Sodium Pump Research Articles

Pivotal role of adipocyte‐Na/K‐ATPase Signaling in the pathogenesis of Experimental Uremic Cardiomyopathy by fat transplantation

ObjectiveMounting evidence highlights the role of adipose tissue in the development of systemic inflammation that contributes to various metabolic abnormalities. We have recently demonstrated that administration of NaKtide, antagonist of Na/K‐ATPase (NKA) signaling, coupled to adipocyte specific promoter can improve adipocyte phenotype and further attenuated experimental uremic cardiomyopathy. Studies have shown that transplanting brown adipose tissue from C57BL6 mice reduced obesity and improved whole energy metabolism. Therefore, the objective of the present study is to explore the specific role of adipocyte function in systemic inflammation and associated pathologies of uremic cardiomyopathy using adipose tissue transplantation studies.HypothesisIn the present study, we hypothesize that, the transplantation of NaKtide transfected subcutaneous fat into mice with partial nephrectomy (PNx) may help to provide a deeper understanding of adipocyte‐NKA signaling, that modulate the progression of uremic cardiomyopathy.MethodsFollowing 4 weeks of PNx or Sham surgery and lenti‐adiponectin‐Naktide in C57BL6 mice, 200 mg of subcutaneous fat pads were dissected and subcutaneously transplanted into the dorsal interscapular region of the recipient mice at the time of the PNx surgery. The tissues were harvested for morphological and molecular analyses after 8 weeks of transplantation. Statistical analysis of the data were performed by one‐way analysis of variance.ResultsThe immunofluorescence staining showed high expression of NaKtide in fat tissues of mice transplanted with NaKtide transfected fat. The pSrc protein expression in the fat tissue was significantly upregulated in PNx, which was significantly down‐regulated by PNx+NaKtide fat transplantation (p<0.01). The adipose tissue dysfunction during PNx was assessed by the level of protein carbonylation and mRNA expression of IL‐1β and Sirt2, which were significantly reversed by NaKtide transfected fat (p<0.01). The transplantation of NaKtide transfected adipose tissue significantly improved the systemic inflammation (p<0.01). The altered plasma miRNA expressions in PNx were also normalized by NaKtide transfected fat transplantation. PNx mice developed cardiomyopathy characterized by increased heart weight and decreased cardiac function, assessed by echocardiography measurements (p<0.01). These alterations were reversed by the transplantation of adipose tissues from PNx+NaKtide donor mice (p<0.01). The mice transplanted with the fat tissue of PNx+NaKtide showed significantly decreased cardiac fibrosis, when compared to PNx heart, which was further confirmed by the mRNA expression of major fibrotic genes in cardiac tissues.ConclusionThe study demonstrates that, the inhibition of adipocyte‐NKA signaling in uremic cardiomyopathy could modulate the adipocyte redox state that resulted in the manipulation of systemic inflammatory milieu and associated pathophysiological mechanisms, which can be used as a clinical target for therapeutic intervention of uremic cardiomyopathy.

Quantitative Expression of Drug, Phospholipid and Nucleoside Transporters in the Lung Tissues across Species

ObjectiveIt has been proven that transporters play a pivotal role in organ distribution of drugs, and subsequently in drug efficacy and toxicity. Active transport processes may affect the local pulmonary disposition of drugs administered directly to the lung by the inhaled route as well as the uptake of drugs to the lung from the systemic circulation. While drug transporter expressions and their roles in drug disposition and elimination in the major elimination organs including intestine, liver and kidney are well‐characterized, the expression of drug transporters and the comparison across species in the lung remain overall poorly understood. For example, azithromycin levels in pulmonary tissue remained to be high for up to 5 days after a single oral dose, in contrast to much lower plasma levels. However, the mechanisms associated with azithromycin lung loading and efficacy remain unknown. The aim of this study was to comprehensively determine the mRNA and protein expression levels of drug transporters in the lung in human and preclinical species using quantitative targeted proteomics approaches. The findings can be useful to further explore potential mechanisms in lung specific disposition.MethodsTotal cellular mRNA was isolated from human lower lung tissues using a Qiagen mRNA isolation kit. qRT‐PCR was performed, and β‐actin mRNA expression was used to normalize target gene expression. The mRNA levels were expressed as a fold change using the 2‐ΔΔCt method. The total membrane proteins of lung tissues from human, monkey, dog and rat were extracted by Native Membrane Protein Extraction Kit, and subsequently digested by trypsin followed by the treatment of DTT and iodoacetamide (IAA). The levels of transporter proteins were quantified using a Sciex 7500 triple‐quadruple mass spectrometer coupled to a Shimadzu LC (SLC‐30A) system with optimized ionization conditions. The Na K‐ATPase level in the lung was measured as the membrane protein marker.Results and conclusionIn the current investigation, thirteen ABC efflux transporters and twenty‐three solute carriers (SLC) were characterized. Among those, mRNAs of eleven ABC transporters and fifteen SLC transporters were detected by the RT‐PCR method. In addition, eight ABC transporter and eleven SLC transporter proteins were quantified using the targeted proteomics LC‐MS/MS method. The mRNA expression pattern agrees with the protein expression; however, no quantitative correlation can be drawn between mRNA and protein expression. While OATP2B1 was only detected in human lung tissues, OATP2A1 and 3A1 are expressed in the lungs of human, monkey dog and rat. The expressions of three equilibrative nucleoside transporters (ENT1/2/3) in human lung tissues were higher than those in monkey. The expression of ABCA1, ABCA3 and ABCG1, the ABC transporters that are the regulators of cellular cholesterol and phospholipid homeostasis, were detected in all tested species, and found to be high in rat. Collectively, the comparison of transporter expression profiles can contribute to a better understanding of drug distribution and transport in the lung and aid pulmonary drug discovery and development.

Preparation and Characterization of Nanohybrid La2O3-K Complexes for Electrochemical Study

In biomedical science, alkali metal potassium plays a conspicuous role in the neurotransmission known as sodium (Na)-potassium (K) pump. The rare earth metal like lanthanum is most applicable in the magnetic resonance imaging technique (MRI) for cancerous cell detection. Keeping in mind this perspective, we synthesized the hybrid of La2O3 nanomaterial and K-complexes [(μ2-4-N,N-dimethylamino benzoate-κO)(μ2-4-N,N-dimethylamino benzoic acid-κO) (4-N,N-dimethylamino benzoic acid-κO) potassium(I) coordination polymer)] (nanohybrid La2O3- K complex) by hydrothermal method, whereas the potassium complex has been synthesized by the 6 hours reflection of the 4-(diethylamino) benzoic acid and potassium hydroxide in the methanol as solvent. The1H, 13C NMR, HRMS, and single-crystal investigation have been carried out for the potassium complex. Moreover, the structural, morphological, and electrochemical estimation of synthesized nanocomposites are also under process, which involves the investigation by X-ray diffraction (XRD), Fourier transform infrared (FTIR), and UV- visible spectroscopy.

Evidence for tissue-specific defence-offence interactions between milkweed and its community of specialized herbivores.

Coevolution between plants and herbivores often involves escalation of defence-offence strategies, but attack by multiple herbivores may obscure the match of plant defence to any one attacker. As herbivores often specialize on distinct plant parts, we hypothesized that defence-offence interactions in coevolved systems may become physiologically and evolutionarily compartmentalized between plant tissues. We report that roots, leaves, flower buds and seeds of the tropical milkweed (Asclepias curassavica) show increasing concentrations of cardenolide toxins acropetally, with latex showing the highest concentration. In vitro assays of the physiological target of cardenolides, the Na+ /K+ -ATPase (hereafter "sodium pump"), of three specialized milkweed herbivores (root-feeding Tetraopes tetrophthalmus, leaf-feeding Danaus plexippus, and seed-feeding Oncopeltus fasciatus) show that they are proportionally tolerant to the cardenolide concentrations of the tissues they eat. Indeed, molecular substitutions in the insects' sodium pumps predicted their tolerance to toxins from their target tissues. Nonetheless, the relative inhibition of the sodium pumps of these specialists by the concentration versus composition (inhibition controlled for concentration, what we term "potency") of cardenolides from their target versus nontarget plant tissues revealed different degrees of insect adaptation to tissue-specific toxins. In addition, a trade-off between toxin concentration and potency emerged across plant tissues, potentially reflecting coevolutionary history or plant physiological constraints. Our findings suggest that tissue-specific coevolutionary dynamics may be proceeding between the plant and its specialized community of herbivores. This novel finding may be common in nature, contributing to ways in which coevolution proceeds in multispecies communities.

Real-time modeling of transient behavior of the primary coolant system in pool-type sodium cooled fast reactor

In the paper, a real-time simulation code THCSA-RT is developed to analyze the primary coolant system of pool-type sodium cooled fast reactor which includes reactor core, hot and cold sodium pools, pumps, and intermediate heat exchangers. Different numerical methods including Gear multi-value method and Quasi-Newton iteration method are used to solve the mass and energy equations set and the momentum equations set, respectively. Several typical design basis conditions of CEFR such as planned shutdown condition and RIA are analyzed, and the calculation results agree well with those of the final safety analysis report calculated by DINROS. The simulation time is presented to prove the effectiveness of the new numerical methods developed in the present work, and the new numerical methods could be applied in the similar system analysis codes as well. The newly developed code THCSA-RT could be used in the system real-time simulation of pool-type sodium cooled fast reactor and has been adopted in the development of CEFR full scope simulator.

Na,K-ATPase α1 and β-subunits show distinct localizations in the nervous tissue of the large milkweed bug

The Na,K-ATPase (NKA) is an essential ion transporter and signaling molecule in all animal tissues and believed to consist at least one α and one ß-subunit to form a functional enzyme. In the large milkweed bug, Oncopeltus fasciatus, adaptation to dietary cardiac glycosides (CGs), which can fatally block the NKA, has resulted in gene duplications leading to four α1-subunits. These differ in sensitivity to CGs, but resistance trades off against ion pumping activity, thus influencing the α1-subunits’ suitability for specific tissues. Besides, O. fasciatus possesses four different ß-subunits that can alter the NKA's kinetics and should play an essential role in the formation of cellular junctions.Proteomic analyses revealed the distribution and composition of α1/ß-complexes in the nervous tissue of O. fasciatus. The highly CG-resistant, but less active α1B and the highly active, but less resistant α1C predominated in the nervous tissue and co-occurred with ß2 and ß3, partly forming larger complexes than just heterodimers. Immunohistochemical analyses provided a fine scale resolution of the subunits’ distribution in different morphological structures of the nervous tissue. This may suggest that α1 as well as ß-subunits occur in isolation without the other subunit, which contradicts the present understanding that the two types of subunits have to associate to form functional complexes. An isolated occurrence was especially prominent for ß3 and βx, the enigmatic fourth and N-terminally largely truncated ß-subunit. We hypothesize that dimerization of these ß-subunits plays a role in cell–cell contacts.

Fluorescence lifetime imaging microscopy reveals sodium pump dimers in live cells

The sodium-potassium ATPase (Na/K-ATPase, NKA) establishes ion gradients that facilitate many physiological functions including action potentials and secondary transport processes. NKA comprises a catalytic subunit (alpha) that interacts closely with an essential subunit (beta) and regulatory transmembrane micropeptides called FXYD proteins. In the heart, a key modulatory partner is the FXYD protein phospholemman (PLM, FXYD1), but the stoichiometry of the alpha–beta–PLM regulatory complex is unknown. Here, we used fluorescence lifetime imaging and spectroscopy to investigate the structure, stoichiometry, and affinity of the NKA-regulatory complex. We observed a concentration-dependent binding of the subunits of NKA–PLM regulatory complex, with avid association of the alpha subunit with the essential beta subunit as well as lower affinity alpha–alpha and alpha–PLM interactions. These data provide the first evidence that, in intact live cells, the regulatory complex is composed of two alpha subunits associated with two beta subunits, decorated with two PLM regulatory subunits. Docking and molecular dynamics (MD) simulations generated a structural model of the complex that is consistent with our experimental observations. We propose that alpha–alpha subunit interactions support conformational coupling of the catalytic subunits, which may enhance NKA turnover rate. These observations provide insight into the pathophysiology of heart failure, wherein low NKA expression may be insufficient to support formation of the complete regulatory complex with the stoichiometry (alpha-beta-PLM)2.

Transfusion-associated graft-versus-host disease, transfusion-associated hyperkalemia, and potassium filtration: advancing safety and sufficiency of the blood supply

Irradiation of cellular blood components is well established as a countermeasure against transfusion-associated graft-versus-host disease (TA-GVHD). Unintended consequences of ionizing radiation are also well established. The red cell “storage lesion” – a progression of metabolic, functional, and morphological changes – may be exacerbated by irradiation rates and doses typically used for TA-GVHD prophylaxis. With or without irradiation, a storage lesion change of clinical concern is the accelerated egress of intracellular potassium. ATP depletion during storage limits the activity of the red cell membrane’s sodium-potassium pump (Na,K-ATPase), which normally maintains intracellular potassium (K+) at levels 30–40 times higher than the extracellular milieu. The natural diffusion of potassium down this concentration gradient proceeds faster if the cell membrane is damaged, and oxidative damage to cellular membranes and membrane proteins – including Na,K-ATPase – is an effect of ionizing radiation.Preventing transfusion-related hyperkalemia is a reason for limiting the shelf life of irradiated red cells. In the absence of specific measurements to assess storage lesion in a particular unit of blood, and in the absence of specific interventions at the time of transfusion to mitigate effects of storage lesion, it is consistent with the precautionary principle to put conservative limits on a blood component’s shelf life. On the other hand, both the safety and sufficiency of a nation’s blood supply might be improved by interventions that benefit specific recipients when they are transfused, and benefit future patients by extending the allowable shelf life of blood components. Potassium filtration of irradiated red blood cell components is one such intervention.

Calcium Binding to TAT Rhodopsin.

Rhodopsin is a large family of retinal-binding photoreceptive proteins found in animals and microbes. The retinal chromophore is normally positively charged by protonation of the Schiff base linkage, which is stabilized by the negatively charged counterion(s) such as aspartates, glutamates, and chloride ions. In contrast, no cation binding was reported near the retinal chromophore under physiological pH, presumably because of the electrostatic repulsion. Sodium binding takes place in light-driven sodium pumps, but the binding near the retinal chromophore is a transient event. Here, we report Ca2+ binding to a wild-type microbial rhodopsin, which is achieved for the neutral retinal chromophore with a deprotonated Schiff base. TAT rhodopsin from marine bacteria contains protonated and deprotonated retinal Schiff bases at physiological pH (pH ∼ 8), which absorb visible and UV light, respectively. We observed that the equilibrium shifted toward the deprotonated state upon increasing Ca2+ concentration, and the Kd value was determined to be 0.17 mM. Site-directed mutagenesis study showed that E54 and D227 constitute the binding site of Ca2+. ATR-FTIR spectroscopy revealed secondary structural changes upon Ca2+ binding to E54 and D227, while they are negatively charged with or without Ca2+ binding.

Thyrotoxic Hypokalemic Periodic Paralysis: Case Report

Introduction: 
 Thyrotoxic Hypokalemic Periodic Paralysis (THPP) is a rare hereditary disorder which is characterized by thyroid hormone elevation, low blood potassium level and recurrent acute muscle weakness. Basic pathology is thought to be the increase in activity in the sodium-potassium pump (Na+/K+ATPase). 
 
 Case report: 
 Here we report the case of a 31-year-old male that presented with weakness in his legs, and inability to walk. The patient had elevated thyroid hormone levels (FT3 and FT4) and lower TSH levels, lower serum potassium levels, and recurrent acute muscle weakness. The diagnosis was made to be Thyrotoxic Hypokalemic Periodic Paralysis precipitated after intense physical activity. 
 
 Conclusion: 
 THPP is a reversible medical emergency. Early diagnosis, and rapid treatment is lifesaving. Although rare, THPP must be considered as a differential diagnosis in patients presenting with hypokalemia and paralysis.

Lactato y catecolaminas: respuesta fisiológica en el paciente crítico

Lactate is a highly dynamic metabolite that is produced, under anaerobic conditions, due to hypoxia or ischemia. Under aerobic conditions, it is synthesized by a mechanism driven by the stimulation of the β2 adrenergic receptor, which increases the activity of the sodium-potassium pump, and by a state of accelerated aerobic glycolysis. This metabolite is capable of being exchanged between different producing and consuming cells, ensuring the raw material for energy production.The sympathetic nervous system responds to stress stimuli through the release of catecholamines, which act as hormones and neurotransmitters in various tissues of the body, allowing an increase in metabolism that raises glucose and available oxygen levels.There is a physiological dependence between catecholamine levels and lactate production, predisposing the body to respond effectively to a stressful situation. However, an exacerbated adrenergic response may cause exaggerated effects on sensitive tissues that increase the probability of failure. Based on the knowledge of these mechanisms, therapeutic strategies focused on regulating the sympathetic activity are proposed.

Molecular characterization of alpha subunit 1 of sodium pump (ATP1A1) gene in Camelus dromedarius: its differential tissue expression potentially interprets the role in osmoregulation.

Dromedary or one-humped camel (Camelus dromedarius) is distinctively acclimatized to survive the arid conditions of the desert environment. It has an excellent ability to compete dehydration with substantial tolerance for rapid dehydration. Therefore, it offers an excellent model for studying osmoregulation. Molecular characterization of Na+/K+ ATPase as a central regulator of electrolyte normohemostasis affords a better understanding of this mechanism in camel. Here is the first to resolve the full-length of alpha-1 subunit of sodium pump (ATP1A1) gene with its differential expression in dromedary tissues. The nucleotide sequence for the recovered full cDNA of ATP1A1was submitted to the GenBank (NCBI GenBank accession #MW628635) and bioinformatically analyzed. The cDNA sequence was of 3760bp length with an open reading frame (ORF) of 3066bp encoding a putative 1021 amino acids polypeptide with a molecular mass of 112696Da. Blast search analysis revealed the shared high similarity of dromedary ATP1A1gene with other known ATP1A1genes in different species. The comparative analysis of its protein sequence confirmed the high identity with other mammalian ATP1A1 proteins. Further transcriptomic investigation for different organs was performed by real-time PCR to compare its level of expression among different organs. The results confirm a direct function between the ATP1A1 gene expression and the order of vital performance of these organs. The expression ofATP1A1mRNA in the adrenal gland and brain was significantly higher than that in the other organs. The noticed down expression in camel kidney concomitant with overexpression in the adrenal cortex might interpret how dromedary expels access sodium without water loss with relative high ability to restrain mineralocorticoid-induced sodium retention on drinking salty water. The results reflect the importance of sodium pump in these organs. Na+/K+ ATPase in the adrenal gland and brain than other organs.

EP291: Disease characterization in sodium-potassium ATPases by reverse genetics in humans

Most genetic disorders are discovered via forward genetics: a shared phenotype is recognized in a group of patients, allowing causative gene(s) to be determined. This can lead to excessively narrow definitions of disease phenotypes due to selection bias, as patients with symptoms outside the “classic” presentation are less likely to be diagnosed. We hypothesized that reverse human genetics, ie, studying variants in a gene without phenotype-based recruitment, could more accurately characterize new diseases. We established the first such approach for ATP1A1, the ubiquitously expressed housekeeping isoform of sodium-potassium ATPase. ATP1A1-associated disease is an emerging entity with several reports of heterozygous variants in individuals with differing neurological phenotypes: peripheral neuropathy, childhood epilepsy-renal hypomagnesemia, or spastic paraplegia. Gain-of-function ion leakage has been proposed as a mechanism for the more severe phenotype in the patients with epilepsy, versus loss-of-function in patients with neuropathy. However, only 14 families in total have been reported with possible ATP1A1 pathogenic variants and the true spectrum of disease remains unknown. Through genotype-based recruitment combined with detailed functional characterization of variants, we broadened the phenotypic spectrum to include incomplete penetrance and identified distinct molecular categories of ATP1A1 variants. We collected variants identified on next-generation sequencing of both clinical and research samples. We recruited patients with heterozygous ATP1A1 variants of uncertain significance found on testing for any indication. In addition, we filtered rare variants found in 7,000 “healthy” volunteers in a population genome sequencing study and prioritized 2 for study based on predicted functional impact. We re-contacted one individual from this cohort for targeted phenotyping at the NIH Clinical Center under an IRB-approved study protocol (ClinicalTrials.gov identifier NCT00369421). We developed a comprehensive panel of 3 assays measuring the ability of ATP1A1 to support survival in transfected cells and its total enzymatic activity and turnover rate per molecule in Xenopus oocytes. In the latter experimental system, we directly measured enzyme activity as steady-state electrical currents in two-electrode voltage clamp. Using sodium transient current measurements, we quantified sodium ion substrate binding by ATP1A1 as well as the number of ATP1A1 molecules reaching the cell surface. We identified 7 rare missense variants with significant impacts on enzyme function, as well as 1 nonsense variant predicted to lead to nonsense-mediated decay. Of 9 missense variants identified in clinically ascertained people, 6 had functional impact and 3 did not. Phenotypes of patients with functionally significant ATP1A1 variants included not only neurological deficits but also previously unreported features such as hypothyroidism (two patients), skin changes, and unusual facial features. Of the 2 variants selected from the population sequencing cohort, 1 was a missense variant with clear loss-of-function impact in multiple assays yet was present in an adult without neurological symptoms. This individual declined research phenotyping. The other was a nonsense variant identified in an asymptomatic adult. This individual underwent detailed phenotyping including nerve conduction/electromyography and was confirmed not to have subclinical neuropathy, magnesium wasting, or spasticity. Molecular characterization of the variants revealed three general patterns of impact on enzyme function. First, some variants decreased protein abundance at the cell membrane with a corresponding loss of enzyme function. Second, other variants resulted in ATP1A1 that reached the cell membrane but had decreased enzyme activity with or without ion leak currents. Third, some variants changed enzyme properties by reducing the voltage dependence of its activity, possibly suggesting gain-of-function at negative voltages. Genotype-based collection of variants for functional characterization was an effective strategy to study the spectrum of ultra-rare ATP1A1 disease. We identified 8 putative pathogenic variants, expanding the total known pool of such variants by more than half. Connecting functional impacts of variants to the phenotypes of the individuals carrying them revealed incomplete penetrance into adulthood in two individuals as well as possible non-neurological disease features for further study. Detailed molecular characterization allowed us to propose distinct classes of variants based on their effects on enzyme function, which could form the basis for future genotype-phenotype correlation.

Ceramide-1-Phosphate as a Potential Regulator of the Second Sodium Pump from Kidney Proximal Tubules by Triggering Distinct Protein Kinase Pathways in a Hierarchic Way.

Kidney proximal tubules are a key segment in the reabsorption of solutes and water from the glomerular ultrafiltrate, an essential process for maintaining homeostasis in body fluid compartments. The abundant content of Na+ in the extracellular fluid determines its importance in the regulation of extracellular fluid volume, which is particularly important for different physiological processes including blood pressure control. Basolateral membranes of proximal tubule cells have the classic Na+ + K+-ATPase and the ouabain-insensitive, K+-insensitive, and furosemide-sensitive Na+-ATPase, which participate in the active Na+ reabsorption. Here, we show that nanomolar concentrations of ceramide-1 phosphate (C1P), a bioactive sphingolipid derived in biological membranes from different metabolic pathways, promotes a strong inhibitory effect on the Na+-ATPase activity (C1P50 ≈ 10 nM), leading to a 72% inhibition of the second sodium pump in the basolateral membranes. Ceramide-1-phosphate directly modulates protein kinase A and protein kinase C, which are known to be involved in the modulation of ion transporters including the renal Na+-ATPase. Conversely, we did not observe any effect on the Na+ + K+-ATPase even at a broad C1P concentration range. The significant effect of ceramide-1-phosphate revealed a new potent physiological and pathophysiological modulator for the Na+-ATPase, participating in the regulatory network involving glycero- and sphingolipids present in the basolateral membranes of kidney tubule cells.

Sodium, Glucose and Dysregulated Glucagon Secretion: The Potential of Sodium Glucose Transporters.

Diabetes is defined by hyperglycaemia due to progressive insulin resistance and compromised insulin release. In parallel, alpha cells develop dysregulation of glucagon secretion. Diabetic patients have insufficient glucagon secretion during hypoglycaemia and a lack of inhibition of glucagon secretion at higher blood glucose levels resulting in postprandial hyperglucagonaemia, which contributes to the development of hyperglycaemia. Sodium-glucose co-transporter 2 (SGLT2) inhibitors are an efficient pharmacologic approach for the treatment of hyperglycaemia in type 2 diabetes. While SGLT2 inhibitors aim at increasing glycosuria to decrease blood glucose levels, these inhibitors also increase circulating glucagon concentrations. Here, we review recent advances in our understanding of how SGLTs are involved in the regulation of glucagon secretion. Sodium plays an important role for alpha cell function, and a tight regulation of intracellular sodium levels is important for maintaining plasma membrane potential and intracellular pH. This involves the sodium-potassium pump, sodium-proton exchangers and SGLTs. While the expression of SGLT2 in alpha cells remains controversial, SGLT1 seems to play a central role for alpha cell function. Under hyperglycaemic conditions, SGLT1 mediated accumulation of sodium results in alpha cell dysregulation due to altered cellular acidification and ATP production. Taken together, this suggests that SGLT1 could be a promising, yet highly underappreciated drug target to restore alpha cell function and improve treatment of both type 1 and 2 diabetes.

Endogenous Cardiac Steroids in Bipolar Disorder: State of the Art.

Bipolar disorder (BD) is a severe psychiatric illness with a poor prognosis and problematic, suboptimal, treatments. Treatments, borne of an understanding of the pathoetiologic mechanisms, need to be developed in order to improve outcomes. Dysregulation of cationic homeostasis is the most reproducible aspect of BD pathophysiology. Correction of ionic balance is the universal mechanism of action of all mood stabilizing medications. Endogenous sodium pump modulators (collectively known as endogenous cardiac steroids, ECS) are steroids which are synthesized in and released from the adrenal gland and brain. These compounds, by activating or inhibiting Na+, K+-ATPase activity and activating intracellular signaling cascades, have numerous effects on cell survival, vascular tone homeostasis, inflammation, and neuronal activity. For the past twenty years we have addressed the hypothesis that the Na+, K+-ATPase-ECS system may be involved in the etiology of BD. This is a focused review that presents a comprehensive model pertaining to the role of ECS in the etiology of BD. We propose that alterations in ECS metabolism in the brain cause numerous biochemical changes that underlie brain dysfunction and mood symptoms. This is based on both animal models and translational human results. There are data that demonstrate that excess ECS induce abnormal mood and activity in animals, while a specific removal of ECS with antibodies normalizes mood. There are also data indicating that circulating levels of ECS are lower in manic individuals, and that patients with BD are unable to upregulate synthesis of ECS under conditions that increase their elaboration in non-psychiatric controls. There is strong evidence for the involvement of ion dysregulation and ECS function in bipolar illness. Additional research is required to fully characterize these abnormalities and define future clinical directions.

De Novo ATP1A1 Variants in an Early-Onset Complex Neurodevelopmental Syndrome.

ATP1A1 encodes the α1 subunit of the sodium-potassium ATPase, an electrogenic cation pump highly expressed in the nervous system. Pathogenic variants in other subunits of the same ATPase, encoded by ATP1A2 or ATP1A3, are associated with syndromes such as hemiplegic migraine, dystonia, or cerebellar ataxia. Worldwide, only 16 families have been reported carrying pathogenic ATP1A1 variants to date. Associated phenotypes are axonal neuropathies, spastic paraplegia, and hypomagnesemia with seizures and intellectual disability. By whole exome or genome sequencing, we identified 5 heterozygous ATP1A1 variants, c.674A>G;p.Gln225Arg, c.1003G>T;p.Gly335Cys, c.1526G>A;p.Gly509Asp, c.2152G>A;p.Gly718Ser, and c.2768T>A;p.Phe923Tyr, in 5 unrelated children with intellectual disability, spasticity, and peripheral, motor predominant neuropathy. Additional features were sensory loss, sleep disturbances, and seizures. All variants occurred de novo and are absent from control populations (MAF GnomAD = 0). Affecting conserved amino acid residues and constrained regions, all variants have high pathogenicity in silico prediction scores. In HEK cells transfected with ouabain-insensitive ATP1A1 constructs, cell viability was significantly decreased in mutants after 72h treatment with the ATPase inhibitor ouabain, demonstrating loss of ATPase function. Replicating the haploinsufficiency mechanism of disease with a gene-specific assay provides pathogenicity information and increases certainty in variant interpretation. This study further expands the genotype-phenotype spectrum of ATP1A1.

Conduction differences between atrial and ventricular working myocardium elucidated through previously unrecognized nanoscale structural organization

The intercalated disc (ID) is a complex, heterogeneous structure that affords electrical (gap junctions; GJ) and mechanical (adherence junctions [AJ], desmosomes [Des]) coupling between cardiomyocytes. Electrogenic proteins underlying the action potential upstroke (cardiac sodium channels [NaV1.5], inward-rectifier potassium channels [Kir2.1] and sodium potassium ATPase [NKA]) enriched within ID nanodomains are emerging as vital machinery for cardiac impulse propagation. ID structure is thus a critical determinant of cardiac conduction.

Intercalated disk nanoscale structure and ion channel localization regulate cardiac conduction

The intercalated disk (ID) is the specialized subcellular region that provides electrical and mechanical connections between myocytes in the heart. The ultrastructure of the ID is composed of mechanical junctions (MJ), gap junctions (GJ) and non-junctional membranes. The junctions and the two apposing cell ID membranes describe a complex, restricted extracellular space between the cells called the intercellular cleft. Recent studies have shown that ID heterogeneity extends to key electrogenic proteins, such as the cardiac voltage-gated sodium channel (NaV1.5), inward-rectifying potassium channel (Kir2.1) and sodium-potassium ATPase (NKA): they are preferentially localized at the IDs, and moreover tend to cluster around MJs and GJs.

Stoichiometry of the sodium pump-phospholemman regulatory complex

The sodium-potassium ATPase (NKA) is the ion motive ATP-dependent transporter that establishes sodium and potassium gradients across the cell membrane, facilitating many physiological processes. In the heart, NKA activity is regulated by its interaction with phospholemman (PLM, FXYD1), a member of the FXYD protein family. PLM inhibits NKA by reducing pump’s apparent affinity of the pump for Na+, which is relieved by PLM phosphorylation. In this study, we used time-correlated single-photon counting, FRET-microscopy and molecular dynamics simulations to investigate the structure, stoichiometry, and affinity of the NKA-PLM regulatory complex.