LITHIUM INHIBITS GROWTH IN A MURINE NEURAL PRECURSOR CELL LINE

The influence of lithium on cell growth and cell viability was studied in short-term cultures of a neural precursor cell line (NT) developed from a murine teratocarcinoma. At very low concentrations ranging from 0.1 m to 1 m Li₂CO₃(equivalent to therapeutic blood concentrations) there was no difference between untreated and treated cultures. 10 m lithium (Li⁺) was found to be toxic with 33% of cell death, while there was inhibition of growth without cell death at concentrations of 2.5 m and 5 m of Li⁺. In experiments where 2.5 m Li⁺was added at the time of seeding, there was growth arrest on day 1 followed by recovery on day 2. Flow cytometric analysis revealed that cells treated with Li⁺were blocked in S phase. At 5 m concentration of Li⁺, the recovery occurred on day 3 and the plating efficiency was significantly low. The ability to form colonies in soft agar was reduced at 2.5 m and 5 m concentrations of Li⁺to an equal extent. Thus, Li⁺has growth inhibitory as well as anchorage-independent growth reducing effects. The NT cell line therefore would be a good model system to study the mechanism of teratogenic effect of Li⁺.     

Lithium transport by fibroblastic mouse cells: Characterization and stimulation by serum and growth factors in quiescent cultures

Serum enhances the rate of Li⁺ entry and exit in quiescent cultures of mouse fibroblasts by 2- to 3-fold. Tertiary cultures of whole mouse embryos as well as established fibroblast lines (3T3, 3T6) show the increase in Li⁺ permeability when serum is added to cultures whose growth has been arrested by serum deprivation. Growing cells are only slightly more permeable to Li⁺ in the presence of serum. Purified compounds which initiate DNA synthesis also rapidly increase Li⁺ entry; mitogenic levels of thrombin and the combination of epidermal growth factor, insulin, and bovine serum albumin were the most effective ones tested. The effect of serum on Li⁺ uptake occurs within a few minutes, is not affected by inhibitors of macromolecular synthesis, and appears mainly to increase the Vmax of entry. Inhibitors of energy production partially reduce Li⁺ entry but do not block the activation by serum. One portion of Li⁺ uptake (?40%), which is inhibited by ouabain, phloretin, or Na⁺ deprivation, is mediated by the Na⁺/K⁺ pump in the plasma membrane. A second mechanism of Li⁺ entry which is blocked by Na⁺ or amiloride appears to be a Na⁺ specific “porter.” The activity of both components is stimulated by serum. The increased activity of the putative Na⁺ porter would increase Na⁺ availability to the Na⁺ pump and may account for its enhancement by serum, which was also noted previously (Rozengurt and Heppel, '75).     

Li/Na exchange and Li active transport in human lymphoid cells U937 cultured in lithium media

Lithium transport across the cell membrane is interesting in the light of general cell physiology and because of its alteration during numerous human diseases. The mechanism of Li⁺ transfer has been studied mainly in erythrocytes with a slow kinetics of ion exchange and therefore under the unbalanced ion distribution. Proliferating cultured cells with a rapid ion exchange have not been used practically in study of Li⁺ transport. In the present paper, the kinetics of Li⁺ uptake and exit, as well as its balanced distribution across the plasma membrane of U937 cells, were studied at minimal external Li⁺ concentrations and after the whole replacement of external Na⁺ for Li⁺. It is found that a balanced Li⁺ distribution attained at a high rate similar to that for Na⁺ and Cl^? and that Li⁺/Na⁺ discrimination under balanced ion distribution at 1–10 mM external Li⁺ stays on 3 and drops to 1 following Na, K-ATPase pump blocking by ouabain. About 80% of the total Li⁺ flux across the plasma membrane under the balanced Li⁺ distribution at 5 mM external Li⁺ accounts for the equivalent Li⁺/Li⁺ exchange. The majority of the Li⁺ flux into the cell down the electrochemical gradient is a flux through channels and its small part may account for the NC and NKCC cotransport influxes. The downhill Li⁺ influxes are balanced by the uphill Li⁺ efflux involved in Li⁺/Na⁺ exchange. The Na⁺ flux involved in the countertransport with the Li⁺ accounts for about 0.5% of the total Na⁺ flux across the plasma membrane. The study of Li⁺ transport is an important approach to understanding the mechanism of the equivalent Li⁺/Li⁺/Na⁺/Na⁺ exchange, because no blockers of this mode of ion transfer are known and it cannot be revealed by electrophysiological methods. Cells cultured in the medium where Na⁺ is replaced for Li⁺ are recommended as an object for studying cells without the Na,K-ATPase pump and with very low intracellular Na⁺ and K⁺ concentration.     

Lithium Induces Apoptosis in Immature Cerebellar Granule Cells but Promotes Survival of Mature Neurons

Lithium (Li⁺) has been used in the treatment of manic—depressive disorders for several decades. More recently, Li⁺ has been shown to affect the signaling pathway of various neurotransmitters and growth/neurotrophic factors. We examined the effect of Li⁺ on the survival of cerebellar granule neurons in culture. Treatment of immature granule cells with Li⁺ resulted in programmed cell death (apoptosis). The death process is accompanied by DNA fragmentation, a hallmark of apoptosis. Following maturation in vitro, granule neurons are dependent on elevated concentrations of extracellular potassium ([K⁺]_o) for survival. Lowering of [K⁺]_o to physiological levels induces apoptosis. Surprisingly, Li⁺ prevents death of mature neurons caused by low [K⁺]_o. Moreover, the concentration range at which Li⁺ exerts its protective effect is the same as that at which it induces apoptosis in immature neurons. Thus, a single agent under similar extracellular conditions has opposing effects on survival, depending on the developmental status of the neuron.     

Regional and Subcellular Localization of Li+ and Other Cations in the Rat Brain Following Long-Term Lithium Administration

Abstract: Rats were given LiCl in their diet (40 mmol/kg dry weight) for at least 3 months to elucidate the regional and subcellular localization of Li⁺ in the brain as well as the effect of chronic lithium administration on the distribution of other cations. At steady-state the mean concentrations of Li⁺ were 0.66 mmol/kg wet weight in the whole brain and 0.52 mM in plasma. The tissue/plasma concentration ratio exceeded unity in all anatomical regions. No region showed excessive accumulation of Li⁺. Whole brain or regional contents of Na⁺ or K⁺ were unaffected by lithium treatment. Subcellular Li⁺ localization was demonstrated in nuclear, crude mitochondrial, and microsomal fractions of whole brain homogenate. Subfractionation of the crude mitochondrial fraction revealed energy-independent intrasynaptosomal and intramitochondrial Li⁺ and K⁺ localization at 0–4°C. Li⁺ administered in vivo disappeared within 10 min from synaptosomes incubated at 37°C. Li⁺ added in vitro at 1 mM attained a synaptosomal steady-state concentration within 30 min at 37°C. In control rats, synaptosomal concentrations and synaptosomal/medium concentration gradients of cations paralleled their respective in vivo concentrations and gradients. Lithium treatment caused synaptosomal depletion of K⁺ and Mg²⁺ and hence probably partial membrane depolarization. Addition of 1 mM Li⁺ in vitro also caused synaptosomal Mg²⁺ depletion. The results indicate that Li⁺ is “accumulated” in brain sediments and synaptosomes following its long-term treatment. The estimated intracellular and intrasynaptosomal Li⁺ concentrations are lower than predicted by passive distribution according to the Nernst equation, evidencing active extrusion of Li⁺.     

Removal by a Trace of Sodium of the Period Lengthening of the Potassium Uptake Rhythm Due to Lithium in Lemna gibba G3

The effect of Li⁺ on the period of the K⁺ uptake rhythm in the flow medium culture of the duckweed (Lemna gibba G3) was investigated under various ionic conditions. In the presence of Li⁺ at 0.2 millimolar or higher concentrations, the period was longer than the normal level of 25.4 hours by 2 hours. Li⁺ also lowered the amplitude of the rhythm. Although Na⁺ itself did not change any parameter of the rhythm, simultaneous application of Na⁺ at a very low concentration (20 micromolar) almost completely removed the effects of 0.5 millimolar Li⁺ on both the period and the amplitude. However, divalent cations (Ca²⁺ and Mg²⁺) or Rb⁺ did not remove the Li⁺ action on the period. The effect of Li⁺ and its removal by Na⁺ corresponded to intracellular Li⁺ and Na⁺ levels. The period was prolonged when the duckweed contained more Li⁺ than 5 micromoles/gram fresh weight. But the Li⁺ effect was cancelled when the in vivo Na⁺ level was greater than one-fifth that of Li⁺, even if the Li⁺ level exceeded over 5 micromoles/gram fresh weight.     

Effect of potassium and lithium ions on protein synthesis in the sea urchin embryo

The effects of K⁺ and Li⁺, alone and in combination, on protein synthesis were determined in two species of sea urchin, from fertilization through gastrulation. Embryos treated with Li⁺ show a decline in rates of [¹⁴C]valine uptake and incorporation beginning after cleavage. Li⁺-induced cecline in protein synthesis is reversed by co-exposure to increasing concentrations of K⁺. K⁺ alone is without effect on protein synthesis. K⁺ is effective in counteracting the effects of Li⁺ only if both ions are present concurrently. The data presented supports the hypothesis that K⁺ permits normal morphological development in the presence of Li⁺ by competitively blocking the effects of Li⁺ at the subcellular level.     

Effects of lithium and potassium on recovery of solute uptake capacity of Acer pseudoplatanus cells after gas-shock

At concentrations of 10-^?3M, Li⁺ inhibits the recovery of solute uptake capacity of Acer pseudoplatanus L. cell suspension cultures after gas-shock (i.e. after rapid exchange of the atmosphere in the culture flasks for ambient air). It also reduces solute uptake capacity of cells having already attained high rates of uptake during recovery from gas-shock. The effects of Li⁺ are much greater in cells which have been cultivated in 7 mM K⁺ solution than in cells cultivated with higher K⁺ levels (19 mM). Increasing K⁺ concentration during recovery reverses the effect of 10^–3M Li⁺ and, with sufficiently high concentrations of K⁺ (≥ 10-^?2M) during recovery, the solute uptake capacity of the fully recovered cells can even become greater than that of the control, at least for the low values of substrate concentration (here sulphate 10-^?5M). Since Li⁺ does not affect the time course of solute uptake measured over 15–20 min, it is thought that it interacts with the synthesis and turnover of the solute uptake machinery of the Acer pseudoplatanus cells. Thermodynamic analysis of the flux data also supports the hypothesis that Li⁺ inhibits the biosynthesis of specific sites of solute permeation, but it does not rule out the possibility that K⁺ interferes rather on the forces acting on the transport of the considered solutes than on the catalytic structures of permeation.     

Phosphate uptake by phosphate-starved Euglena

Phosphate-deprived Euglena acquire the ability to rapidly in-corporate added phosphate and, also, synthesize an induced acid phosphatase localized in the pellicle. The phosphate uptake system is saturated at low concentrations of phosphate and is inhibited by dinitrophenol, by low temperature, by K⁺, Li⁺, and Na⁺ ions, and competitively by arsenate. The orthophosphate incorporated into the cell is rapidly converted into organic forms but enough remains unesterified to suggest that the uptake is an active transport process. The data do not rule out the possibility that the induced phosphatase is involved in the transport process.     

Microsomal (Na + K)-activated ATPase from frog skin epithelium. Cation activations and some effects of inhibitors

A method is described for the extraction of microsomal ouabain-sensitive (Na⁺ + K⁺)-activated ATPase from separated frog skin epithelium. The method yields a microsomal fraction containing (Na⁺ + K⁺)-stimulated activity in the range of 30–40 nmol · mg⁻¹ · min⁻¹ at 26 °C. This portion, which is also ouabain sensitive, is about half of the total activity in media containing Mg²⁺, Na⁺ and K⁺. These preparations also contain Mg²⁺-dependent or Ca²⁺-dependent activities which are not additive and which are not significantly affected by ouabain, Na⁺, K⁺ or Li⁺.The activations of the ouabain-sensitive ATPase activity by Mg²⁺, Na⁺, and K⁺ are similar to those described in other tissues. It is found that Li⁺ does not substitute for Na⁺ as an activator but in high concentrations does produce partial activation in the presence of Na⁺ with no K⁺. These results are pertinent to the reported observations of ouabain-sensitive Li⁺ flux across frog skin. It is concluded that this flux is not apparently due to a direct activating effect of Li⁺ on the sodium pump.     

Lithium treatment of affective disorders: effects of lithium on the inositol phospholipid and cyclic AMP signalling pathways

The effects of lithium (Li⁺) on the adenylyl cyclase and inositol phospholipid receptor signalling pathways were compared directly in noradrenergic and carbachol stimulated rat brain cortical tissue slices. Li⁺ was a comparatively weak inhibitor of noradrenaline-stimulated cyclic AMP accumulation with an IC₅₀ of approx. 20 mM. By contrast, half-maximal effects of Li⁺ on inositol monophosphate (InsP) accumulation in [³H]inositol labelled tissue slices occurred at about 1 mM. A similar IC₅₀ for Li⁺ of about 1 mM was also obtained for noradrenaline-stimulated accumulation of CMP-phosphatidate (CMPPA), a sensitive indicator of intracellular inositol depletion, in tissue slices that had been prelabelled with [³H]cytidine. The effect of myo-inositol (inositol) depletion on the prolonged activity of phosphoinositidase C (PIC) was examined in carbachol-stimulated corticol slices using a novel mass assay fro InsP. Exposure to a maximal dose of carbachol for 30 min in the presence of 5 mM Li⁺ caused a 10-fold increase in the level of radioactivity associated with the InsP fraction, but only a 2-fold increase in InsP mass. During prolonged incubations in the presence of both carbachol and Li⁺ the accumulation of InsP mass was enhanced if 30 mM inositol was included in the medium. The results are comptable with the inositol depletion hypothesis of Li⁺ action but do not support the concept that adenylyl cyclase or guanine nucleotide dependent proteins represent therapeutically relevant targets of this drug.     

THE UPTAKE OF LOW CONCENTRATIONS OF LITHIUM IONS INTO RAT CEREBRAL CORTEX SLICES AND ITS DEPENDENCE ON CATIONS

—Rat cerebral slices were incubated in oxygenated Krebs-Ringer bicarbonate glucose saline, and the uptake of Li⁺ was measured after periods of 15 s to 5 min. Saturation was not seen within the concentrations of Li⁺ employed (0·5-2·0 mm ). The half-time of the uptake was 7·9 min. At steady state, after 1 h incubation, the concentration of Li⁺ in the tissue was linearly related to that of the medium (0·5-1·5 mm Li⁺) with a concentration ratio of 1·29–1·66. The concentrations of K⁺ and Na⁺ in the slices incubated without Li⁺ were found to be (μmol/g incubated wt, mean ±s.d .) 63·8 ± 9·6 and 96·2 ± 7·8 respectively (n = 28). In the presence of media with 1·5 mm -Li⁺, the K⁺ and Na⁺ in the slices were 56·2 ± 8·8 and 101·0 ± 7·7 respectively (n = 37). The concentration of Li⁺ in the slices, after 1 h incubation, increased in a non linear way as the concentration of K⁺ in the medium was decreased within a range of 0·10 mm -K⁺. In the absence of K⁺ in the medium the uptake of Li⁺ was approx 50% higher than in the presence of 4·9 mm -K⁺. There was an inverse linear relationship between the concentration of Li⁺ in the slices and that of Ca²⁺ in the medium within the range of 0-5·2 mm (-0·13 mm -Li⁺/mm Ca²⁺). The concentration of Li⁺ in the slices increased by approx 10% when the Mg²⁺ in the medium was increased from 1·3 mm to 2·6 mm . Changes of the concentration of Na⁺ between 120 mm and 170 mm in the medium had no significant effect on the Li⁺ uptake.     

Intersubunit bridging by Na+ ions as a rationale for the unusual stability of the c-rings of Na+-translocating F1F0 ATP synthases

The oligomeric c-rings of Na⁺-translocating F₁F₀ ATP synthases exhibit unusual stability, resisting even boiling in SDS. Here, we show that the molecular basis for this remarkable property is intersubunit crossbridging by Na⁺ or Li⁺ ions. The heat stability of c₁₁ was dependent on the presence of Na⁺ or Li⁺ ions. For equal stability, 10 times higher Li⁺ than Na⁺ concentrations were required, reflecting the 10 times lower binding affinity for Li⁺ than for Na⁺. In a recent structural model of c₁₁, the Na⁺ or Li⁺ binding ligands are located on neighboring c-subunits, which thus become crossbridged by the binding of either alkali ion with a concomitant increase in the stability of the ring. Site-directed mutagenesis strengthens the essential role of glutamate 65 in the crossbridging of the subunits and also corroborates the proposed stabilizing effect of an ion bridge including aspartate 2.     

Lithium induced changes in intracellular free magnesium concentration in isolated rat ventricular myocytes

We have examined the effect of exposing isolated rat ventricular myocytes to lithium while measuring cytosolic free magnesium ([Mg²⁺]_i) and calcium ([Ca²⁺]_i) levels with the fluorescent, ion sensitive probes mag-fura-2 and fura-2. There was a significant rise in [Mg²⁺]_i after a 5 min exposure to a solution in which 50% of the sodium had been replaced by Li⁺, but not when the sodium had been replaced by bis-dimethylammonium (BDA). However, there were significant increases in [Ca²⁺]_i when either Na⁺ substitute was used. The possibility that Li⁺, which enters the cells, interferes with the signal from mag-fura-2 was eliminated as Li⁺ concentrations up to 10 mM had no effect on the dye's fluorescence signal. A possible explanation for these findings is that Li⁺ displaces Mg²⁺ from intracellular binding sites. Having considered the binding constants for Mg²⁺ and Li⁺ to ATP, we conclude that Li⁺ can displace Mg²⁺ from Mg-ATP, thus causing a rise in [Mg²⁺]_i. This work has implications for other studies where Li⁺ is used as a Na⁺ substitute.     

Lithium Transport in the Kidney of the Herring Gull Larus argentatus: Induction of Renal Li+ Net Secretion by a Salt Loading

To find out whether salt loading can induce Li⁺ net secretion in the kidney of the birds that have an extrarenal organ (the salt gland) for excretion of NaCl excess (earlier, such effect was revealed in the pigeon and chicken, the birds that do not have the salt gland), the effect of intravenous NaCl injections (4–5 injections of 14–20 mmol/kg at 20–30-min intervals) on Li⁺ transport in the gull kidney was studied. Prior to the salt loading, above 99% of the filtered Li⁺ were reabsorbed in the kidney: fractional excretion of Li⁺ (FE_Li) was 0.0024 ± 0.0007 (mean ± SD), the Li⁺ reabsorption occurring not only in the proximal, but also in the distal tubule. Under conditions of the salt loading, two essentially different regimes of the kidney functioning were observed: net Li⁺ reabsorption (FE_Li = 0.63 ± 0.26) and net Li⁺ secretion (FE_Li = 1.26 ± 0.12). In the absence of the salt loading, Li⁺ (due to its distal reabsorption) does not fit requirements of an indicator of proximal reabsorption of Na⁺ and water. However, in the regimes of the salt net reabsorption and the salt net secretion, FE_Li probably can serve as an indicator of delivery of these substances to the end of the proximal tubule (the lithium clearance method). If this suggestion is correct, transition from the net Li⁺ reabsorption (FE_Li < 1) to its net secretion (FE_Li > 1) with rise of the salt loading indicates the appearance of net Na⁺ secretion in the proximal tubule under these conditions. A comparison of the results for the gull and the chicken has shown that although the presence of the salt gland did not prevent transition of the kidney to the net Li⁺ secretion, its duration in the gull was significantly shorter. The comparative data are presented on parameters of renal function in the regimes of net Li⁺ secretion and net Li⁺ reabsorption. In both regimes, a linear correlation was observed between FE_Li and FE_Na; however, regression coefficients considerably differed. An abrupt break of the curve occurred at FE_Li 1.     

Ability of monovalent cations to replace potassium during stimulation of lymphoid cells

Stimulation of hamster thymocytes, splenocytes, or lymph node cells occurred to a minimal extent in the absence of K⁺. This observation was found for stimulation by T-cell mitogens (phytohemagglutinin and concanavalin A), A B-cell mitogen (lipopolysaccharide), or antigen (KLH). Marginal restoration of the responses to these stimulants occurred in the presence of 0.1 mM K⁺ and responsiveness returned to near maximal levels on addition of 1 mM K⁺ to the cultures. Attempts to restore the responsiveness with other monovalent cations revealed an order of effectiveness of K⁺ ≥ Rb⁺ ? NH₄⁺ ≥ Li⁺. At the 1 mM level K⁺ and Rb⁺ were equally effective in supporting stimulation by phytohemagglutinin while all concentrations of Li⁺ tested (0.1–10 mM) would not support stimulation. However, addition of Li⁺ to cultures reconstituted with 1 mM K⁺ or Rb⁺ revealed that this ion could enhance the phytohemagglutinin response by approximately 100% in the presence of K⁺ and only 30% in the presence of Rb⁺. These data support the hypotheses that the Na,K ATPase must be active for lymphocyte stimulation to occur and that some of the biological effects of Li⁺ on lymphocyte stimulation are mediated at the level of the Na,K ATPase.     

L'induction du rythme endogène de sporulation chez Aspergillus niger: Rõles du rapport glucose/potassium et de micro-éléments

In Aspergillus niger Van Tieghem cultivated on a synthetic medium, the induction of an endogenous rhythm of sporulation and its perpetuation depend on the glucose K⁺ balance in the medium, an excess of one of them suppressing the oscillations. In its inducing effect on the rhythm K⁺ is partially replaced by Rb⁺, but not by Na⁺, Li⁺ or Cs⁺. While the glucose K⁺ balance is favourable for the manifestation of the rhythm, the addition of increasing levels of Na⁺, Li⁺ or Cs⁺ do not modify the period length. Nevertheless, at 0.3 M of Na⁺ or Li⁺ and 0.03 M of Cs⁺ rhythm disappears. The amplitude of oscillations depends on the level of the micro-elements furnished, especially on Mn²⁺. EDTA (1 × 10^?3M) inhibits the rhythm.     

Effects of alkali ions on the circadian leaf movements of Oxalis regnellii

The influence of alkali ions on the circadian leaf movements of Oxalis regnellii Mig. was investigated. Ions were given to the oscillating system via the transpiration stream of cut stalks in nutrient medium. Chloride solutions of Rb⁺, Cs⁺, Na⁺ and K⁺ were tested and the results compared to previously published LiCl-results. The period of the circadian leaf movements was unaffected by a continual addition of Na⁺ or K⁺ to the nutrient medium (at least up to 40 mM). Rb⁺, in the concentration of 2.5 or 5 mM, caused a shortening of the period when applied continuously. Rb⁺ concentrations up to 60 mM were tested. Cs⁺ ions caused only lengthenings of the circadian period. Cs⁺ concentrations up to 40 mM were tested. Cs⁺ resembled Li⁺ in producing period lengthenings, but was not as effective as Li⁺ when compared on a concentration basis. Toxicity of the effective ions was in the following order: Li⁺Cs⁺Rb⁺, Rb⁺ pulses (50 mM, 4 h) phase-shifted the rhythm and caused advances. A phase response curve was determined and the maximum steady state advances were of the order of 1 h. The dual effect of the Rb⁺ ions is discussed and is assumed to be due to two counteracting processes, exemplified by Rb⁺-sensitive ATPase-controlled pumping processes and protein synthesis. For comparison, the effects of Rb⁺ and Li⁺ in human depressive disorders is also discussed in relation to their influence on circadian systems. It is emphasized that Rb⁺ and K⁺ behave differently and are not interchangeable in their action on circadian systems.     

On the mechanism of action of polyether XXVIII at site I of the electron-transfer chain in rat liver mitochondria

In rat liver mitochondria, the macrocyclic polyether, dibenzo-18-crown-6 (polyether XXVIII) inhibits the oxidation of NAD-dependent substrates, as stimulated by ADP, uncouplers and valinomycin plus K⁺. It does not inhibit the oxidation of succinate. It is concluded that polyether XXVIII inhibits electron transfer in the NADH-CoQ span of the respiratory chain. This is a process that is reversed by menadione. Inhibition of oxidation of NAD-dependent substrates in K⁺-depleted mitochondria induced by the polyether is reversed by concentrations of K⁺ higher than 60 mM, and also by Li⁺, a cation that does not complex with polyether XXVIII. As assayed by swelling mitochondria, reversal of the inhibition of electron transfer is accompanied by influx of monovalent cations. Polyether XXVIII also inhibits in submitochondrial particles the aerobic oxidation of NADH, but not that of succinate; this inhibition is also reversed by K⁺ at high concentrations, and Li⁺. The data are consistent with the hypothesis that a monovalent cation is required for maximal rates of electron transport in the NADH-CoQ span of the respiratory chain.     

The Na+/H+ Exchanger Present in Trout Erythrocyte Membranes is Not Responsible for the Amiloride-insensitive Na+/Li+ Exchange Activity

The protein responsible for the Na⁺/Li⁺ exchange activity across the erythrocyte membrane has not been cloned or isolated. It has been suggested that a Na⁺/H⁺ exchanger could be responsible for the Na⁺/Li⁺ exchange activity across the erythrocyte membrane. Previously, we reported that in the trout erythrocyte, the Li⁺/H⁺ exchange activity (mediated by the Na⁺/H⁺ exchanger βNHE) and the Na⁺/Li⁺ exchange activity respond differently to cAMP, DMA (dimethyl-amiloride) and O₂. We concluded that the DMA insensitive Na⁺/Li⁺ exchange activity originates from a different protein. To further examine these findings, we measured Li⁺ efflux in fibroblasts expressing the βNHE as the only Na⁺/H⁺ exchanger. Moreover, the internal pH of these cells was monitored with a fluorescent probe. Our findings indicate that acidification of fibroblasts expressing the Na⁺/H⁺ exchanger βNHE, induces a Na⁺ stimulated Li⁺ efflux activity in trout erythrocytes. This exchange activity, however, is DMA sensitive and therefore differs from the DMA insensitive Na⁺/Li⁺ exchange activity. In these fibroblasts no significant DMA insensitive Na⁺/Li⁺ exchange activity was found. These results support the hypothesis that the trout erythrocyte Na⁺/Li⁺ exchange activity is not mediated by the Na⁺/H⁺ exchanger (βNHE) present in these membranes. Received: 6 December 1996/Revised: 11 August 1997