Changes In Intracellular Sodium Research Articles

Intracellular Sodium Changes in Cancer Cells Using a Microcavity Array-Based Bioreactor System and Sodium Triple-Quantum MR Signal

The sodium triple-quantum (TQ) magnetic resonance (MR) signal created by interactions of sodium ions with macromolecules has been demonstrated to be a valuable biomarker for cell viability. The aim of this study was to monitor a cellular response using the sodium TQ signal during inhibition of Na/K-ATPase in living cancer cells (HepG2). The cells were dynamically investigated after exposure to 1 mM ouabain or K+-free medium for 60 min using an MR-compatible bioreactor system. An improved TQ time proportional phase incrementation (TQTPPI) pulse sequence with almost four times TQ signal-to-noise ratio (SNR) gain allowed for conducting experiments with 12–14 × 106 cells using a 9.4 T MR scanner. During cell intervention experiments, the sodium TQ signal increased to 138.9 ± 4.1% and 183.4 ± 8.9% for 1 mM ouabain (n = 3) and K+-free medium (n = 3), respectively. During reperfusion with normal medium, the sodium TQ signal further increased to 169.2 ± 5.3% for the ouabain experiment, while it recovered to 128.5 ± 6.8% for the K+-free experiment. These sodium TQ signal increases agree with an influx of sodium ions during Na/K-ATPase inhibition and hence a reduced cell viability. The improved TQ signal detection combined with this MR-compatible bioreactor system provides a capability to investigate the cellular response of a variety of cells using the sodium TQ MR signal.

Sevoflurane Preconditioning Attenuates the Fall in Adenosine Triphosphate Levels, but Does Not Alter the Changes in Sodium and Potassium Levels during Hypoxia in Rat Hippocampal Slices

Sevoflurane preconditioning improves recovery after hypoxia. Sevoflurane administered before and during hypoxia improved recovery and attenuated the changes in intracellular sodium, potassium, and adenosine triphosphate (ATP) levels during hypoxia. In this study, the authors examine the effects of sevoflurane applied only before hypoxia on sodium, potassium, and ATP. Hippocampal slices from adult male Sprague-Dawley rats were pretreated with 4% sevoflurane, washed, and then subjected to hypoxia (n≥8 animals/group). The cornus ammonis 1 regions of the hippocampal slices were micro-dissected and sodium, potassium, and ATP concentrations measured. Pretreatment with sevoflurane for 15 or 60 min did not attenuate the increase in intracellular sodium or the decrease in intracellular potassium during hypoxia. After 60 min of preconditioning and 5 min of hypoxia, sodium increased 57% (vs. nonpreconditioned hypoxia 54% increase) and potassium decreased 31% (vs. 26%). These changes were not statistically significant versus untreated hypoxia. The 60-min sevoflurane preconditioning group had statistically significant higher ATP levels at 5 min of hypoxia (3.8 nmol/mg dry wt.) when compared to untreated hypoxic tissue (2.1 nmol/mg). There was no significant difference in ATP levels between the sevoflurane preconditioned and the untreated tissue before hypoxia (8.9 vs. 8.5 nmoles/mg, respectively). Preconditioning with sevoflurane for 60 min before hypoxia does not alter changes in intracellular sodium and potassium during hypoxia but does attenuate the fall in intracellular ATP levels during hypoxia. Thus, there are differences between anesthetic preconditioning and when anesthetics are present before and during hypoxia.

Modulation of canine cardiac sodium current by Apelin

Apelin, a ligand of the G protein-coupled putative angiotensin II-like receptor (APJ-R), exerts strong vasodilating, cardiac inotropic and chronotropic actions. Its expression is highly up-regulated during heart failure. Apelin also increases cardiac conduction speed and excitability. While our knowledge of apelin cardiovascular actions is growing, our understanding of the physiological mechanisms behind the cardiac effects remains limited. We tested the effects of apelin on the cardiac sodium current (INa) using patch clamp technique on cardiac myocytes acutely dissociated from dog ventricle. We found that apelin-13 and apelin-17 increased peak INa by 39% and 61% and shifted its mid-activation potential by −6.8±0.6 mV and −17±1 mV respectively thus increasing channel opening at negative voltage. Apelin also slowed INa recovery from inactivation. The effects of apelin on INa amplitude were linked to activation of protein kinase C. Apelin also increased INa “window” current by up to 600% suggesting that changes in intracellular sodium may contribute to the apelin inotropic effects. Our results reveal for the first time the effects of apelin on INa. These effects are likely to modulate cardiac conduction and excitability and may have beneficial antiarrhythmic action in sodium chanelopathies such as Brugada Syndrome where INa amplitude is reduced.

Changes in intracellular sodium, chlorine, and potassium concentrations in staurosporine‐induced apoptosis

Ion gradients across the plasma membrane, fundamentally K(+), play a pivotal role in the execution phase of apoptosis. However, little is known about other monovalent anions (Cl(-)) or cations (Na(+)) in apoptosis. In addition, the relationship between changes in total ion composition and morphological and biochemical events are poorly understood. We investigated simultaneous changes in sodium (Na), chlorine (Cl), and potassium (K) concentrations in stauroporine-induced apoptosis by quantitative electron probe X-ray microanalysis (EPXMA) in single cells. Apoptotic cells identified unequivocally from the presence of chromatin condensation in backscattered electron images were characterized by an increase in intracellular Na, a decrease in intracellular Cl and K concentrations, and a decrease in K/Na ratio. The ouabain-sensitive Rb-uptake assay demonstrated a net decrease in Na(+)/K(+)-ATPase activity, suggesting that increases in Na and decreases in K and the K/Na ratio in apoptotic cells were related with inhibition of the Na(+)/K(+)-ATPase pump. These changes in diffusible elements were associated with externalization of phosphatidyl serine and oligonucleosomal fragmentation of DNA. This alteration in ion homeostasis and morphological hallmarks of apoptosis occur in cells that have lost their inner mitochondrial transmembrane potential and before the plasma membrane becomes permeable.

Functional innervation in tissue engineered models for in vitro study and testing purposes.

The biotechnology industry is rapidly expanding and the emerging field of tissue engineering is projected to have a high impact in the near future. Recently the field of cellular, drug, and prosthetic delivery has melded with the field of tissue engineering to make simulated tissues. In addition to their roles as tissue substitutes for transplantation, these simulated tissues may provide more accurate models and environments for toxicology testing and the study of peripheral nerves. The current study demonstrates the importance of innervation, in general, for the function of engineered tissues. We observe that the presence of nerves in a tissue engineered (TE) human cornea model enhances the growth of the epithelium and the formation of its protective mucin layer. Innervation also confers protection to the epithelium from chemical insult, as determined by the level of post-treatment epithelial cell death. We demonstrate differential responses of the nerves to chemical stimuli by changes in intracellular sodium as measured by 2-photon microscopy. The 2-photon imaging techniques also allow for the visualization and study of the fine sensory axon fibers within the 3-dimensional tissue. This work demonstrates a role for innervation in the protective quality and function of the engineered tissue, and the potential to use the nerves themselves as indicators of the severity of an insult. These results are important to consider for the development of any optimized TE models for in vitro study and testing purposes.

Regulation of Na,K-ATPase expression by endothelin-1 in transformed human ciliary non-pigmented epithelial (HNPE) cells.

Endothelin-1 (ET-1) (1-100 nM) decreases the activity of Na,K-ATPase, a key enzyme responsible for aqueous humor formation, in transformed human non-pigmented ciliary epithelial (HNPE) cells. The present study sought to determine if ET-1 alters the expression of the catalytically active alpha subunit of Na,K-ATPase in HNPE cells and identify mechanisms underlying these effects. We report that acute (15 and 30 min) treatment with ET-1 results in an increase in mRNA expression of the alpha 1 subunit of Na,K-ATPase. Similar to ET-1's effects, ouabain (100 microM), a selective inhibitor of Na,K-ATPase, and monensin (10 microM), a sodium ionophore, also increased Na,K-ATPase expression in HNPE cells. The increase in Na,K-ATPase expression by short-term treatment with ouabain and monensin was dependent on their ability to elevate intracellular sodium concentrations. However, acute ET-1 treatment mediated increase in Na,K-ATPase expression was independent of changes in intracellular sodium. A prolonged (24 hr) ET-1 treatment results in an increase in both mRNA and protein levels of the alpha 1 subunit of Na,K-ATPase. These observations suggest that ET-1 could play a homeostatic role in modulating aqueous humor formation by having differential effects on the activity and expression of Na,K-ATPase by the ciliary epithelium in the eye.

Changes in intracellular sodium, potassium, and calcium concentrations in transplanted mouse pancreatic islets.

Our previous studies have suggested a chronically low oxygen tension in transplanted pancreatic islets. The present study tested the hypothesis that this may be coupled to changes in intracellular concentrations of crucial ions within the transplanted islet cells and, thus, their function. X-ray microanalysis was used for studies of native islet cells and islet grafts residing for 1 day or 1 month in nondiabetic or diabetic recipients. Markedly increased sodium concentrations and decreased potassium concentrations were recorded in all transplanted islet cells, irrespective of whether the grafts had been implanted into nondiabetic or diabetic recipients or whether they were investigated 1 day or 1 month after transplantation. The calcium concentration in 1-day-old islet grafts was similar to that in native islet cells, but it decreased markedly between 1 day and 1 month after transplantation. Again this was seen in both nondiabetic and diabetic recipients. Most probably, the disturbances in graft sodium and potassium concentrations reflect ATP depletion and inhibition of the Na/K-ATPase in the plasma membrane as a result of impeded oxygen supply. The decreased calcium concentrations developing over time in the transplanted islet cells might be potentially detrimental, because calcium plays a fundamental role in the control of a variety of cellular functions, including insulin secretion, in beta cells.

Intracellular sodium changes during the speract response and the acrosome reaction in sea urchin sperm.

The sperm-activating peptide speract and fucose-sulphate glycoconjugate (FSG) are sea urchin egg-envelope components that modulate sperm ion permeability. They influence motility and induce acrosomal reaction (AR), respectively. A fluorescent Na(+)-sensitive dye (Na(+)-binding benzofuran isophthalate, SBFI) was used to determine how these egg envelope components influence sperm Na(+) permeability. [Ca(2+)](i) and pH(i) were also measured to correlate their changes in response to speract and FSG with those observed in [Na(+)](i). SBFI determinations indicate that the resting [Na(+)](i) is 20 +/- 8 mM in sea urchin sperm. Saturating levels of speract increased [Na(+)](i) by approximately 15 mM, while similar levels of FSG caused a further elevation of approximately 30 mM. The kinetics of the [Na(+)](i), [Ca(2+)](i) and pH(i) changes induced by saturating levels of speract were faster than those induced by FSG. Both egg ligands appeared to activate more than one Na(+) transport system. Nifedipine, Ni(2+) and TEA(+) inhibited the ionic changes and the AR induced by FSG but, importantly, did not alter those caused by speract. Thus, there are differences in some of the ionic transport mechanisms that operate in the speract and FSG responses. ZD2788, a blocker of hyperpolarization and cyclic-nucleotide-gated (HCN) channels such as SpHCN present in sea urchin sperm, did not decrease the speract-induced [Na(+)](i) increase, but slowed its kinetics. Therefore, SpHCN does not play a major role in the uptake of Na(+) triggered by this decapeptide. KB-R7943, an inhibitor of Na(+)/Ca(2+) exchangers, decreased the resting [Na(+)](i) and did not change significantly the speract-induced [Ca(2+)](i) increase, but slowed its recovery.

Apoptotic volume decrease and the incredible shrinking cell.

Apoptotic volume decrease and the incredible shrinking cell.

Metabolic Modulation

featured in this issue of Circulation presents data showingthat pyruvate, which dose-dependently improved perfor-mance in myocardium from failing human hearts, may be onesuch agent.A number of abnormalities in both energy metabolism andcontractile function have been identified in failing cardiacmuscle. Metabolic changes include decreases in high energyphosphorylation potential, decreases in mitochondrial oxida-tive phosphorylation capacity, and a switch to a more fetalglycolytic metabolism.

Enhancement of glutamate uptake transport by CO2/bicarbonate in the leech giant glial cell

Glutamate uptake into glial cells via the excitatory amino acid transporter (EAAT) is accompanied by an influx of sodium and acid equivalents into the cells. The sodium-bicarbonate cotransport (NBC) in glial cells moves sodium and base equivalents across the glial membrane in both directions. We have studied possible interactions between these two electrogenic transporters in the giant glial cell of isolated ganglia of the leech Hirudo medicinalis. Changes in membrane potential, membrane current, intracellular sodium, and intracellular pH evoked by aspartate (1 mM), an EAAT agonist, were measured both in the absence and in the presence of CO(2)/bicarbonate. When 5% CO(2) and 24 mM bicarbonate was added to the saline (at constant pH 7.4), the aspartate-induced membrane current was increased, while the change in intracellular sodium was decreased. The acid influx evoked by aspartate was enhanced by CO(2)/bicarbonate but, because of the increased intracellular CO(2)/bicarbonate-dependent buffering power, the change in intracellular pH was decreased. 4,4'-Diisothiocyanatostilbene-2, 2'-disulfonic acid (DIDS, 0.5 mM), which inhibits the NBC, reversed the effects of CO(2)/bicarbonate on the aspartate-induced current and pH change. Our results suggest that the NBC helps counteract dissipation of the sodium and the acid-base gradients induced by the EAAT, enhancing the rate and capacity of glutamate uptake by glial cells.

Attenuation of ischemia induced increases in sodium and calcium by the aldose reductase inhibitor zopolrestat.

We have previously demonstrated that zopolrestat, an inhibitor of the enzyme aldose reductase, reduces ischemic injury in hearts from diabetic and non-diabetic rats. To further explore potential cardioprotective mechanisms of zopolrestat, we measured changes in intracellular sodium, calcium, and Na+,K(+)-ATPase activity in zopolrestat treated hearts during ischemia and reperfusion. Hearts from acute diabetic (Type I) and age-matched control rats were isolated and retrogradely perfused. Hearts had either control perfusion or exposure to 1 microM zopolrestat for 10 min, followed by 20 min of global ischemia and 60 min of reperfusion. Changes in intracellular sodium and calcium were measured using 23Na and 19F magnetic resonance spectroscopy, respectively, while the activity of Na+,K(+)-ATPase was measured using biochemical assays. Zopolrestat blunted the rise in [Na]i during ischemia in both diabetic hearts and non-diabetic hearts. The end-ischemic [Na]i was 21.3 +/- 2.6 mM in the zopolrestat treated diabetics and 25.9 +/- 2.3 in zopolrestat treated non-diabetics, versus 31.6 +/- 2.6 mM and 32.9 +/- 2.8 mM in the untreated diabetics and untreated non-diabetics, respectively, (P = 0.002). Similarly, the rise in [Ca]i at the end of ischemia was significantly reduced in zopolrestat treated diabetic and non-diabetic hearts (P = 0.005). Zopolrestat increased the activity of Na-,K(+)-ATPase in diabetic hearts under baseline conditions (11.70 +/- 0.95 versus 7.28 +/- 0.98 mumol/h/mg protein, P = 0.005) as well as during ischemia and reperfusion. Similar changes in Na+,K(+)-ATPase activity were also observed in non-diabetic hearts. The data provide additional support to the protective effects of zopolrestat and suggest that a possible mechanism of action may be associated with the attenuation of the rise in [Na]i and [Ca]i during ischemia and reperfusion.

Alpha-Adrenergic effects on Na+-K+ pump current in guinea-pig ventricular myocytes.

1. The whole-cell patch clamp was employed to study Na+-K+ pump current (Ip) in acutely isolated myocytes. alpha-Adrenergic receptors were activated with noradrenaline (NA) after blocking beta-adrenergic receptors with propranolol. Ip was measured as the current blocked by strophanthidin (Str). 2. Activation of alpha-receptors by NA increased Ip in a concentration-dependent manner. The K0.5 depended on intracellular calcium ([Ca2+]i), however maximal stimulation did not. At 15 nM [Ca2+]i the K0.5 was 219 nM NA whereas at 1.4 microM [Ca2+]i it was 3 nM. 3. The voltage dependence of Ip was not shifted by NA at either high or low [Ca2+]i. At each voltage, maximal stimulation of Ip was 14-15 %. 4. Staurosporine (St), an inhibitor of protein kinase C (PKC), eliminated the alpha-receptor-mediated stimulation of Ip at either high or low[Ca2+]i. 5. The stimulation of Ip was independent of changes in intracellular sodium or external potassium concentrations, and did not reflect a change in affinity for Str. 6. Phenylephrine, methoxamine and metaraminol, three selective alpha1-adrenergic agonists, stimulate Ip in a similar manner to NA. Stimulation of Ip by NA was eliminated by prazosin, an alpha1-antagonist, but was unaffected by yohimbine, an alpha2-antagonist. 7. We conclude noradrenaline activates ventricular alpha1-receptors, which are specifically coupled via PKC to increase Na+-K+ pump current. The sensitivity of the coupling is [Ca2+]i dependent, however the maximal increase in pump current is [Ca2+]i and voltage independent.

Heat-Induced Changes in Intracellular Sodium and Membrane Potential: Lack of a Role in Cell Killing and Thermotolerance

Hyperthermia induces transient changes in intracellular free sodium levels and membrane potential. The possible role of these changes in cell killing by hyperthermia and thermotolerance has been evaluated using Chinese hamster ovary IS1 and HeLa cells. Intracellular sodium was measured with Sodium Green and SBFI, while membrane potential was measured with the oxonol dye diBAC4(3). Heating at either 42.0 or 45.0 degrees C caused nearly the same decrease in free [Na+]i from about 20 mM in unheated cells to 5-7 mM in heated cells. However, survival differed by over two orders of magnitude after heating for 30 min at these two temperatures. In addition, blockage of the heat-induced decrease in [Na+]i using ouabain and/or amiloride did not affect the survival curves for heated cells. Hyperthermia also induced a membrane hyperpolarization of 15 mV after 15 min at 42.0 degrees C or 35 mV after 15 min at 45.0 degrees C which could be blocked with ouabain and amiloride. Both the free [Na+]i and membrane potential recovered to near baseline levels within 30-40 min after heating. Induction of thermotolerance using a 45.0 degrees C, 10-min heat treatment also was not affected by ouabain and/or amiloride. Finally, thermotolerant cells experienced the same heat-induced changes in free [Na+]i and membrane potential as non-thermotolerant cells. We conclude that the heat-induced changes in free [Na+]i and membrane potential are not directly related to cell killing by hyperthermia or thermotolerance.

Diacylglycerol activation of protein kinase C results in a dual effect on Na+,K(+)-ATPase activity from intact renal proximal tubule cells.

This study evaluated the effect of L-1-oleoyl-2-acetyl-sn-3-glycerol (OAG) on ouabain-sensitive Na,K-dependent oxygen consumption (Na,K-QO2) in intact renal proximal tubule cells (RPTC). Basal Na,K-QO2 (nmol O2/mg protein per min) was 20.0 +/- 1.0. Incubation with 10 nM of OAG induced a dual effect on Na,K-QO2, with an initial stimulation (maximal at 10 min, 37.1 +/- 5.0), followed by an inhibition (significant at 20 min, 16.3 +/- 1.0). No changes in ouabain-insensitive QO2 were observed in any of the protocols. The effects were abolished by sphingosine, a protein kinase C inhibitor. Stimulation was abolished by amiloride 0.1 mM. Amiloride had no effect on Na,K-QO2 at the concentration used. Stimulation was not potentiated by the sodium ionophore, amphotericin B, and the later inhibition was still observed in the presence of amphotericin B. The initial stimulation was attributed to an increase in sodium permeability, while the later inhibition was attributed to a direct effect on the Na,K-pump. Regulation of Na+,K(+)-ATPase activity by protein kinase C in intact RPTC can be accomplished by a direct effect on the protein or as a secondary effect consequent upon changes in intracellular sodium.

Changes in intracellular sodium during the hydroosmotic response to vasopressin

During vasopressin (VP)-induced water movement, toad urinary bladder epithelial cells undergo unique morphological changes. The osmolality within these responding cells remains relatively stable despite the large transcellular transport of water. We hypothesized that the hydroosmotic response to VP may be associated with a net increase in sodium either as an aid in maintaining the intracellular osmolality or as part of a Na-Ca exchange process. Changes in intracellular sodium (Nai) were monitored over time in individual hemibladders using 23Na NMR. Hemibladders were mounted as bags on glass pipets and filled with deionized water. During NMR studies, the serosal bath consisted of aerated 2.4 mM HCO3 amphibian Ringer's (pH 8.1) made up with 15% D2O containing the shift reagent, dysprosium tripolyphosphate (1 mM). This reagent allowed for visualization of Nai by shifting the extracellular Na signal; it did not affect basal or VP stimulated water flow, short-circuit current, or high energy phosphate metabolism as seen by 31P NMR. Changes in Nai were determined by integrating the area under the unshifted Na peak at each measurement and expressing differences as a ratio relative to baseline. The initial Nai signal from unstimulated hemibladders remained stable in these tissues over at least 180 minutes. Within 30 minutes of VP (20 mU/ml) exposure, however, the Nai peak increased 2.47 times above pretreatment baseline (N = 16, P less than 0.001). The Nai signal returned toward baseline values with removal of VP from the serosal bath but only after approximately 90 minutes. When change in cell shape and water movement were prevented by having isotonic sorbitol in the mucosal bath, VP produced no change in the Nai signal (N = 10).(ABSTRACT TRUNCATED AT 250 WORDS)

Local Myocardial Biochemical and Ionic Alterations During Myocardial Ischaemia and Reperfusion

Acute myocardial ischaemia and reperfusion result in a series of inhomogeneous metabolic, ionic and neurohumoral events that explain the associated mechanical and electrical events, including cardiac death. The time course of the hydrolysis of high energy phosphates, the rise in extracellular potassium and the fall in intracellular and extracellular pH induced by acute no-flow ischaemia have been well characterised. However, the time course of the changes in intracellular sodium, calcium and magnesium levels is less clear. It appears that the changes in intracellular calcium may be pivotal to many of the biochemical and electrophysiological changes produced by the abrupt cessation of coronary arterial inflow and the associated interruption of venous washout. Consequently, agents that modify the handling of calcium by the sarcolemma and the sarcoplasmic reticulum have a significant impact on many of the metabolic, ionic and electrical abnormalities characterising acute ischaemia and reperfusion.

A review of 23Na nuclear magnetic resonance spectroscopy for the in vitro study of cellular sodium metabolism

Changes in intracellular sodium have been associated with a number of different diseases. Consequently, various methods have been used to quantify the level of intracellular sodium concentrations. Traditional methods like flame photometry and ion-selective electrodes are destructive or invasive, thereby potentially altering the intracellular sodium levels. There has been an increasing interest in evaluating the method of 23Na nuclear magnetic resonance in recent years, since this method allows for non-invasive continuous monitoring of intracellular sodium in cell suspensions and tissues. A phenomenological approach to basic theory, review of methodology, applications to the in vitro study of cellular sodium metabolism, and difficulties of interpretation of this analytical modality is presented.

A review of23Na nuclear magnetic resonance spectroscopy for thein vitrostudy of cellular sodium metabolism

Changes in intracellular sodium have been associated with a number of different diseases. Consequently, various methods have been used to quantify the level of intracellular sodium concentrations. Traditional methods like flame photometry and ion-selective electrodes are destructive or invasive, thereby potentially altering the intracellular sodium levels. There has been an increasing interest in evaluating the method of 23Na nuclear magnetic resonance in recent years, since this method allows for non-invasive continuous monitoring of intracellular sodium in cell suspensions and tissues. A phenomenological approach to basic theory, review of methodology, applications to the in vitro study of cellular sodium metabolism, and difficulties of interpretation of this analytical modality is presented.

NMR studies of intracellular free calcium, free magnesium and sodium in the guinea pig reticulocyte and mature red cell

During the maturation process reticulocytes lose their intracellular organelles and undergo changes in membrane lipid composition and ion transport properties. While several reports indicate differences in the levels of magnesium, sodium and calcium in reticulocytes and erythrocytes, controversy remains concerning the actual magnitude and direction of ionic alterations during reticulocyte maturation. One problem with all of these studies is that the techniques used are invasive and are limited to measuring only the total cell ion content. We have used 31P, 23Na and 19F nuclear magnetic resonance (NMR) spectroscopy to compare the intracellular free ion and phosphometabolite levels in guinea pig reticulocytes and mature red blood cells. In contrast to a sharply decreased concentration of ATP in erythrocytes in comparison to reticulocytes, the intracellular free magnesium, measured using 31P-NMR, was increased by about 65% upon maturation (150 mumol/l cell water in reticulocytes in comparison to 250 mumol/l cell water in erythrocytes). Sizeable but opposite changes in intracellular sodium (5.5 mumol/ml cells in reticulocytes vs. 8.5 mumol/ml cells in erythrocytes) and intracellular free calcium (99 nM vs. 31 nM in reticulocytes and mature red cells, respectively) were also observed, suggesting that alterations in the kinetics of membrane ion transport systems, accompanying changes in phospholipid and cholesterol content, occur during the process of red cell maturation. However, in contrast to dog red blood cells, there was no evidence for the presence of a Na+/Ca2+ exchanger in guinea pig reticulocytes or erythrocytes.