Among the compensatory mechanisms restoring circulating blood volume after severe haemorrhage, increased vasopressin secretion enhances water permeability of distal nephron segments and stimulates Na(+) reabsorption in cortical collecting tubules via epithelial sodium channels (ENaC). The ability of vasopressin to upregulate ENaC via a cAMP-dependent mechanism in the medium to long term is well established. This study addressed the acute regulatory effect of cAMP on human ENaC (hENaC) and thus the potential role of vasopressin in the initial compensatory responses to haemorrhagic shock. The effects of raising intracellular cAMP (using 5 mmol/L isobutylmethylxanthine (IBMX) and 50 μmol/L forskolin) on wild-type and Liddle-mutated hENaC activity expressed in Xenopus oocytes and hENaC localisation in oocyte membranes were evaluated by dual-electrode voltage clamping and immunohistochemistry, respectively. After 30 min, IBMX + forskolin had stimulated amiloride-sensitive Na(+) current by 52% and increased the membrane density of Na(+) channels in oocytes expressing wild-type hENaC. These responses were prevented by 5 μmol/L brefeldin A, which blocks antegrade vesicular transport. By contrast, IBMX + forskolin had no effects in oocytes expressing Liddle-mutated hENaC. cAMP stimulated rapid, exocytotic recruitment of wild-type hENaC into Xenopus oocyte membranes, but had no effect on constitutively over-expressed Liddle-mutated hENaC. Extrapolating these findings to the early cAMP-mediated effect of vasopressin on cortical collecting tubule cells, they suggest that vasopressin rapidly mobilises ENaC to the apical membrane of cortical collecting tubule cells, but does not enhance ENaC activity once inserted into the membrane. We speculate that this stimulatory effect on Na(+) reabsorption (and hence water absorption) may contribute to the early restoration of extracellular fluid volume following severe haemorrhage.

译文

在严重出血后恢复循环血容量的代偿机制中,加压素分泌增加可增强远端肾单位段的水通透性,并通过上皮钠通道 (ENaC) 刺激皮质收集小管中的Na () 重吸收。加压素在中长期内通过cAMP依赖性机制上调ENaC的能力已得到充分确立。这项研究探讨了cAMP对人ENaC (hENaC) 的急性调节作用,从而探讨了加压素在对出血性休克的初始代偿反应中的潜在作用。通过双电极电压钳制和免疫组织化学评估了提高细胞内cAMP (使用5 mmol/L异丁基甲基黄嘌呤 (IBMX) 和50 μ mol/L forskolin) 对非洲爪蟾卵母细胞中表达的野生型和Liddle突变的hENaC活性以及卵母细胞膜中hENaC定位的影响。30分钟后,IBMX forskolin通过52% 刺激了对阿米洛利敏感的Na () 电流,并增加了表达野生型hENaC的卵母细胞中Na () 通道的膜密度。5 μ mol/L brefeldin A阻止了这些反应,从而阻止了顺行的水泡运输。相比之下,IBMX forskolin对表达Liddle突变的hENaC的卵母细胞没有影响。cAMP刺激野生型hENaC迅速,胞外募集到非洲爪蟾卵母细胞膜中,但对组成型过度表达的Liddle突变的hENaC没有影响。将这些发现外推到加压素对皮质收集小管细胞的早期cAMP介导的作用,他们表明加压素可迅速将ENaC动员到皮质收集小管细胞的顶膜上,但一旦插入膜中就不会增强ENaC的活性。我们推测这种对Na () 重吸收 (并因此吸收水) 的刺激作用可能有助于严重出血后细胞外液量的早期恢复。

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