Effects of drugs on water permeability of erythrocyte membranes (original) (raw)
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Journal of Membrane Biology, 1983
The changes in water diffusion across human erythrocyte membranes following exposure to various inhibitors and proteolytic enzymes have been studied on isolated erythrocytes suspended in isotonic buffered solutions. An important issue was to investigate whether the sulfhydryl reacting reagents that have been applied in osmotic experiments showed similar effects on diffusional permeability. It was found that mercurials, including mersalyl, were the only sulfhydryl reacting reagents that were efficient inhibitors. Under optimal conditions a similar degree of inhibition (around 45%) was found with all mercury-containing sulfhydryl reagents. Other reagents, including the sulfhydryl reagent DTNB, phloretin, or H2DIDS, the specific inhibitor of the anion transport system in erythrocyte membrane, did not appear to inhibit significantly the diffusional permeability. No changes in water diffusion were noticed after exposure of erythrocytes to trypsin and chymotrypsin. A new kind of experiment was that in which the effects of exposure of erythrocytes to two or more agents were studied. It was found that none of the chemical manipulations of membranes that did not affect water diffusion hampered the inhibitory action of mercurials. These findings show that the SH groups involved in water diffusion across erythrocyte membranes do not react with any of the other SH reagents aside from mercurials and that the molecular mechanism of water transport is not affected by chymotryptic cleavage of band 3 protein into the 60 and 35 kD fragments. The NMR method appears as a useful tool for studying changes in water diffusion in erythrocyte membranes following various chemical manipulations of the membranes with the aim of locating the water channel.
Bioscience Reports, 1990
The characteristics of water diffusional permeability (P) of human red blood cells were studied on isolated erythrocytes by a doping nuclear magnetic resonance technique. In order to estimate the basal permeability the maximal inhibition of water diffusion was induced by exposure of red blood cells to p-chloromercuribenzene sulfonate (PCMBS) under various conditions (concentration, duration, temperature). The lowest values of P were around 0.7×10−3 cm s−1 at 10°C, 1.2×10−3 cm s−1 at 15°C, 1.4×10−3 cm s−1 at 20°C, 1.8×10−3 cm s−1 at 25°C, 2.1×10−3 cm s−1 at 30°C and 3.5×10−3 cm s−1 at 37°C. The mean value of the activation energy of water diffusion (Ea,d) was 25 kJ/mol for control and 43.7 kJ/mol for PCMBS-inhibited erythrocytes. The values of P and Ea,d obtained after induction of maximal inhibition of water diffusion by PCMBS can be taken as references for the basal permeability to water of the human red blood cell membrane.
Na-nitroprusside and HgCl2 modify the water permeability and volume of human erythrocytes
Bioelectrochemistry, 2007
The passage of water through the aquaporin-1 (AQP1) transmembrane channel protein of the human erythrocyte is known to be inhibited by organic mercurials such as p-chloromercuribenzoate (pCMB), which react with the free SH-group of the critical cysteine (Cys189) located near the constriction of the AQP1 water-specific channel. Sodium nitroprusside (SNP), which is known as a nitric oxide (NO) donor in interactions with SH-containing molecules, is shown here to suppress the diffusional water permeability (P d ) of the erythrocyte membrane, presumably as a result of reaction with the Cys189 of the human erythrocyte AQP1 water channels. Further, treatment of erythrocytes with HgCl 2 is found to result in a cell volume decrease that can be related to activation of membrane K + -selective Gárdos channels and subsequent loss of intracellular K + and cell shrinkage. The variations in P d and volume of the erythrocyte were deduced from induced variations in the measured proton ( 1 H) nuclear magnetic resonance (NMR) transverse (T 2 ) relaxation functions of water exchanging between diamagnetic intracellular and paramagnetic extracellular compartments of the 20-25% hematocrit samples. The extracellular solvent contained 10 mM membrane-impermeable paramagnetic Mn 2+ ions. The 1 H-T 2 NMR technique allows determination of the time constant τ exch (for exchange of the erythrocyte intracellular water) that is inversely proportional to the permeability coefficient P d when the intracellular water volume is left unmodified, as in the case of SNP-treated erythrocytes. However, for HgCl 2 -treated erythrocytes, this technique showed simultaneous variation of both τ exch and the volume ratio V in /V out of intracellular and extracellular water in proportions suggesting that P d was left unmodified. The HgCl 2 effect has been found to be partly reversible by the reducing activity of added mercaptoethanol.
African Journal of Biomedical Research, 2010
The effect of membrane potential change of the human erythrocytes on cationic drugs tetracaine and chlorpromazine and neutral drug benzyl alcohol induced cell shape change and red cell uptake of drug has been quantitated using light microscopy and spectrophotometry respectively. At the drug concentration necessary to cause cell membrane cell shape change membrane potential change from -7.1 mV to 1 6.4mV let to the reversal of the cup-forming property of chlorpromazine and tetracaine to that of a crenetor at both 20°C and 37°C. The effect of altering the membrane potential from -7. lmV to 16.4mV also led to the decrease of cellular uptake of drug with increasing membrane diffusion potential. The membrane potential dependent drug induced cell shape change with also reversible on reversing the membrane potential. The results therefore suggest that the cellular uptake of drug and drug induced cell shape change in human erythrocytes was dependent on change in extracellular chloride concentration
Journal of Membrane Biology, 1989
The water permeability of human red blood cell (RBC) membrane has been monitored by a doping nuclear magnetic resonance (NMR) technique on intact cells and resealed ghosts following exposure to various sulfydryl-reacting (SH) reagents and proteolytic enzymes. The main conclusions are the following: (i) When appropriate conditions for exposure of erythrocytes or ghosts to mercury-containing SH reagents (concentration, temperature and duration of incubation) were found, the maximal inhibition of water diffusion could be obtained with all mercurials (including HgCl2 and mersalyl that failed to show their inhibitory action on RBC water permeability in some investigations). While previous studies claimed that long incubation times are required for the development of maximal inhibition of water diffusion by mercurials, the present results show that it can be induced in a much shorter time (5–15 min at 37°C) if relatively high concentrations of PCMBS (2–4mm) are used and no washings of the inhibitor are performed after incubation. Higher than optimal concentrations of mercurials and/or longer incubation times result in lower values of inhibition, sometimes a loss of inhibition, or can even lead to higher values of permeability compared to control RBCs. (ii) The conditions for inhibition by mercurials are drastically changed by preincubation of erythrocytes with noninhibitory SH reagents (such as NEM or IAM) or by exposure to proteolytic enzymes. If the cells are digested with papain, the duration of incubation with PCMBS should be decreased in order for inhibition to occur. This explains the lack of inhibition reported previously, when a relatively long duration of incubation with PCMBS was used subsequent to papain digestion. (iii) The degree of inhibition of water diffusion induced by mercurials appeared to be dependent upon the temperature of which the water permeability was measured. The values of maximal inhibition ranged from 45–50% at 37°C, increased 10–15% at 20°C and further increased at lower temperatures, reaching values above 75% below 10°C; these results clarify the conflicting reports of various authors. (iv) The inhibition of water diffusion, either reversible, or irreversible, was not accompanied by significant changes in the pattern of RBC membrane polypeptides fractionated by polyacrylamide gel electrophoresis. (v) The mean value of the activation energy of water diffusion (E a,d) obtained on 42 donors was 25.6 kJ/mol. The values ofE a,d increased in parallel with the values of the inhibition of water diffusion induced by PCMBS until the maximal inhibition was reached (whenE a,d=41 kJ/mol) and then both sets of values decreased in parallel.
Journal of Pharmacological Methods, 1985
The electron spin resonance (EPR) spectrum of erythrocyte membranes labeled with 2,2,6,6-tetra-methyl-4-maleimido-piperidin-l-oxyl (MAL6) indicates both weakly and strongly immobilized labeling sites. Changes in the ratio of the membrane's low field spectral peaks (W/S) have often been used to monitor drugerythrocyte interactions. We have investigated a number of experimental factors that may influence this ratio even in the absence of drug. Instrumental settings on the EPR spectrometer had no obvious effect. As the weight ratio of label/protein decreased, the W/S ratio increased. Similarly an increase in labeling time and temperature lead to an increase in the ratio. The ratio also increased with time after labeling; this change was most marked in samples kept at 37"C, but was insignificant in samples kept at 4°C. Increasing the viscosity of the sample with low-molecular-weight substances such as sucrose or glycerol, but not with those with high molecular weight such as dextran and PVP, caused a reduction in the ratio. Increasing the pH and changing the buffer concentrations also lead to a small increase in W/S. These results suggest that it is very important that all of these factors be kept constant and at some optimal value if reliable and consistent results are to be obtained using this method to monitor drug-erythrocyte interactions.
Water transport in human red cells: effects of ‘non-inhibitory’ sulfhydryl reagents
Biochimica Et Biophysica Acta-biomembranes, 1991
The water diffusional permeability of human red blood cells following exposure to various sulfhydryl group (SH) reagents have been studied using a nuclear magnetic resonance technique. Exposure of red blood cells up to 12 mM N-ethylmaleimide (NEM) or 10 mM 5,5'-dithio-bis(2-nitrobenzoic acid) (DTNE) alone does not affect water diffusion. in contrast, when DTNB treatment follows a preincubation of the cells with NEM, a small (18% at 37°C) but significant inhibition of water permeability occurs. The NEM and DTNB treatment of the cells caused no change of the cell shape and volume or of the cell water volume. Consequently, the inhibition observed after NEM and DTNB treatment has a real significance.
The effect of selected membrane active substances on erythrocyte deformability
Bratislavske lekarske listy
Aminoguanidine improved the erythrocyte filterability by 4%, pyridoxyliden-aminoguanidine by 11% and pyridoxal by 13% in healthy subjects. In diabetic patients the aminoguanidine effect on erythrocyte filterability was improved by 7%, PAG effect by 9% and pyridoxal effect by 15% in comparison to the control group. The other investigated haematological variables in both groups were within the range of the physiological standard. All of the tested substances demonstrated a mild protective influence on erythrocyte elasticity both in healthy subjects and diabetic patients. Significant elasticity improvement was obtained only by pyridoxal (p<0.01) in patients with diabetes mellitus. (Fig. 4, Ref. 18.).