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Glutamate levels and lifetime in the brain extracellular space are dinamically regulated by a family of Na+- and K+-dependent glutamate transporters, which thereby control numerous brain functions and play a role in numerous neurological and psychiatric diseases. Migraine is a neurological disorder characterized by recurrent attacks of typically throbbing and unilateral headache and by a global dysfunction in multisensory processing. Familial hemiplegic migraine type 2 (FHM2) is a rare monogenic form of migraine with aura caused by loss-of-function mutations in the α2 Na/K ATPase (α2NKA). In the adult brain, this pump is expressed almost exclusively in astrocytes where it is colocalized with glutamate transporters. Knockin mouse models of FHM2 (FHM2 mice) show a reduced density of glutamate transporters in perisynaptic astrocytic processes (mirroring the reduced expression of α2NKA) and a reduced rate of glutamate clearance at cortical synapses during neuronal activity and sensory stimulation. Here we review the migraine-relevant alterations produced by the astrocytic glutamate transport dysfunction in FHM2 mice and their underlying mechanisms, in particular regarding the enhanced brain susceptibility to cortical spreading depression (the phenomenon that underlies migraine aura and can also initiate the headache mechanisms) and the enhanced algesic response to a migraine trigger. [ABSTRACT FROM AUTHOR]

This study describes the changes in ion homeostasis of human endometrial mesenchymal stem/stromal cells (eMSCs) during the formation of three-dimensional (3D) cell structures (spheroids) and investigates the conditions for apoptosis induction in 3D eMSCs. Detached from the monolayer culture, (2D) eMSCs accumulate Na+ and have dissipated transmembrane ion gradients, while in compact spheroids, eMSCs restore the lower Na+ content and the high K/Na ratio characteristic of functionally active cells. Organized as spheroids, eMSCs are non-proliferating cells with an active Na/K pump and a lower K+ content per g cell protein, which is typical for quiescent cells and a mean lower water content (lower hydration) in 3D eMSCs. Further, eMSCs in spheroids were used to evaluate the role of K+ depletion and cellular signaling context in the induction of apoptosis. In both 2D and 3D eMSCs, treatment with ouabain (1 µM) results in inhibition of pump-mediated K+ uptake and severe K+ depletion as well as disruption of the mitochondrial membrane potential. In 3D eMSCs (but not in 2D eMSCs), ouabain initiates apoptosis via the mitochondrial pathway. It is concluded that, when blocking the Na/K pump, cardiac glycosides prime mitochondria to apoptosis, and whether a cell enters the apoptotic pathway depends on the cell-specific signaling context, which includes the type of apoptotic protein expressed. [ABSTRACT FROM AUTHOR]

This paper describes changes in ionic homeostasis associated with premature senescence in human endometrial mesenchymal stem cells (eMSCs). Changes in the intracellular potassium and sodium content and potassium fluxes through the plasma membrane have been examined/ with flame photometry, It was found that, during oxidative stress-induced cell-cycle arrest and the development of premature aging, eMSCs retained the high ionic heterogeneity that is common for functionally active animal cells. Premature cell aging is accompanied by increased intracellular sodium content and transmembrane potassium fluxes associated with the functioning of the Na/K pump, but does not affect the passive transport of potassium across the plasma membrane. One distinctive feature of stress-induced arrested eMSCs is reduced specific intracellular potassium content (500–600 μmol per 1 g of protein) compared to proliferating eMSCs (800–900 μmol per 1 g of protein). It is suggested that decreased intracellular potassium content associated with the development of premature aging shows the participation of potassium ions in the regulation of cell volume and may indicate a decreased hydration of aging eMSCs. [ABSTRACT FROM AUTHOR]

This study describes the changes in ion homeostasis of human endometrial mesenchymal stem/stromal cells (eMSCs) during the formation of three-dimensional (3D) cell structures (spheroids) and investigates the conditions for apoptosis induction in 3D eMSCs. Detached from the monolayer culture, (2D) eMSCs accumulate Na+ and have dissipated transmembrane ion gradients, while in compact spheroids, eMSCs restore the lower Na+ content and the high K/Na ratio characteristic of functionally active cells. Organized as spheroids, eMSCs are non-proliferating cells with an active Na/K pump and a lower K+ content per g cell protein, which is typical for quiescent cells and a mean lower water content (lower hydration) in 3D eMSCs. Further, eMSCs in spheroids were used to evaluate the role of K+ depletion and cellular signaling context in the induction of apoptosis. In both 2D and 3D eMSCs, treatment with ouabain (1 µM) results in inhibition of pump-mediated K+ uptake and severe K+ depletion as well as disruption of the mitochondrial membrane potential. In 3D eMSCs (but not in 2D eMSCs), ouabain initiates apoptosis via the mitochondrial pathway. It is concluded that, when blocking the Na/K pump, cardiac glycosides prime mitochondria to apoptosis, and whether a cell enters the apoptotic pathway depends on the cell-specific signaling context, which includes the type of apoptotic protein expressed.

Magnesium (Mg 2 +) is an essential mineral for several cellular functions. The concentration of this ion below the physiological concentration induces recurrent neuronal discharges both in slices of the hippocampus and in neuronal cultures. These epileptiform discharges are initially sensitive to the application of N -methyl-D-aspartate (NMDA) receptor antagonists, but these antagonists may lose their effectiveness with prolonged exposure to low [Mg 2 + ], when extracellular Ca 2 + reduction occurs, typical of ictal periods, indicating the absence of synaptic connections. The study herein presented aimed at investigating the effect of reducing the [Mg 2 + ] during the induction of Nonsynaptic Epileptiform Activities (NSEA). As an experimental protocol, NSEA were induced in rat hippocampal dentate gyrus (DG), using a bath solution containing high-K + and zero-added-Ca 2 + . Additionally, computer simulations were performed using a mathematical model that represents electrochemical characteristics of the tissue of the DG granular layer. The experimental results show that the reduction of [Mg 2 + ] causes an increase in the duration of the ictal period and a reduction in the interictal period, intensifying epileptiform discharges. The computer simulations suggest that the reduction of the Mg 2 + level intensifies the epileptiform discharges by a joint effect of reducing the surface charge screening and reducing the activity of the Na/K pump. [ABSTRACT FROM AUTHOR]

Background: In recent times, noncommunicable diseases (NCDs) have attained epidemic status in India and several other countries. Globally, each year, NCDs cause premature deaths of nearly 40 million people younger than 60 years accounting for 70% of all deaths; 80% of these premature deaths are reportedly in the low- and middle-income countries. Many studies have reported increasing prevalence of NCDs such as type 2 diabetes, coronary and pulmonary diseases, and cancer. Incidentally, all these studies besides discussing environmental pollution have grossly ignored dietary profiling of affected populace despite the fact that several studies have established a direct correlation between food quality and good health. Objective: Besides pollutants, an attempt is made to analyze mathematically effects of dietary changes over the last four decades on constantly increasing prevalence of NCDs at cellular level. A relationship seems to exist between increasing prevalence of NCD and large-scale consumption of synthetic salt. Conclusions: Elimination of essential trace elements and micro- and macro-minerals in regular diet through synthetic salt fortified with iodine alone appears to cause more harm than benefits associated with iodization of salt than pollutants. Regular intake of essential trace elements is necessary for normal functioning of many fundamental functions of the body such as Na-K pump, electron–proton transport, biochemistry, and thermodynamics. An attempt has been made to study effects of salt constituents used in food at the most fundamental cellular level by means of disturbances in body electrolyte through heat and ion transport mechanism that is fundamental to various underlying processes in human body. Interestingly, almost all the families who switched over to unprocessed rock salt from synthetic iodized salt reported improvement in general health and reported reduction in medical visits.

The capability of the synchronization modulation (SM) technique in enhancing the function of Na/K pumps has been demonstrated in various cells and tissues, including cardiomyocytes, a monolayer of cultured MDCK kidney cells, peripheral blood vessels, and frog skeletal muscles. This study characterized the membrane potential hyperpolarization induced by SM in both physiological and high [K+]o conditions on single skeletal muscle fibers. The results showed that SM could consistently induce membrane potential hyperpolarization by a few millivolts, and this hyperpolarization was not possible in the presence of ouabain. In contrast, the same electrical pulses but with random frequencies, constant frequencies, or synchronization with backward-modulation could not hyperpolarize the membrane potential. Prolonged field application and higher field intensity enhanced the effects of SM-induced hyperpolarization. Finally, the effect of SM was tested on skeletal muscle fibers incubated in a solution with high external potassium. Results showed that the SM electric field could hyperpolarize the membrane potential even if the external K+ concentration was higher than the normal, which implied the therapeutic effects of the SM electric field on the hyperkalemic situation. [ABSTRACT FROM AUTHOR]

The sodium–potassium pump (Na+/K+ pump) is crucial for cell physiology. Despite great advances in the understanding of this ionic pumping system, its mechanism is not completely understood. We propose the use of a statistical model checker to investigate palytoxin (PTX)-induced Na+/K+ pump channels. We modelled a system of reactions representing transitions between the conformational substates of the channel with parameters, concentrations of the substates and reaction rates extracted from simulations reported in the literature, based on electrophysiological recordings in a whole-cell configuration. The model was implemented using the UPPAAL-SMC platform. Comparing simulations and probabilistic queries from stochastic system semantics with experimental data, it was possible to propose additional reactions to reproduce the single-channel dynamic. The probabilistic analyses and simulations suggest that the PTX-induced Na+/K+ pump channel functions as a diprotomeric complex in which protein–protein interactions increase the affinity of the Na+/K+ pump for PTX.

The Na + /K + -ATPase interacts with the non-selective cation channels TRPC6 but the functional consequences of this association are unknown. Experiments performed with HEK cells over-expressing TRPC6 channels showed that inhibiting the activity of the Na + /K + -ATPase with ouabain reduced the amount of TRPC6 proteins and depressed Ca 2 + entry through TRPC6. This effect, not mimicked by membrane depolarization with KCl, was abolished by sucrose and bafilomycin-A, and was partially sensitive to the intracellular Ca 2 + chelator BAPTA/AM. Biotinylation and subcellular fractionation experiments showed that ouabain caused a multifaceted redistribution of TRPC6 to the plasma membrane and to an endo/lysosomal compartment where they were degraded. The amyloid beta peptide Aβ 1–40 , another inhibitor of the Na + /K + -ATPase, but not the shorter peptide Aβ 1–16 , reduced TRPC6 protein levels and depressed TRPC6-mediated responses. In cortical neurons from embryonic mice, ouabain, veratridine (an opener of voltage-gated Na + channel), and Aβ 1–40 reduced TRPC6-mediated Ca 2 + responses whereas Aβ 1–16 was ineffective. Furthermore, when Aβ 1–40 was co-added together with zinc acetate it could no longer control TRPC6 activity. Altogether, this work shows the existence of a functional coupling between the Na + /K + -ATPase and TRPC6. It also suggests that the abundance, distribution and activity of TRPC6 can be regulated by cardiotonic steroids like ouabain and the naturally occurring peptide Aβ 1–40 which underlines the pathophysiological significance of these processes. [ABSTRACT FROM AUTHOR]

Objective: The mechanisms associated with nerve dysfunction and axonal loss in diabetes has not been fully clarified. Excitability and pathological aspects in nerves from diabetic mice were studied in order to explore the pathophysiology of diabetic neuropathy. Methods: Myelinated nerve fibres from the sciatic nerve of BKS.Cg-m+/+ Leprdb/J mice were studied by registering the CMAP controlled by an automated threshold tracking method. The sciatic nerve was also studied pathologically. Results: Diabetic mice displayed longer latencies, higher thresholds and lower amplitudes compared to controls and had a rightward shift in the stimulus response curves. Strength-duration time constant was lower in diabetic mice but not reaching statistical significance (p = 0.09). Diabetics displayed an increase in accommodation, with a smaller change in excitability in threshold electrotonus. Refractoriness, mean superexcitability and late subexcitability were reduced in diabetic mice. Diabetic mice had a larger number of myelinated fibres compared to controls (p < 0.05), but larger than 9 µm were virtually absent, accounting for near 7 % in control animals. Conclusions: Db/db mice develop electrophysiological changes suggestive of membrane depolarization as the result of Na+ /K+ pump impairment. Loss of large myelinated fibres might also contribute to the nerve excitability profiles in this model. [ABSTRACT FROM AUTHOR]

In rodents, ubiquitous α1-Na, K-ATPase is inhibited by ouabain and other cardiotonic steroids (CTS) at ~10-fold higher concentrations than those effective in other mammals. To examine the specific roles of the CTS- sensitive α1 S- and CTS- resistant α1 R-Na, K-ATPase isoforms, we compared the effects of ouabain on intracellular Na and K content, cell survival, and mitogen-activated protein kinases (MAPK) in human and rat vascular smooth muscle cells (HASMC and RASMC), human and rat endothelial cells (HUVEC and RAEC), and human and rat brain astrocytes. 6-h exposure of HASMC and HUVEC to 3 μM ouabain dramatically increased the intracellular [Na]/[K] ratio to the same extend as in RASMC and RAEC treated with 3000 μM ouabain. In 24, 3 μM ouabain triggered the death of all types of human cells used in this study. Unlike human cells, we did not detect any effect of 3000-5000 μM ouabain on the survival of rat cells, or smooth muscle cells from mouse aorta (MASMC). Unlike in the wild-type α1 mouse, ouabain triggered death of MASMC from α1 mouse expressing human α1-Na, K-ATPase. Furthermore, transfection of HUVEC with rat α1 R-Na, K-ATPase protected them from the ouabain-induced death. In HUVEC, ouabain led to phosphorylation of p38 MAPK, whereas in RAEC it stimulated phosphorylation of ERK1/2. Overall, our results, demonstrate that the drastic differences in cytotoxic action of ouabain on human and rodent cells are caused by unique features of α1S/α1R-Na, K-ATPase, rather than by any downstream CTS-sensitive/resistant components of the cell death machinery. [ABSTRACT FROM AUTHOR]

The sodium-potassium pump (Na + /K + pump) is crucial for cell physiology. Despite great advances in the understanding of this ionic pumping system, its mechanism is not completely understood. We propose the use of a statistical model checker to investigate palytoxin (PTX)-induced Na + /K + pump channels. We modelled a system of reactions representing transitions between the conformational substates of the channel with parameters, concentrations of the substates and reaction rates extracted from simulations reported in the literature, based on electrophysiological recordings in a whole-cell configuration. The model was implemented using the UPPAAL-SMC platform. Comparing simulations and probabilistic queries from stochastic system semantics with experimental data, it was possible to propose additional reactions to reproduce the single-channel dynamic. The probabilistic analyses and simulations suggest that the PTX-induced Na + /K + pump channel functions as a diprotomeric complex in which protein-protein interactions increase the affinity of the Na + /K + pump for PTX., Competing Interests: We declare we have no competing interests.