School of Life Sciences, Sun Yat-sen University, Guangzhou, China.
PLoS One. 2011;6(8):e22283. doi: 10.1371/journal.pone.0022283. Epub 2011 Aug 22.
The epithelium lining the epididymis provides an optimal acidic fluid microenvironment in the epididymal tract that enable spermatozoa to complete the maturation process. The present study aims to investigate the functional role of Na(+)/HCO(3)(-) cotransporter in the pH regulation in rat epididymis.
METHOD/PRINCIPAL FINDINGS: Immunofluorescence staining of pan cytokeratin in the primary culture of rat caput epididymal epithelium showed that the system was a suitable model for investigating the function of epididymal epithelium. Intracellular and apical pH were measured using the fluorescent pH sensitive probe carboxy-seminaphthorhodafluor-4F acetoxymethyl ester (SNARF-4F) and sparklet pH electrode respectively to explore the functional role of rat epididymal epithelium. In the HEPES buffered Krebs-Henseleit (KH) solution, the intracellular pH (pHi) recovery from NH(4)Cl induced acidification in the cultured caput epididymal epithelium was completely inhibited by amiloride, the inhibitor of Na(+)/H(+) exchanger (NHE). Immediately changing of the KH solution from HEPES buffered to HCO(3)(-) buffered would cause another pHi recovery. The pHi recovery in HCO(3)(-) buffered KH solution was inhibited by 4, 4diisothiocyanatostilbene-2,2-disulfonic acid (DIDS), the inhibitor of HCO(3)(-) transporter or by removal of extracellular Na(+). The extracellular pH measurement showed that the apical pH would increase when adding DIDS to the apical side of epididymal epithelial monolayer, however adding DIDS to the basolateral side had no effect on apical pH.
The present study shows that sodium coupled bicarbonate influx regulates intracellular and apical pH in cultured caput epididymal epithelium.
附睾管中的上皮为精子提供了一个优化的酸性液体微环境，使精子能够完成成熟过程。本研究旨在探讨钠/碳酸氢盐共转运体在调节大鼠附睾 pH 值中的功能作用。
方法/主要发现：对大鼠头段附睾上皮的原代培养进行泛细胞角蛋白免疫荧光染色，结果表明该体系是研究附睾上皮功能的合适模型。使用荧光 pH 敏感探针羧基-半花青染料-4F 乙脂（SNARF-4F）和火花 pH 电极分别测量细胞内和顶端 pH 值，以探讨大鼠附睾上皮的功能作用。在 HEPES 缓冲的 Krebs-Henseleit（KH）溶液中，用氨氯吡咪诱导培养的头段附睾上皮酸化，细胞内 pH（pHi）的恢复完全被 NHE 的抑制剂阿米洛利所抑制。立即将 KH 溶液从 HEPES 缓冲液转换为 HCO3-缓冲液会引起另一次 pHi 恢复。在 HCO3-缓冲 KH 溶液中的 pHi 恢复被 HCO3-转运体的抑制剂 4,4-二异硫氰酸基-2,2-二磺酸（DIDS）或去除细胞外 Na+所抑制。细胞外 pH 值测量显示，当将 DIDS 添加到附睾上皮单层的顶端侧时，顶端 pH 值会升高，而将 DIDS 添加到底侧侧对顶端 pH 值没有影响。
本研究表明，钠偶联碳酸氢盐内流调节培养的头段附睾上皮的细胞内和顶端 pH 值。
