Damkier Helle Hasager, Prasad Vikram, Hübner Christian Andreas, Praetorius Jeppe
Dept. of Anatomy, Aarhus University, DK-8000 Aarhus C, Denmark.
Am J Physiol Cell Physiol. 2009 Jun;296(6):C1291-300. doi: 10.1152/ajpcell.00062.2009. Epub 2009 Apr 15.
The choroid plexus epithelium (CPE) secretes the major fraction of the cerebrospinal fluid (CSF). The Na(+)-HCO(3)(-) transporter Ncbe/Nbcn2 in the basolateral membrane of CPE cells is important for Na(+)-dependent pH(i) increases and probably for CSF secretion. In the current study, the anion transport inhibitor DIDS had no effect on the residual pH(i) recovery in acidified CPE from Ncbe/Nbcn2 knockout mouse by 2',7'-bis(2-carboxyethyl)-5(6)-carboxyfluorescein (BCECF)-fluorescence microscopy in the presence of CO(2)/HCO(3)(-) (Ncbe/Nbcn2-ko+DIDS 109% of control, P = 0.76, n = 5). Thus Ncbe/Nbcn2 mediates the DIDS-sensitive Na(+)-dependent pH(i) recovery in the CPE. The Na(+)/H(+) exchanger-1 Nhe1 is proposed to mediate similar functions as Ncbe/Nbcn2 in CPE. Here, we immunolocalize the Nhe1 protein to the luminal membrane domain in mouse and human CPE. The Na(+)-dependent pH(i) recovery of Nhe1 wild-type (Nhe1-wt) mice in the absence of CO(2)/HCO(3)(-) was abolished in the Nhe1 knockout CPE (Nhe1-ko 0.37% of Nhe1-wt, P = 0.0007, n = 5). In Ncbe/Nbcn2-ko mice, Nhe1 was targeted to the basolateral membrane. Nevertheless, the luminal Na(+)-dependent pH(i) recovery was increased in Ncbe/Nbcn2-ko compared with wild-type littermates (Nhe1-ko 146% of Nhe1-wt, P = 0.007, n = 5). Whereas the luminal Nhe activity was inhibited by the Nhe blocker EIPA (10 microM) in the Ncbe/Nbcn2-wt, it was insensitive to the inhibitor in Ncbe/Nbcn2-ko (Ncbe/Nbcn2-ko+EIPA 100% of control, P = 0.98, n = 5). This indicates that a luminal EIPA-insensitive Nhe was induced in Ncbe/Nbcn2-ko CPE and that EIPA-sensitive Nhe activity was basolateral. The Nhe1 translocation in Ncbe/Nbcn2-ko CPE may reflect a compensatory response, which provides the cells with better means of regulating pH(i) or transporting Na(+) after Ncbe/Nbcn2 disruption.
脉络丛上皮（CPE）分泌脑脊液（CSF）的主要成分。CPE细胞基底外侧膜中的Na(+)-HCO(3)(-)转运体Ncbe/Nbcn2对于Na(+)依赖的细胞内pH值（pH(i)）升高很重要，可能也对脑脊液分泌很重要。在本研究中，通过2',7'-双（2-羧乙基）-5（6）-羧基荧光素（BCECF）荧光显微镜观察，在存在CO(2)/HCO(3)(-)的情况下，阴离子转运抑制剂DIDS对来自Ncbe/Nbcn2基因敲除小鼠的酸化CPE中残余的pH(i)恢复没有影响（Ncbe/Nbcn2基因敲除+DIDS为对照的109%，P = 0.76，n = 5）。因此，Ncbe/Nbcn2介导了CPE中对DIDS敏感的Na(+)依赖的pH(i)恢复。有人提出Na(+)/H(+)交换体-1 Nhe1在CPE中发挥与Ncbe/Nbcn2类似的功能。在此，我们通过免疫定位将Nhe1蛋白定位于小鼠和人CPE的腔面膜结构域。在没有CO(2)/HCO(3)(-)的情况下，Nhe1野生型（Nhe1-wt）小鼠的Na(+)依赖的pH(i)恢复在Nhe1基因敲除的CPE中消失（Nhe1基因敲除为Nhe1-wt的0.37%，P = 0.0007，n = 5）。在Ncbe/Nbcn2基因敲除小鼠中，Nhe1定位于基底外侧膜。然而，与野生型同窝小鼠相比，Ncbe/Nbcn2基因敲除小鼠的腔内Na(+)依赖的pH(i)恢复增加（Nhe1基因敲除为Nhe1-wt的146%，P = 0.007，n = 5）。在Ncbe/Nbcn2野生型中，腔内Nhe活性被Nhe阻滞剂EIPA（10 microM）抑制，而在Ncbe/Nbcn2基因敲除小鼠中对该抑制剂不敏感（Ncbe/Nbcn2基因敲除+EIPA为对照的100%，P = 0.98，n = 5）。这表明在Ncbe/Nbcn2基因敲除的CPE中诱导了一种对EIPA不敏感的腔内Nhe，且对EIPA敏感的Nhe活性位于基底外侧。Ncbe/Nbcn2基因敲除的CPE中Nhe1的易位可能反映了一种代偿反应，在Ncbe/Nbcn2破坏后为细胞提供了更好的调节pH(i)或转运Na(+)的方式。
