Intracellular Sodium Concentration Research Articles

Two sides of the same coin: Sodium homeostasis and signaling in astrocytes under physiological and pathophysiological conditions

The intracellular sodium concentration of astrocytes is classically viewed as being kept under tight homeostatic control and at a relatively stable level under physiological conditions. Indeed, the steep inwardly directed electrochemical gradient for sodium, generated by the Na⁺/K⁺-ATPase, contributes to maintain the electrochemical gradient of K⁺ and the highly K⁺-based negative membrane potential, and is a central element in energizing membrane transport. As such it is tightly coupled to the homeostasis of extra- and intracellular potassium, calcium or pH and to the reuptake of transmitters such as glutamate. Recent studies, however, have demonstrated that this picture is far too simplistic. It is now firmly established that transmitters, most notably glutamate, and excitatory neuronal activity evoke long-lasting sodium transients in astrocytes, the properties of which are distinctly different from those of activity-related glial calcium signals. From these studies, it emerges that sodium homeostasis and signaling are two sides of the same coin: sodium-dependent transporters, primarily known for their role in ion regulation and homeostasis, also generate relevant ion signals during neuronal activity. The functional consequences of activity-related sodium transients are manifold and are just coming into view, enabling surprising and important new insights into astrocyte function and neuron-glia interaction in the brain. The present review will highlight current knowledge about the mechanisms that contribute to sodium homeostasis in astrocytes, present recent data on the spatial and temporal properties of activity-related glial sodium signals and discuss their functional consequences with a special emphasis on pathophysiological conditions.

Angiotensin II rapidly stimulates the short‐circuit current in opossum kidney cells expressing rat Na,K‐ATPase

The binding of angiotensin II (Ang II) to the angiotensin receptor (AT1) stimulates Na,K‐ATPase activity that drives sodium reabsorption in the proximal tubule, which reabsorbs 2/3 of the filtered sodium. To determine how quickly Ang II can stimulate Na,K‐ATPase in the proximal tubule, the α‐1 subunit of rat kidney Na,K‐ATPase and the AT1a receptor were stably co‐expressed in opossum kidney cells. Cells were grown on permeable supports and mounted in an Ussing Chamber containing Ringer's solution. The addition of 10 μM Ang II to the basolateral side increased the short‐circuit current (Isc), which is proportional to net sodium transport, in ≤ ~10 sec. Isc peaked at ~20 sec and declined to a plateau above baseline for over 3 min. The subsequent addition of 200 μM nystatin to increase the sodium permeability of the apical membrane, and thereby the concentration of intracellular sodium, increased Isc to the level of the Ang II‐dependent peak. Subsequently adding 600 μM ouabain, which inhibits Na,K‐ATPase, reduced Isc to its original baseline value. Thus, Ang II significantly stimulated Na,K‐ATPase activity and transcellular sodium transport in 10 seconds or less. We hypothesize that such rapid Ang II‐dependent stimulation of sodium reabsorption in the proximal tubule is part of a mechanism to help the body to recover from a sudden drop in blood pressure. Supported in part by HL096800 to F.S.

A specific and essential role for Na,K‐ATPase α3 in neurons co‐expressing α1 and α3

Most neurons co‐express two catalytic isoforms of Na,K‐ATPase, the ubiquitous α1, and the more selectively expressed α3. Although neurological syndromes are associated with α3 mutations, the specific role of this isoform is not completely understood. Here we used electrophysiology and sodium imaging to study the role of α3 in neurons expressing both isoforms. Under basal conditions, selective inhibition of α3 using a low concentration of the cardiac glycoside ouabain resulted in a modest increase in the intracellular sodium concentration ([Na+]i), accompanied by membrane potential depolarization. When neurons were challenged with a large rapid increase in [Na+]i, similar to what could be expected following suprathreshold neuronal activity, inhibition of α3 almost completely abolished the capacity to restore [Na+]i in dendrites. Recordings of Na,K‐ATPase‐specific current supported the notion that α3 is the predominant isoform, responsible for rapid extrusion of Na+. Low concentrations of ouabain were also found to disrupt cortical network oscillations, providing further support for the importance of α3 function in the central nervous system. The α isoforms express a well conserved protein kinase A (PKA) consensus site, which is structurally associated with a Na+ binding site. Activation of PKA significantly attenuated both the α3 dependent current and restoration of dendritic [Na+]i, indicating that α3 is a target for phosphorylation and may participate in short‐term regulation of neuronal function.

Application of Mathematical Modeling for Analysis of Experimental Data on Cell Water-Electrolyte Balance

The study of osmotic and ion balance is of particular importance for transporting epithelial cells that have to maintain stable intracellular medium and cell volume upon intensive transcellular osmolyte and water fluxes. We propose an approach based on mathematical modeling that allows quantitative estimation of the rate of transmembrane ion transport in rat renal collecting duct principal cells. Our goals were:1) to estimate membrane permeabilities for sodium, potassium and chloride ions;2) to estimate rates of passive transmembrane fluxes for sodium, potassium and chloride ions and the flux through Na/K-pump;3) to assess the activity of KCC and NKCC cotransporters in the cells under examination.The mathematical model describing transmembrane ion fluxes was used to analyze the results of experimental measurements of cell volume and intracellular sodium concentration dynamics. The use of high-performance calculations on computer cluster made it possible to obtain the correspondence between the values of model permeability parameters and experimentally measured cell physiological characteristics.Quantitative estimates of permeability parameters and the rates of transmembrane fluxes were obtained for renal collecting duct principal cells. Also the model made it possible to detect the decrease in sodium and potassium membrane permeability of collecting duct cells from animals kept on high salt diet.This approach sets grounds for interpretation of experimental data to estimate cell physiological characteristics whose accurate experimental measurement is a hardly achievable goal.

Regulation of the sodium pump during cardiomyocyte adaptation to pregnancy

Regulation of the sodium pump during normal pregnancy and its effect on the function of cardiomyocytes is poorly understood. Our objective was to evaluate the possible implication of the Na(+)-K(+)-ATPase, the sodium pump which controls cellular ionic and metabolic homeostasis, in the adaptations of cardiomyocytes to normal pregnancy. We have used Western blots and patch-clamp measurements to identify changes in the sodium pump proteins. Confocal microscopy was applied to estimate intracellular sodium concentration. Time-resolved spectroscopy was employed to measure mitochondrial NAD(P)H fluorescence and estimate oxidative metabolic state. Optical microscopy was adopted to study the contractility responses of cardiomyocytes. Cells from non-pregnant and pregnant rats (1 day prior parturition) were studied. Our results showed lower protein expression of the α1 Na(+)-K(+)-ATPase isoform in cardiomyocytes in pregnant rats, decreased sodium pump membrane current and elevated steady-state sodium concentration. In addition, ouabain, the inhibitor of the sodium pump capable of increasing cardiomyocyte contractility in non-pregnant rats in a concentration-dependent manner, failed to affect cell contractions in pregnant rats. We also noted modified responsiveness of the mitochondrial metabolic state to ouabain in cardiac cells. The gathered data confirmed that in pregnant rats, the sodium pump protein content and transmembrane flux are decreased, while the sensitivity of cardiomyocyte contractility and the sensitivity of mitochondrial metabolic redox state to ouabain are modified, pointing to regulation of the Na(+)-K(+)-ATPase during cardiac cell adaptations to normal pregnancy.

A novel SGLT is expressed in the human kidney

Selective inhibitors of sodium-glucose cotransporter 2 (SGLT2)-mediated reabsorption of glucose in the proximal tubule of the kidney are being developed for the treatment of diabetes. SGLT2 shares high degree of homology with SGLT3; however, very little is known about the expression and functional role of SGLT3 in the human kidney. Indeed, the SGLT2 inhibitors that are currently in clinical trials might affect the expression and/or the activity of SGLT3. Therefore, the present study examined the expression of SGLT3 mRNA and protein in human kidney and in a human proximal tubule HK-2 cell line. The results indicated that human SGLT3 (hSGLT3) message and protein are expressed both in vivo and in vitro. We also studied the activity of hSGLT3 protein following its over-expression in mammalian kidney-derived COS-7 cells and in HK-2 cells treated with the imino sugar deoxynojirimycin (DNJ), a potent agonist of hSGLT3. Over-expression of hSGLT3 in COS-7 cells increased intracellular sodium concentration by 3-fold without affecting glucose transport. Activation of hSGLT3 with DNJ (50μM) increased sodium uptake in HK-2 cells by 5.5 fold and this effect could be completely blocked with SGLT inhibitor phlorizin (50μM). These results suggest that SGLT3 is expressed in human proximal tubular cells where it serves as a novel sodium transporter. Up-regulation of the expression of SGLT3 in the proximal tubule in diabetic patients may contribute to the elevated sodium transport in this segment of the nephron that has been postulated to promote hyperfiltration and renal injury.

Noninvasive quantification of intracellular sodium in human brain using ultrahigh–field MRI

In vivo sodium magnetic resonance imaging (MRI) measures tissue sodium content in living human brain but current methods do not allow noninvasive quantitative assessment of intracellular sodium concentration (ISC) - the most useful marker of tissue viability. In this study, we report the first noninvasive quantitative in vivo measurement of ISC and intracellular sodium volume fraction (ISVF) in healthy human brain, made possible by measuring tissue sodium concentration (TSC) and intracellular sodium molar fraction (ISMF) at ultra-high field MRI. The method uses single-quantum (SQ) and triple-quantum filtered (TQF) imaging at 7 Tesla to separate intra- and extracellular sodium signals and provide quantification of ISMF, ISC and ISVF. This novel method allows noninvasive quantitative measurement of ISC and ISVF, opening many possibilities for structural and functional metabolic studies in healthy and diseased brains.

Towards deciphering dynamic changes and evolutionary mechanisms involved in the adaptation to low salinities in Ectocarpus (brown algae).

Colonizations of freshwater by marine species are rare events, and little information is known about the underlying mechanisms. Brown algae are an independent lineage of photosynthetic and multicellular organisms from which few species inhabit freshwater. As a marine alga that is also found in freshwater, Ectocarpus is of particular interest for studying the transition between these habitats. To gain insights into mechanisms of the transition, we examined salinity tolerance and adaptations to low salinities in a freshwater strain of Ectocarpus on physiological and molecular levels. We show that this isolate belongs to a widely distributed and highly stress-resistant clade, and differed from the genome-sequenced marine strain in its tolerance of low salinities. It also exhibited profound, but reversible, morphological, physiological, and transcriptomic changes when transferred to seawater. Although gene expression profiles were similar in both strains under identical conditions, metabolite and ion profiles differed strongly, the freshwater strain exhibiting e.g. higher cellular contents of amino acids and nitrate, higher contents of n-3 fatty acids, and lower intracellular mannitol and sodium concentrations. Moreover, several stress markers were noted in the freshwater isolate in seawater. This finding suggests that, while high stress tolerance and plasticity may be prerequisites for the colonization of freshwater, genomic alterations have occurred that produced permanent changes in the metabolite profiles to stabilize the transition.

Increases in intracellular sodium activate transcription and gene expression via the salt-inducible kinase 1 network in an atrial myocyte cell line

Cardiac hypertrophy (CH) generally occurs as the result of the sustained mechanical stress caused by elevated systemic arterial blood pressure (BP). However, in animal models, elevated salt intake is associated with CH even in the absence of significant increases in BP. We hypothesize that CH is not exclusively the consequence of mechanical stress but also of other factors associated with elevated BP such as abnormal cell sodium homeostasis. We examined the effect of small increases in intracellular sodium concentration ([Na(+)](i)) on transcription factors and genes associated with CH in a cardiac cell line. Increases in [Na(+)](i) led to a time-dependent increase in the expression levels of mRNA for natriuretic peptide and myosin heavy chain genes and also increased myocyte enhancer factor (MEF)2/nuclear factor of activated T cell (NFAT) transcriptional activity. Increases in [Na(+)](i) are associated with activation of salt-inducible kinase 1 (snflk-1, SIK1), a kinase known to be critical for cardiac development. Moreover, increases in [Na(+)](i) resulted in increased SIK1 expression. Sodium did not increase MEF2/NFAT activity or gene expression in cells expressing a SIK1 that lacked kinase activity. The mechanism by which SIK1 activated MEF2 involved phosphorylation of HDAC5. Increases in [Na(+)](i) activate SIK1 and MEF2 via a parallel increase in intracellular calcium through the reverse mode of Na(+)/Ca(2+)-exchanger and activation of CaMK1. These data obtained in a cardiac cell line suggest that increases in intracellular sodium could influence myocardial growth by controlling transcriptional activation and gene expression throughout the activation of the SIK1 network.

Magnesium and Vascular Changes in Hypertension

Many factors have been implicated in the pathogenesis of hypertension, including changes in intracellular concentrations of calcium, sodium, potassium, and magnesium. There is a significant inverse correlation between serum magnesium and incidence of cardiovascular diseases. Magnesium is a mineral with important functions in the body such as antiarrhythmic effect, actions in vascular tone, contractility, glucose metabolism, and insulin homeostasis. In addition, lower concentrations of magnesium are associated with oxidative stress, proinflammatory state, endothelial dysfunction, platelet aggregation, insulin resistance, and hyperglycemia. The conflicting results of studies evaluating the effects of magnesium supplements on blood pressure and other cardiovascular outcomes indicate that the action of magnesium in the vascular system is present but not yet established. Therefore, this mineral supplementation is not indicated as part of antihypertensive treatment, and further studies are needed to better clarify the role of magnesium in the prevention and treatment of cardiovascular diseases.

Effects of Nifedipine on Cation Transport and Na - K-ATPase Activity in Erythrocytes and Electrolyte Homeostasis in Rats

Calcium channel antagonists have been reported to reduce blood pressure in those individuals at risk of cardiac and cerebrovascular events. There is no specific study available regarding the role of electrolyte alterations in blood pressure lowering effects of nifedipine. The present study was designed to investigate the role of electrolyte homeostasis, changes in ouabain-sensitive Na+ K+ adenosine triphosphatase (ATPase) activity, and net sodium efflux and potassium influx across blood cell membranes. Rats were divided into two experimental groups . Nifedipine (20mg/ kg body weight ) was administered by gastric tube to the test group . Control group received same volume of deionize water .The intra-erythrocyte sodium, serum sodium, potassium, calcium and sodium, calcium content of heart and kidney tissues were decreased significantly. Whereas, intra-erythrocyte potassium was slightly decreased or remained normal in nifedipine treated rats as compared to normal healthy rats. The Na - K - ATPase activity, serum magnesium, potassium and magnesium content in heart and kidney tissues were increased significantly. Results confirmed that nifedipine represses ion channels, transporters and calcium-binding proteins in tissues. Erythrocyte studies indicate that nifedipine blocks the entrance of calcium into the cells but also stimulate Na - K - ATPase activity, resulting in reduction of intracellular sodium concentration, thus suggesting direct nifedipine-induced blood pressure reduction.

3 Tesla Sodium Inversion Recovery Magnetic Resonance Imaging Allows for Improved Visualization of Intracellular Sodium Content Changes in Muscular Channelopathies

To implement different sodium (²³Na)-magnetic resonance imaging (MRI) contrasts at 3 Tesla and to evaluate if a weighting toward intracellular sodium can be achieved, using 2 rare muscular channelopathies as model diseases. Both lower legs of 6 patients with hypokalemic periodic paralysis (HypoPP), 5 patients with paramyotonia congenita (PC), and 5 healthy volunteers were examined on a 3 Tesla system with 3 different ²³Na-MRI pulse sequences. HypoPP and PC are rare muscle diseases, which are well characterized by elevated myoplasmic sodium at rest and after cooling, respectively. Intra- and interindividual comparisons were performed before and after provocation of one lower leg muscle. Three different ²³Na-MRI sequences were applied: (i) The total tissue sodium concentration was measured using a spin-density sequence (²³Na-TSC). (ii) A T1-contrast was applied to assess whether the known changes of the intracellular sodium concentration can be visualized by T1-weighting (²³Na-T1). (iii) An inversion recovery (²³Na-IR) sequence was used to utmost suppress the sodium signal from extracellular or vasogenic edema. Furthermore, a potential influence of the temperature dependency of the sodium relaxation times was considered. Additionally, H-MRI was performed to examine potential lipomatous or edematous changes. In HypoPP, all Na sequences showed significantly (P<0.05) higher signal intensities compared with PC patients and healthy subjects. In muscles of PC patients, provocation induced a significant (P=0.0007) increase (>20%) in the muscular ²³Na-IR signal and a corresponding decrease of muscle strength. Additionally, a tendency to higher ²³Na-T1 (P=0.07) and ²³Na-TSC (P=0.07) signal intensities was observed. Provocation revealed no significant changes in ¹H-MRI. In volunteers and in the contralateral, not cooled lower leg, there were no significant signal intensity changes after provocation. Furthermore, the ²³Na-IR sequence allows for a suppression of signal emanating from intravascular sodium and vasogenic edema. Our results indicate that the ²³Na-IR sequence allows for a weighting toward intracellular sodium. The combined application of the ²³Na-TSC and the ²³Na-IR sequence enables an improved analysis of pathophysiological changes that occur in muscles of patients with muscular channelopathies.

Bilirubin Attenuates Bufadienolide-Induced Ventricular Arrhythmias and Cardiac Dysfunction in Guinea-Pigs by Reducing Elevated Intracellular Na+ Levels

Bufadienolides, known ligands of the sodium pump, have been shown to inhibit the proliferation of several cancer cell types. However, their development to date as anticancer agents has been impaired by a narrow therapeutic margin resulting from their potential to induce cardiotoxicity. In the present study, we examined the effects of bilirubin, an endogenous antioxidant, on the cardiotoxicity of bufadienolides (derived from toad venom) in guinea-pigs. The results showed that bufadienolides (8mg/kg) caused ventricular arrhythmias, conduction block, cardiac dysfunction and death in guinea-pigs. Pretreatment with bilirubin (75 and 150mg/kg) significantly prevented bufadienolide-induced premature ventricular complexes, ventricular tachycardia, ventricular fibrillation and death. Bilirubin also markedly improved the inhibition of cardiac contraction in bufadienolide-treated guinea-pigs as evidenced by increases in left ventricular systolic pressure and decreases in left ventricular diastolic pressure in vivo. Furthermore, bilirubin significantly reduced the intracellular sodium content ([Na(+)]( i )) in ex vivo bufadienolide-stimulated guinea-pig ventricular myocytes loaded with the sodium indicator Sodium Green. An antitumor study showed that bilirubin did not compromise the ability of bufadienolides to inhibit gastric cancer cell MGC-803 proliferation. These results suggested that bilirubin can attenuate bufadienolide-induced arrhythmias and cardiac dysfunction in guinea-pigs by reducing elevated [Na(+)]( i ) and may improve bufadienolide therapeutic index in cancer treatment.

A human ventricular cell model for investigation of cardiac arrhythmias under hyperkalaemic conditions

In this study, several modifications were introduced to a recently proposed human ventricular action potential (AP) model so as to render it suitable for the study of ventricular arrhythmias. These modifications were driven by new sets of experimental data available from the literature and the analysis of several well-established cellular arrhythmic risk biomarkers, namely AP duration at 90 per cent repolarization (APD(90)), AP triangulation, calcium dynamics, restitution properties, APD(90) adaptation to abrupt heart rate changes, and rate dependence of intracellular sodium and calcium concentrations. The proposed methodology represents a novel framework for the development of cardiac cell models. Five stimulation protocols were applied to the original model and the ventricular AP model developed here to compute the described arrhythmic risk biomarkers. In addition, those models were tested in a one-dimensional fibre in which hyperkalaemia was simulated by increasing the extracellular potassium concentration, [K(+)](o). The effective refractory period (ERP), conduction velocity (CV) and the occurrence of APD alternans were investigated. Results show that modifications improved model behaviour as verified by: (i) AP triangulation well within experimental limits (the difference between APD at 50 and 90 per cent repolarization being 78.1 ms); (ii) APD(90) rate adaptation dynamics characterized by fast and slow time constants within physiological ranges (10.1 and 105.9 s); and (iii) maximum S1S2 restitution slope in accordance with experimental data (S(S1S2)=1.0). In simulated tissues under hyperkalaemic conditions, APD(90) progressively shortened with the degree of hyperkalaemia, whereas ERP increased once a threshold in [K(+)](o) was reached ([K(+)](o)≈6 mM). CV decreased with [K(+)](o), and conduction was blocked for [K(+)](o)>10.4 mM. APD(90) alternans were observed for [K(+)](o)>9.8 mM. Those results adequately reproduce experimental observations. This study demonstrated the value of basing the development of AP models on the computation of arrhythmic risk biomarkers, as opposed to joining together independently derived ion channel descriptions to produce a whole-cell AP model, with the new framework providing a better picture of the model performance under a variety of stimulation conditions. On top of replicating experimental data at single-cell level, the model developed here was able to predict the occurrence of APD(90) alternans and areas of conduction block associated with high [K(+)](o) in tissue, which is of relevance for the investigation of the arrhythmogenic substrate in ischaemic hearts.

Levetiracetam exhibits protective properties on rat Schwann cells in vitro

Oxidative stress and inflammation represent pathways causing substantial damage to the peripheral nervous system. Levetiracetam (LEV) is a commonly used antiepileptic drug targeting high-voltage activated N-type calcium channels. Recent evidence suggests that LEV may also act as a histone deacetylase inhibitor, suggesting that this drug exhibits both anti-inflammatory and anti-oxidative effects, and as such may represent an interesting candidate for treating inflammatory diseases affecting the peripheral nerve. Therefore, we analysed the influence of LEV ex vivo on purified Schwann cells from neonatal P3 rats as well as on dorsal root ganglia prepared from E15 rat embryos. LEV diminished a lipopolysaccharide (LPS)-induced increase of the pro-inflammatory signature molecules tumour necrosis factor alpha, matrix metalloproteinase 9 (MMP-9), and caspase 6. Furthermore, LEV decreased LPS-induced cell death and protected cells against oxidative stress in a glutamate-based oxidative stress model. MMP-2 activity, usually elevated during myelination and repair, was also found to be up-regulated following LEV, while LEV exhibited no negative effects on myelination. Intracellular sodium or calcium concentrations were unaltered by LEV. Thus, LEV may be a promising, well-tolerated drug that - besides its antiepileptic potential - mediates anti-inflammatory, anti-oxidative, and anti-apoptotic properties that may potentially be useful in treating diseases of the peripheral nerve.

Sodium influx induced by external calcium chelation decreases human sperm motility

Calcium removal from the medium promptly reduces human sperm motility and induces a Na(+)-dependent depolarization that is accompanied by an increase in intracellular sodium concentration ([Na(+)](i)) and a decrease in intracellular calcium concentration ([Ca(2+)](i)). Sodium loading activates a Na(+)/K(+)-ATPase. Membrane potential (Vm) and [Ca(2+)](i) were simultaneously detected in human sperm populations with the fluorescent probes diSC(3)(5) and fura 2. [Na(+)](i) and was measured independently in a similar fashion using sodium-binding benzofuran isophthalate. Motility was determined in a CASA system, ATP was measured using the luciferin-luciferase assay, and cAMP was measured by radioimmunoassay. Human sperm motility reduction after calcium removal is related to either Na(+)-loading or Na(+)-dependent depolarization, because, under conditions that inhibit the calcium removal-induced Na(+)-dependent depolarization and [Na(+)](i) increase, sperm motility was unaffected. By clamping sperm Vm with valinomycin, we found that the motility reduction associated with the calcium removal was related to sodium loading, and not to membrane potential depolarization. Mibefradil, a calcium channel blocker, markedly inhibited the Na(+)-dependent depolarization and sodium loading, and also preserved sperm motility. In the absence of calcium, both ATP and cAMP concentrations were decreased by 40%. However ATP levels were unchanged when calcium removal was performed under conditions that inhibit the calcium removal-induced Na(+)-dependent depolarization and [Na(+)](i) increase. Human sperm motility arrest induced by external calcium removal is mediated principally by sodium loading, which would stimulate the Na(+)/K(+)-ATPase and in turn deplete the ATP content.

Effects and Mechanisms of Action of Polyene Macrolide Antibiotic Nystatin on Babesia gibsoni In Vitro

Nystatin is a membrane-active polyene macrolide antibiotic and a channel-forming ionophore. Nystatin exhibits in vitro activity against Babesia gibsoni infecting normal canine erythrocytes containing low potassium (LK) and high sodium concentrations, i.e., LK erythrocytes. The calculated IC(50) value of nystatin against B. gibsoni infecting LK erythrocytes was 31.96 µg/ml. The anti-babesial activity of nystatin disappeared when B. gibsoni in LK erythrocytes were incubated in culture media containing high potassium concentrations (HK). Moreover, when the parasites were harbored in canine HK erythrocytes, which contained high potassium and low sodium concentrations as a result of high Na-K-ATPase activity, the in vitro anti-babesial activities of nystatin also disappeared, apparently due to protection by HK erythrocytes. This suggested that nystatin could show in vitro anti-babesial activity against B. gibsoni by its ionophorous activity, the same as other ionophores such as valinomycin. Subsequently, the effects of nystatin on the host cells were observed. Nystatin could not modify the intracellular concentrations of potassium, sodium, adenosine triphosphate, or glucose in either LK or HK erythrocytes, although it caused weak hemolysis in HK erythrocytes. In addition, nystatin did not affect the survival of canine peripheral polymorphonuclear leukocytes. In conclusion, nystatin destroyed B. gibsoni by ionophorous activity but did not affect either canine erythrocytes or leukocytes in vitro.

In vivo magnetic resonance imaging of sodium and diffusion in rat glioma at 21.1 T

Sodium and diffusion magnetic resonance imaging (MRI) in intracranial rat 9L gliomas were evaluated over 6-8 days using the advanced sensitivity of sodium MRI at 21.1 T. Glioma doubling time was 2.4-2.6 days. Glioma sodium signal was detected using the ultra-short echo time of 0.15 ms. The high resolution 3D sodium MRI with pixels of 0.125 μL allowed for minimizing a partial volume effect often relevant to the MRI of low intensity signals. Tumor sodium and diffusion MRI were evaluated for two separate subclones of 9L cells with different resistance to 1,3-bis(2-chloroethyl)-1-nitrosurea detected by pre-surgery assays. In vivo, after implantation, resistant 9L cells created tumors with significantly reduced sodium concentrations (57 ± 3 mM) compared with nonresistant 9L cells (78 ± 3 mM). The corresponding differences in diffusion were less, but also statistically significant. During tumor progression, an increase of glioma sodium concentration was observed in both cell types with a rate of 2.4-5.8 %/day relative to normal brain. Tumor diffusion was not significantly changed at this time, indicative of no alterations in glioma cellularity. Thus, changes in sodium during tumor progression reflect increasing intracellular sodium concentration and mounting metabolic stress. These experiments also demonstrate an enhanced sensitivity of sodium MRI to reflect tumor cell resistance.

Ionic dynamics mediate spontaneous termination of seizures and postictal depression state.

Epileptic seizures are characterized by periods of recurrent, highly synchronized activity that spontaneously terminates, followed by postictal state when neuronal activity is generally depressed. The mechanisms for spontaneous seizure termination and postictal depression remain poorly understood. Using a realistic computational model, we demonstrate that termination of seizure and postictal depression state may be mediated by dynamics of the intracellular and extracellular ion concentrations. Spontaneous termination was linked to progressive increase of intracellular sodium concentration mediated by activation of sodium channels during highly active epileptic state. In contrast, an increase of intracellular chloride concentration extended seizure duration making possible long-lasting epileptic activity characterized by multiple transitions between tonic and clonic states. After seizure termination, the extracellular potassium was reduced below baseline, resulting in postictal depression. Our study suggests that the coupled dynamics of sodium, potassium, and chloride ions play a critical role in the development and termination of seizures. Findings from this study could help identify novel therapeutics for seizure disorder.

Salt-inducible kinase 1 is present in lung alveolar epithelial cells and regulates active sodium transport

Salt-inducible kinase 1 (SIK1) in epithelial cells mediates the increases in active sodium transport (Na+, K+-ATPase-mediated) in response to elevations in the intracellular concentration of sodium. In lung alveolar epithelial cells increases in active sodium transport in response to β-adrenergic stimulation increases pulmonary edema clearance. Therefore, we sought to determine whether SIK1 is present in lung epithelial cells and to examine whether isoproterenol-dependent stimulation of Na+, K+-ATPase is mediated via SIK1 activity. All three SIK isoforms were present in airway epithelial cells, and in alveolar epithelial cells type 1 and type 2 from rat and mouse lungs, as well as from human and mouse cell lines representative of lung alveolar epithelium. In mouse lung epithelial cells, SIK1 associated with the Na+, K+-ATPase α-subunit, and isoproterenol increased SIK1 activity. Isoproterenol increased Na+, K+-ATPase activity and the incorporation of Na+, K+-ATPase molecules at the plasma membrane. Furthermore, those effects were abolished in cells depleted of SIK1 using shRNA, or in cells overexpressing a SIK1 kinase-deficient mutant. These results provide evidence that SIK1 is present in lung epithelial cells and that its function is relevant for the action of isoproterenol during regulation of active sodium transport. As such, SIK1 may constitute an important target for drug discovery aimed at improving the clearance of pulmonary edema.