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. 2004 Mar 15;164(6):911–921. doi: 10.1083/jcb.200308103

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Copyright © 2004, The Rockefeller University Press

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Figure 7. — Alterations in K8-null ion transporter dynamics. (A) Immunoblot analysis of K8+/+, +/−, and −/− distal colon. Total homogenates of distal colon were loaded in equal amounts (based on protein assay; further verified by blotting using anti-tubulin Ab, not depicted) and then analyzed by blotting using antibodies to the indicated proteins. For each genotype, homogenates from two independent mice are shown. (B) Immunofluorescence staining of AE1/2 and ENaCγ in colon of K8+/+ and −/− mice. Distal colon AE1/2 (red, a–d), F-actin (green, a and b; not depicted in c and d in order to highlight the AE1/2 red staining), ENaCγ (red, e–h), and nuclei (blue, a–h) were stained. Panels c, d, g, and h show higher magnifications of the base of the tubular crypts of the corresponding a, b, d, and f panels. L, lumen; M, muscle layer. Asterisks in f highlight the apical expression of ENaCγ. Bars: (a and b) 50 μm; (c and d) 10 μm; (e and f) 50 μm; (g and h) 10 μm. (C) Surface cell pH and Cl/HCO₃ exchange activity in K8+/+ and −/− colon. Distal colon from K8+/+ and −/− mice was loaded with the pH_i-sensitive dye BCECF-AM, and individual surface epithelial cells were imaged with a low-light CCD camera. Representative pH_i (vs. time) of surface cells in K8+/+ and −/− colon during transient removal of extracellular Cl (0-Cl⁻) are shown. Bar graphs show estimated Cl/HCO₃ exchange activity based on the average alkalinization observed over 5 min after Cl removal, expressed as mean ± SEM. The number of cells counted/experiments are 170/14 (K8+/+) and 120/10 (K8−/−).