Digitonin-permeabilized colonic cell layers: Demonstration of Calcium-activated Basolateral K+ and Cl Conductances

Dean Chang, David C. Dawson

Research output: Contribution to journalArticle

19 Scopus citations

Abstract

Sheets of isolated turtle colon were exposed to digitonin on the mucosal side to chemically remove the apical membrane as a permeability barrier. Increases in the mucosal uptake of 86Rb, [3H]mannitol, and 45Ca-EGTA, and the appearance of the cytosolic marker enzyme lactate dehydrogenase in the mucosal bath confirmed the permeabilizing effect of the detergent. Basolateral K+ and Cl currents were generated by imposing transmural ion gradients, and cytosolic free Ca2+ was manipulated by means of a Ca2+ buffer system in the mucosal bathing solution. Raising the cytosolic free Ca2+ concentration from the nanomolar to the micromolar range activated basolateral conductances for K+ and Cl. Differences in ion selectivity, blocker specificity, calcium activation kinetics, and divalent cation activation selectivity indicated that the Ca2+ increases in the K+ and Cl conductances were due to separate populations of channels. The results are consistent with the notion that the apical membranes of turtle colon epithelial cells can be functionally removed under conditions that preserve some of the conductive properties of the basolateral membrane, specifically Ca2+-activated conductive pathways for K+ and Cl. This permeabilized preparation should offer a means for the identification of macroscopic currents that are due to presumed Ca2+ channels, and may also provide a model system for the functional reconstitution of channel regulatory mechanisms.

Original languageEnglish (US)
Pages (from-to)281-306
Number of pages26
JournalJournal of General Physiology
Volume92
Issue number3
DOIs
StatePublished - Sep 1 1988

ASJC Scopus subject areas

  • Physiology

Fingerprint Dive into the research topics of 'Digitonin-permeabilized colonic cell layers: Demonstration of Calcium-activated Basolateral K<sup>+</sup> and Cl<sup>−</sup> Conductances'. Together they form a unique fingerprint.

  • Cite this