Two pathways for choline transport in eel erythrocytes: a saturable carrier and a volume-activated channel

1994 ◽  
Vol 267 (3) ◽  
pp. R773-R779 ◽  
Author(s):  
S. E. Joyner ◽  
K. Kirk
Keyword(s):  
Red Blood Cells ◽  
Blood Cells ◽  
Anguilla Anguilla ◽  
Anion Transport ◽  
Response Curves ◽  
Choline Transport ◽  
Human Red Blood Cells ◽  
Selective Channel ◽  
Transport Inhibitors ◽  
In Cells

Choline transport in eel (Anguilla anguilla) erythrocytes was investigated in cells suspended in isotonic and hypotonic media. In cells in isosmotic solution choline transport was mediated by a saturable system with a Michaelis constant (Km; 62 +/- 6 microM) similar to that of the choline carrier of human erythrocytes but a maximal transport rate (Vmax; 4.5 +/- 0.4 mmol.1 red blood cells-1.h-1) almost two orders of magnitude higher than that in human red blood cells. This pathway was inhibited by hemicholinium-3 and dodecyltrimethylammonium, but not by any of a range of anion transport inhibitors tested. Swelling the cells by suspending them in hyposmotic media activated a second choline transport component that was kinetically and pharmacologically distinct from the saturable system. The volume-activated component was nonsaturable (up to 50 mM choline). It was not inhibited by hemicholinium-3 or dodecyltrimethylammonium but was inhibited by anion transport inhibitors, the most potent of which was 5-nitro-2-(3-phenylpropylamino)benzoic acid (NPPB; half-maximal inhibitory concentration = 14 microM). Dose-response curves for the effect of NPPB on swelling-activated choline transport and the swelling-activated transport of taurine, a sulfonic amino acid, were superimposable. It is postulated that the transport of choline and taurine (as well as that of other small organic solutes) in osmotically swollen fish erythrocytes is mediated by a volume-activated, anion-selective channel.

Proton fluxes associated with the Ca pump in human red blood cells

1990 ◽  
Vol 258 (3) ◽  
pp. C552-C562 ◽  
Author(s):  
M. A. Milanick
Keyword(s):  
Red Blood Cells ◽  
Blood Cells ◽  
Disulfonic Acid ◽  
Surprising Result ◽  
Exchange System ◽  
Response Curves ◽  
Ca Pump ◽  
Human Red Blood Cells ◽  
Pump Activity ◽  
Ph Stat

Ca fluxes and H fluxes were measured in human red blood cells at 37 degrees C to characterize the effects of extracellular protons (Hout) on the Ca pump and to determine the stoichiometry of Ca-H exchange. A pH-stat technique was used to measure the rate of H influx, and 45Ca was used to determine the rate of Ca efflux. 4,4'-Diisothiocyanostilbene-2,2'-disulfonic acid (DIDS) was used to reduce proton permeability. A La-sensitive H influx was observed in Ca-loaded cells (Ca = 2 mmol/l packed cells) and was not observed in the cells loaded with vanadate as well as Ca. Similar results were obtained in Ca-loaded ghosts. The La dose-response curves for H influx and for Ca efflux were similar [50% inhibitory concentration (IC50) = approximately 5 microM] in intact red blood cells. The stoichiometry of the La-sensitive fluxes among different experiments ranged from 1.7 to 2.1 H/Ca when extracellular pH (pHout) = 6.3. Thus the Ca pump in intact red blood cells mediates Ca-2H exchange at pHout = 6.3. A 100-fold decrease in Hout [from pH 6.5 to 8.5; intracellular pH (pHin) approximately 7.4] only decreased Ca efflux 1.5- to 3-fold, hence Hout had little effect on the overall rate under the conditions studied. The small effect of Hout was a surprising result for a Ca-H exchange system, since one would have expected a steep dependence of Ca pump on Hout at Hout less than the Michaelis constant (Km). However, no La-sensitive H influx was observed when pHout = 8. On the basis of these data, it is suggested that the Ca pump also mediates Ca efflux uncoupled from H influx (Ca2+/phi H+). Ca efflux in the presence of 11 mM extracellular Ca (Caout) was one-fifth the value obtained in the absence of Caout at pHout = 8.5; this inhibition was reversed by increasing Hout (to pH 6.1). These results are consistent with a model in which 1) the Ca pump mediates Ca2+/2H+ exchange at high Hout; 2) the Ca pump mediates Ca2+/phi H+ exchange at low pHout; 3) the rates of the two processes are less than or equal to 4-fold different; 4) Caout inhibits pump activity at low Hout; and 5) Caout competes with Hout for binding.

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