Development of Small Intestinal Preparation with Intact Epithelial Polarity for Patch-Clamp Study

哺乳动物小肠上皮细胞参与cl的分泌和Na+的吸收以及有机溶质的吸收。一般认为肠道分泌是由隐窝上皮细胞完成的，而吸收是由绒毛上皮细胞完成的。然而，几乎没有直接的证据支持这一假设。为了在单细胞水平上直接检验这一假设，我们开始使用从豚鼠小肠分离的单个绒毛上皮细胞进行实验。然而，单独分离的肠细胞的刷状边界结构会迅速丢失，因此膜片钳技术无法从它们记录葡萄糖依赖的离子电流。本研究旨在建立一种制备具有完整上皮极性的成年豚鼠小肠绒毛和隐窝标本的方法。通过在含edta的生理盐水溶液中振动后分离上皮和切片分别获得该制剂。应用膜片钳技术，可以记录这些离体上皮细胞的离子电流，特别是来自离体绒毛的葡萄糖依赖性离子电流和来自离体隐窝的环amp激活（cftr型）cl通道电流。

The epithelial cells of the mammalian small intestine partakes in both secretion of Cl-and absorption of Na+as well as organic solutes.It is generally thought that the intestinal secretion is carried out by the crypt epithelial cell whereas the absorption is by the villus epithelial cell. However, little direct level evidence has been provided for the hypothesis. To directly examine the hypothesis at the single cell level, we had started experiments using single villus epithelial cells isolated from the guinea pig small intestine.The brush-border structure of singly isolated enterocytes, however, became rapidly lost, and therefore no glucose-dependent ionic currents could be recorded from them by the patch-clamp technique. The present study is aimed at developing the method to prepare the villus and crypt specimen with the intact epithelial polarity from the small intestine of the adult guinea pig. The preparations could be separately obtained by isolating the epithelia after vibration in the EDTA-containing saline solution and by slicing the tissue. By applying the patch-clamp technique, numbers of ionic currents could be recorded from these isolated epithelia, especially glucose-dependent ionic currents from isolated villi and cyclic AMP-activated (CFTR-typed)Cl-channel currents from isolated crypts.