Microtubule-dependent relocation of branchial V-H+-ATPase to the basolateral membrane in the Pacific spiny dogfish (Squalus acanthias): a role in base secretion-猫眼课题宝

太平洋角鲨 (Squalus acanthias) 鳃 V-H -ATP 酶依赖于微管重新定位至基底外侧膜：在碱分泌中的作用

DOI：

发表时间：

2006

期刊：

Journal of Experimental Biology

影响因子：

2.8

通讯作者：

中科院分区：

生物学2区

文献类型：

作者：

M. Tresguerres;S. Parks;F. Katoh;G. Goss

摘要我们之前已经证明，连续静脉输注 NaHCO3 24 小时（∼1000 μmol kg-1 h-1）会导致太平洋角鲨鳃中 V-H+-ATP 酶从细胞质迁移到基底外侧膜。为了进一步研究这种假定的碱基分泌过程，我们进行了类似的实验，添加了秋水仙碱（一种细胞骨架依赖性细胞运输过程的抑制剂）。与单独注入 HCO3- 的鱼相比，经秋水仙碱处理、注入 HCO3- 的鱼的血液 pH 值和血浆总 CO2 显着更高。输注24小时后秋水仙碱的效果最高（8.33±0.06 vs 8.02±0.03 pH单位，15.72±3.29 vs 6.74±1.34 mmol CO2 l-1，N=5）。免疫组织化学和蛋白质印迹证实秋水仙碱阻断 V-H+-ATPase 转运至基底外侧膜。此外，对整个鳃和细胞膜样本的蛋白质印迹分析表明，对碱性胁迫的短期（6小时）反应包括细胞中已存在的V-H+-ATP酶重新定位到基底外侧膜，而长期（24小时）既存在先前存在的酶的重新定位，又存在新单位合成中的上调。我们的结果强烈表明，V-H+-ATP酶的细胞重新定位对于增强太平洋角鲨鳃中HCO3-的分泌是必要的。

SUMMARY We have previously shown that continuous intravenous infusion of NaHCO3 for 24 h (∼1000 μmol kg-1 h-1) results in the relocation of V-H+-ATPase from the cytoplasm to the basolateral membrane in the gills of the Pacific dogfish. To further investigate this putative base-secretive process we performed similar experiments with the addition of colchicine, an inhibitor of cytoskeleton-dependent cellular trafficking processes. Blood pH and plasma total CO2 were significantly higher in the colchicines-treated, HCO3--infused fish compared with fish infused with HCO3- alone. The effect of colchicine was highest after 24 h of infusion (8.33±0.06 vs 8.02±0.03 pH units, 15.72±3.29 vs 6.74±1.34 mmol CO2 l-1, N=5). Immunohistochemistry and western blotting confirmed that colchicine blocked the transit of V-H+-ATPase to the basolateral membrane. Furthermore, western blotting analyses from whole gill and cell membrane samples suggest that the short-term (6 h) response to alkaline stress consists of relocation of V-H+-ATPases already present in the cell to the basolateral membrane, while in the longer term (24 h) there is both relocation of preexistent enzyme and upregulation in the synthesis of new units. Our results strongly suggest that cellular relocation of V-H+-ATPase is necessary for enhanced HCO3- secretion across the gills of the Pacific dogfish.