Chloride Transport Across Rat Ileal Basolateral Membrane Vesicles

Marlene Daher, Sari Acra, Wendy Dykes, Fayez K. Ghishan

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Abstract

The present study was designed to investigate CI” transport across rat ileal basolateral membranes. Basolateral membrane vesicles were prepared by a well-validated technique. The purity of the basolateral membrane vesicles was verified by marker enzyme studies and by studies of d-glucose and calcium uptake. Cl- uptake was studied by a rapid filtration technique. Neither an outwardly directed pH gradient, nor a HCO3 gradient, or their combination could elicit any stimulation of Cl transport when compared with no gradient. 4,4-Diisothiocyanostilbene-2,2-disulfonic acid at 5 mM concentration did not inhibit Cl uptake under gradient condition. Similarly, the presence of the combination of outwardly directed Na+ and HCO3 gradients did not stimulate Cl uptake compared with the combination of K+ and HCO3 gradients or no HCO3 gradient. This is in contrast to our results in the brush border membranes, where an outwardly directed pH gradient caused an increase in Cl uptake. Cl uptake was stimulated in the presence of combined Na+ and K+ gradient. Bumetanide at 0.1 mM concentration inhibited the initial rate of Cl uptake in the presence of combined Na+ and K+ gradients. Kinetic studies of bumetanide-sensitive Cl uptake showed a Vmax of 5.6 × 0.7 nmol/mg protein/5 sec and a Km of 30 × 8.7 mM. Cl uptake was stimulated by an inside positive membrane potential induced by the ionophore valinomycin in the setting of inwardly directed K+ gradient compared with voltage clamp condition. These studies demonstrate two processes for Cl transport across the rat ileal basolateral membrane: one is driven by an electrogenic diffusive process and the second is a bumetanide-sensitive Na+/K+/2 Cl process. Cl uptake is not enhanced by pH gradient, HCO3 gradient, their combination, or outwardly directed HCO3 and Na+ gradients.

Original languageEnglish (US)
Pages (from-to)254-260
Number of pages7
JournalProceedings of the Society for Experimental Biology and Medicine
Volume201
Issue number3
DOIs
StatePublished - Dec 1992
Externally publishedYes

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ASJC Scopus subject areas

  • Biochemistry, Genetics and Molecular Biology(all)

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