Transmembrane exchange of chloride with bicarbonate ion in mammalian red blood cells: evidence for a sulphonamide‐sensitive "carrier".

Abstract

SUMMARY1. It is well known that red blood cells suspended in isotonic NH4C1 solution swell because penetration of NH3 induces a transmembrane exchange between Cl-0 and OH-, (or HCO3-j). The rate of swelling thus depends on the speed of the transmembrane exchanges and on the amount of anions available for exchange.2. It has been demonstrated in experiments carried out in a C02-free medium that OH-ions are poorly permeating whereas the permeability for HCO3-is very high. Thus the rate of swelling is largely dependent on the intracellular HCO3-concentration. In this context the well-known inhibitory effect of sulphonamides upon swelling can be interpreted, and always has been until now, as being due to the inhibitory action of the drug on the intracellular carbonic anhydrase. However, this inhibitory effect could also result from a direct action of the drug on the transmembrane exchange; it would explain why under conditions of total carbonic anhydrase inhibition we have shown that the inhibition of swelling is far from maximal.3. A direct experimental evidence of such an effect of carbonic anhydrase inhibitors on the transmembrane exchange of Chwith HCO3-was obtained with benzolamide (Cl 11,366), Cl 13,580 and ethoxzolamide. Surprisingly enough, however, acetazolamide (Diamox) does not affect the transmembrane exchange process.4. The inhibitory effect of sulphonamides on HCO3-transport process is discussed in terms of an interaction of the drug with a transport system common to HCO3-and organic anions.

A Transport Metabolon

“…AE1 bicarbonate efflux activity was maximally inhibited by 70 Ϯ 2% at an acetazolamide concentration of 250 M. The apparent K i was 54 M, which is higher than the 10 Ϫ8 M value measured using purified CAII (20). The concentration of carbonic anhydrase inhibitors required to have an effect on anion exchange in erythrocytes were also found to be several orders of magnitude higher than required to inhibit CAII (24,25). In these studies, CAII did not become rate-limiting for anion exchange until the enzyme was inhibited to greater than 99%.…”

“…Effect of CAII Inhibition on Cl Ϫ /HCO 3 Ϫ Exchange Activity-To determine the role of CAII in facilitating AE1 transport activity, we compared the Cl Ϫ /HCO 3 Ϫ exchange activity of cells transiently transfected with AE1 cDNA before and after incubation with acetazolamide, a sulfonamide that inhibits CA enzymatic activity without direct effect on anion exchange (25,48). To measure AE1 transport activity, HEK293 cells grown on coverslips were transiently transfected with AE1 cDNA, loaded with BCECF-AM, a pH-sensitive fluorescent dye, and mounted in a fluorescence cuvette.…”

Studies on the relationship between rat renal medullary cell volume and external anion concentration in hyperosmolal media.

“…Acetazolamide does not affect Cl-HCO3 exchange across red cell membranes (Cousin, Motais & Sola, 1975) nor, uniquely among a wide range of carbonic anhydrase inhibitors studied, does it impair Cl-Cl self-exchange (Cousin & Motais, 1976). If these findings are applicable also to renal medullary cells, the effects of acetazolamide upon cell volume (Fig.…”

The role of carbonic anhydrase inhibitors on anion permeability into ox red blood cells.

“…It has recently been shown that benzolamide (Cl 11,366) and Cl 13,580 inhibit the transmembrane Cl-/HCO3-exchanges by acting directly at membrane level Cousin et al 1975). The present results show that the self-exchanges of organic anions (AO-/Ai-) or chloride (Cl-/Cl1-), in which carbonic anhydrase could not be involved, are also inhibited by sulphonamides.…”