethane-N,N,N',N'-tetraacetic acid. In addition, Ca2+i measured with fura 2 did not change during cell swelling, indicating that increases in Ca2+i do not regulate inositol efflux. Exposure of C6 cells to 20 nM phorbol 12-myristate 13-acetate, 0.5 mM adenosine 3',5'-cyclic monophosphate (cAMP), or 50 microM forskolin had no effect on basal inositol efflux but stimulated swelling-induced inositol loss by 2.6-, 2.2-, and 3.4-fold, respectively. Exposure to the protein kinase inhibitors 1-(5-isoquinolinylsulfonyl)-2-methylpiperazine or staurosporine or downregulation of protein kinase C (PKC) activity, however, had no inhibitory effect on inositol efflux, and cellular cAMP levels were not altered by cell swelling. Taken together, these results indicate that stimulation of PKC and protein kinase A modulates the activity of the efflux pathway but is not required for swelling-induced activation. Ketoconazole, cinnamyl-3,4-dihydroxy-alpha-cyanocinnamate, and gossypol, inhibitors of lipoxygenase enzymes, blocked both basal and swelling-induced inositol efflux, suggesting indirectly that lipoxygenase metabolites may be responsible for swelling-induced activation of the efflux mechanism. The characteristics of inositol efflux in C6 cells are similar to those described for volume regulatory sorbitol and taurine efflux in a number of cell types, suggesting the existence of a common transport mechanism.
长期适应高渗培养基的大鼠C6胶质瘤细胞会积累大量肌醇。当恢复到等渗条件时，细胞会肿胀，并通过被动肌醇外流速率增加4至5倍而缓慢丢失肌醇。肌醇外流途径是一种不依赖Na⁺的转运机制，对肌醇亲和力低，且受奎尼丁、奎宁、各种阴离子转运阻滞剂和顺式不饱和脂肪酸抑制。离子霉素诱导的细胞内Ca²⁺（Ca²⁺i）升高对基础或肿胀诱导的肌醇外流没有影响。用1,2-双（2-氨基苯氧基）乙烷-N,N,N',N'-四乙酸螯合Ca²⁺i不会抑制肌醇外流。此外，用fura 2测量的Ca²⁺i在细胞肿胀过程中没有变化，表明Ca²⁺i的增加不调节肌醇外流。将C6细胞暴露于20 nM佛波醇12-肉豆蔻酸酯13-乙酸酯、0.5 mM腺苷3',5'-环磷酸（cAMP）或50 μM福斯高林对基础肌醇外流没有影响，但分别刺激肿胀诱导的肌醇丢失2.6倍、2.2倍和3.4倍。然而，暴露于蛋白激酶抑制剂1-（5-异喹啉磺酰基）-2-甲基哌嗪或星形孢菌素或下调蛋白激酶C（PKC）活性对肌醇外流没有抑制作用，且细胞肿胀不会改变细胞内cAMP水平。综上所述，这些结果表明PKC和蛋白激酶A的刺激调节了外流途径的活性，但肿胀诱导的激活并不需要它们。酮康唑、肉桂基-3,4-二羟基-α-氰基肉桂酸酯和棉酚是脂氧合酶的抑制剂，它们阻断了基础和肿胀诱导的肌醇外流，间接表明脂氧合酶代谢产物可能是肿胀诱导的外流机制激活的原因。C6细胞中肌醇外流的特征与许多细胞类型中描述的体积调节性山梨醇和牛磺酸外流的特征相似，这表明存在一种共同的转运机制。
