Pump Activity Research Articles

An overview of molecular mechanisms in cancer drug resistance and therapeutic strategies

Cancer resistance refers to the ability of cancer cells to evade the effects of treatments, such as chemotherapy and radiation, making it challenging to eliminate tumors. Cancer cells develop resistance through genetic modifications, alterations in cellular pathways, or microenvironmental adaptations that allow cancer cells to persist, proliferate, and even thrive despite the application of therapies designed to eradicate them. Despite efforts to create more potent target-based drugs, the flexibility of acquired resistance necessitates advanced options. Combination therapy and precision immunotherapy has revolutionized treatment but is limited by patient specificity and requires further development. New approaches like small molecules, peptides, and nanotherapeutics aim to overcome resistance by enhancing site-specific delivery and increasing drug concentration inside cancer cells. This review provides a comprehensive overview of the molecular mechanisms underlying cancer drug resistance, including genetic mutations, epigenetic alterations, efflux pump activity, and the influence of the tumor microenvironment. Additionally, the roles of microRNAs, long noncoding RNAs, and cellular processes such as autophagy and hypoxia in mediating resistance are examined. This review seeks to improve patient care and help to the development of more effective cancer medicines by explaining these complicated systems and investigating novel therapeutic alternatives.

Recent Advances in Marine-Derived Compounds as Potent Antibacterial and Antifungal Agents: A Comprehensive Review.

The increase in antimicrobial resistance (AMR) in microorganisms is a significant global health concern. Various factors contribute to AMR, including alterations in cell membrane permeability, increased efflux pump activity, enzymatic modification or inactivation of antibiotics, target site changes, alternative metabolic pathways, and biofilm formation. Marine environments, with their extensive biodiversity, provide a valuable source of natural products with a wide range of biological activities. Marine-derived antimicrobial compounds show significant potential against drug-resistant bacteria and fungi. This review discusses the current knowledge on marine natural products such as microorganisms, sponges, tunicates and mollusks with antibacterial and antifungal properties effective against drug-resistant microorganisms and their ecological roles. These natural products are classified based on their chemical structures, such as alkaloids, amino acids, peptides, polyketides, naphthoquinones, terpenoids, and polysaccharides. Although still in preclinical studies, these agents demonstrate promising in vivo efficacy, suggesting that marine sources could be pivotal in developing new drugs to combat AMR, thereby fulfilling an essential medical need. This review highlights the ongoing importance of marine biodiversity exploration for discovering potential antimicrobial agents.

In silico and in vitro screening of selected antimicrobial compounds for inhibiting drug efflux pumps to combat threatening MRSA

The rapid emergence of antibiotic-resistant bacteria is occurring worldwide due to severe antibiotic abuse in the past few decades; among them, Methicillin-resistant Staphylococcus aureus (MRSA) is of major concern. Study examines potent antimicrobial compounds against Methicillin resistance Staphylococcus aureus (MRSA) efflux system. Interactome data serves as a primary focal point for global MRSA infection control initiatives. We employed CB Dock and Discovery Studio for performing docking analysis on specific antimicrobial compounds against NorA, NorB, and NorG. Additionally, we conducted molecular dynamics simulations using myPresto. In vitro validation was done by agar-based EtBr cartwheel method to assess the efflux pump activity of MRSA. Out of the eight screened antimicrobial compounds, quercetin exhibited favorable binding energies. Subsequently, molecular dynamics simulations were conducted on quercetin complexes with NorA, NorB, and NorG. In vitro result revealed that the quercetin is active at concentration of 60 µg/ml ±10 against MRSA to inhibit efflux mechanisms. Quercetin demonstrated the most promising binding affinity with the efflux pumps and their regulators of MRSA, making it the prime candidate for use with antibiotics. In-vivo experiments must be performed to validate the in-silico and in vitro analysis.

Evaluation of chlorogenic acid and carnosol for anti-efflux pump and anti-biofilm activities against extensively drug-resistant strains of Staphylococcus aureus and Pseudomonas aeruginosa.

In summary, CGA and CL demonstrated promising potentiating antimicrobial effects against XDR strains of Staphylococcus aureus and Pseudomonas aeruginosa, suggesting their probably potential as candidates for addressing nosocomial pathogens. They exhibited significant suppression of efflux pump activity, indicating a possible successful inhibition of this mechanism. Moreover, all substances effectively inhibited biofilm formation, while showing minimal cytotoxicity. However, further advancement to clinical trials is needed to evaluate the feasibility of utilizing CGA and CL for reversing bacterial XDR efflux and determining their efficacy against biofilms. These trials will provide valuable insights into the practical applications of these compounds in combating drug-resistant infections.

Siliceous deposition in limestone-marl alternations of the Yangtze Carbonate platform during the Permian Chert Event

During the Permian Chert Event (PCE), limestone-marl alternations (LMAs), contemporaneously developing in the Yangtze region with chert-mudstone alternations, differed from typical LMAs by their enrichment in authigenic siliceous components (quartziferous components and Mg-phyllosilicates). However, the factors driving the transition from distal chert deposits to proximal siliceous-rich carbonate deposits remain unclear, and the genesis of authigenic siliceous components is puzzling. Resolving these issues could provide a deeper understanding of the paleoclimate and paleoenvironment during the PCE. Optical and SEM observations and geochemical analyses revealed that Si4+, derived from seawater, silica-secreting organisms, and terrigenous clasts, precipitated as authigenic quartziferous components under relatively acidic conditions, while it precipitated as Mg-phyllosilicates through reverse weathering by combining with the dissolved high-Mg calcites under relatively alkaline conditions. Through the facies division of LMAs, its variations describe relative and short-term sea-level fluctuations in the Yangtze Carbonate Platform (YCP). By integrating tendency comparisons and time series analyses of sea level fluctuations, continental weathering intensity, authigenic siliceous content, and relative proportions of quartz vs. Mg-phyllosilicates, four proxies exhibit covariation and similar astronomical signals, indicating that the high sea level period of the YCP, corresponding to intensified continental weathering, favored the deposition of authigenic siliceous components and reverse weathering. Because the facies variations encompass short eccentricity, obliquity, and precession cycles, sea level fluctuations are considered to have been influenced by Gondwana glaciation P3. The seawater in the continental shelf cooled and became rich in silica and nutrients under the effect of upwelling. Interglacial periods may trigger high sea levels and intensified continental weathering in the YCP, facilitating the influx of Si-rich eutrophic seawater and stimulating biological silica secretion by radiolarians. However, the direct precipitation of activated silica resulted in weakened reverse weathering. Consequently, the lithological transformation from distal to proximal areas during the PCE was attributed to differences in silica-secreting organism abundance, upwelling intensity, biological pump activity, reverse weathering intensity, OM abundance, pH conditions, and seawater temperature between the continental shelf and carbonate platform.

Thymoquinone’ potent impairment of multidrug-resistant Staphylococcus aureus NorA efflux pump activity

The drug efflux pump is a crucial mechanism implicated in resistance to multiple antimicrobials. Thymoquinone (TQ) has evidently demonstrated multiple activities, antibacterial being the most effective. Knowledge about TQ activity against multidrug-resistant Staphylococcus aureus is very scarce. Therefore, the present study was conducted to investigate TQ resistance modulation in ciprofloxacin (CIP) and doxycycline (DO) multidrug-resistant S. aureus. Forty-seven samples were collected from different sources, and S. aureus was isolated and identified. Then, S. aureus resistance profiles to antimicrobials, N. sativa essential oil, and TQ; the correlation between TQ-MIC readings and disc diffusion; cartwheel and ethidium bromide (EtBr) accumulation assays; and norA gene expression were all described within silico molecular docking for TQ interactions with norA efflux pump protein. TQ-MICs ranged from 5–320 µg/ml. TQ down-regulated norA gene expression, resulting in a drop in efflux pump activity of 77.5–90.6% in the examined strains, comparable to that observed with verapamil. Exposure of S. aureus strains to CIP and DO raises the initial basal efflux pumping expression to 34.2 and 22.9 times, respectively. This induced efflux pumping overexpression was substantially reduced by 97.7% when TQ was combined with CIP or DO. There was a significant reduction of MICs of CIP and DO MICs by 2–15 and 2–4 folds, respectively, after treatment with 0.5XMIC-TQ in resistance modulation assays. These results refer to TQ ligand inhibitory interactions with NorA protein in molecular docking. Interpretations of inhibition zone diameters (IZDs) of disc diffusion and TQ-MICs exhibit independence of MICs from IZDs, as indicated by invalid linear regression analysis. TQ significantly reduced efflux pumping S. aureus induced by CIP and DO, but further investigations are needed to improve TQ-pharmacokinetics to restore CIP and DO activity and suppress fluoroquinolone and doxycycline-resistant S. aureus selection in clinical and animal settings.

Ccr4-not ubiquitin ligase signaling regulates ribosomal protein homeostasis and inhibits 40S ribosomal autophagy

The Ccr4-Not complex contains the poorly understood Not4 ubiquitin ligase that functions in transcription, mRNA decay, translation, proteostasis, and endolysosomal nutrient signaling. To gain further insight into the in vivo functions of the ligase, we performed quantitative proteomics in Saccharomyces cerevisiae using yeast cells lacking Not4, or cells overexpressing wild-type Not4 or an inactive Not4 mutant. Herein, we provide evidence that balanced Not4 activity maintains ribosomal protein (RP) homeostasis independent of changes to RP mRNA or known Not4 ribosomal substrates. Intriguingly, we also find that Not4 loss activates 40S ribosomal autophagy independently of canonical Atg7-dependent macroautophagy, indicating that microautophagy is responsible. We previously demonstrated that Ccr4-Not stimulates target of rapamycin complex 1 (TORC1) signaling, which activates RP expression and inhibits autophagy, by maintaining vacuole V-ATPase H+ pump activity. Importantly, combining Not4 deficient cells with a mutant that blocks vacuole H+ export fully restores RP expression and increases 40S RP autophagy efficiency. In contrast, restoring TORC1 activity alone fails to rescue either process, indicating that Not4 loss disrupts additional endolysosomal functions that regulate RP expression and 40S autophagy. Analysis of the Not4 regulated proteome reveals increases in endolysosomal and autophagy-related factors that functionally interact with Not4 to control RP expression and affect 40S autophagy. Collectively, our data indicate that balanced Ccr4-Not ubiquitin ligase signaling maintains RP homeostasis and inhibits 40S autophagy via the ligase’s emerging role as an endolysosomal regulator.

P-113 Comparison of mitochondrial function between expanded and hatching human blastocysts

Abstract Study question Is there a difference in mitochondrial function between expanded and hatching fresh and vitrified blastocysts? Summary answer Freezing and thawing may lead to mitochondrial dysfunction. What is known already Mitochondria, as crucial cellular organelles, play role in energy production through the electron transport chain, cell motility and organelle movement. They also have a significant impact on embryonic development. Furthermore, during the process of embryonic hatching, the energy generated by these organelles can affect ion pump activity, blastocyst expansion, and collapse. Therefore, factors that stress embryonic mitochondria may contribute to embryogenesis and hatching. Cooling and warming, as stress factors, maybe result in organelle damage, protein denaturation, membrane disruption, and DNA fragmentation. Additionally, they can cause changes in mitochondrial function, leading to decreased ATP levels and increased ROS levels. Study design, size, duration In this study, vitrified and fresh human 8-cell embryos were collected from April 18, 2021, to July 19, 2023. The embryos were cultured until the blastocyst stage, and then four samples were assessed in four groups. These groups included vitrified expanded blastocysts, vitrified hatching blastocysts, fresh expanded blastocysts, and fresh hatching blastocysts. Participants/materials, setting, methods Vitrified embryos were sourced from the embryo bank of the Royan Institute, while fresh embryos were obtained from good-quality 3PN embryos or excess embryos from couples who did not wish to freeze them. A stock solution containing lipophilic cationic JC-1 dye in dimethyl sulfoxide was prepared. Blastocysts were then exposed to a concentration of 1 μg/ml of JC-1 diluted in HamsF10-HSA. After washing with HF-HSA, imaging was performed using a 2-photon confocal microscope. Main results and the role of chance Our results showed that mitochondrial membrane potential in expanded blastocysts of both fresh and vitrified eight-cell embryos was significantly higher than in hatching blastocysts of fresh and vitrified embryos (P < 0.05). However, there was no significant difference in mitochondrial function between the vitrified and fresh groups, neither in expanded nor hatching blastocysts. Limitations, reasons for caution Due to the human origin of the samples, the minimum necessary replicates were considered for each experimental group. Wider implications of the findings In this study, it appears that plump zona breakers can be detected in hatching blastocysts using confocal microscopy. Previous studies, such as the one conducted by Sathananthan, identified these cells using a TEM microscope. Trial registration number -

Chlorophyllin Supplementation of Medicated or Unmedicated Swine Diets Impact on Fecal Escherichia coli and Enterococci.

Considering that certain catabolic products of anaerobic chlorophyll degradation inhibit efflux pump activity, this study was conducted to test if feeding pigs a water-soluble chlorophyllin product could affect the antibiotic resistance profiles of select wild-type populations of fecal bacteria. Trial 1 evaluated the effects of chlorophyllin supplementation (300 mg/meal) on fecal E. coli and enterococcal populations in pigs fed twice daily diets supplemented without or with ASP 250 (containing chlortetracycline, sulfamethazine and penicillin at 100, 100 and 50 g/ton, respectively). Trial 2, conducted similarly, evaluated chlorophyllin supplementation in pigs fed diets supplemented with or without 100 g tylosin/ton. Each trial lasted 12 days, and fecal samples were collected and selectively cultured at 4-day intervals to enumerate the total numbers of E. coli and enterococci as well as populations of these bacteria phenotypically capable of growing in the presence of the fed antibiotics. Performance results from both studies revealed no adverse effect (p > 0.05) of chlorophyllin, antibiotic or their combined supplementation on average daily feed intake or average daily gain, although the daily fed intake tended to be lower (p = 0.053) for pigs fed diets supplemented with tylosin than those fed diets without tylosin. The results from trial 1 showed that the ASP 250-medicated diets, whether without or with chlorophyllin supplementation, supported higher (p < 0.05) fecal E. coli populations than the non-medicated diets. Enterococcal populations, however, were lower, albeit marginally and not necessarily significantly, in feces from pigs fed the ASP 250-medicated diet than those fed the non-medicated diet. Results from trial 2 likewise revealed an increase (p < 0.05) in E. coli and, to a lesser extent, enterococcal populations in feces collected from pigs fed the tylosin-medicated diet compared with those fed the non-medicated diet. Evidence indicated that the E. coli and enterococcal populations in trial 1 were generally insensitive to penicillin or chlortetracycline, as there were no differences between populations recovered without or with antibiotic selection. Conversely, a treatment x day of treatment interaction observed in trial 2 (p < 0.05) provided evidence, albeit slight, of an enrichment of tylosin-insensitive enterococci in feces from the pigs fed the tylosin-medicated but not the non-medicated diet. Under the conditions of the present study, it is unlikely that chlorophyllin-derived efflux pump inhibitors potentially present in the chlorophyllin-fed pigs were able to enhance the efficacy of the available antibiotics. However, further research specifically designed to optimize chlorophyll administration could potentially lead to practical applications for the swine industry.

Heated environment increases blood pressure drop and postural sway during initial orthostasis in healthy subjects.

We tested the hypothesis that heat stress influences the closed-loop cardio-postural control by an increased blood pressure (BP) drop and postural sway. Fourteen healthy individuals (eight women) performed two orthostatic tests under thermal reference (TC; ~ 24ºC) and HOT (~ 38ºC) conditions. The center-of-pressure (COP) displacements and the electromyography (EMG) activity of the calf muscles (medial gastrocnemius and tibialis anterior) were recorded during the initial orthostasis (ORT onset) after the supine-to-stand challenge. At the same period, BP (beat-to-beat) was continuously monitored, and supine-to-stand variations (∆%) were calculated. Sublingual temperature (Tsl) was measured as a surrogate of internal temperature. Tsl increased in HOT compared to TC (TC 36.5 ± 0.3 vs. HOT 36.7 ± 0.3ºC; p < 0.01). COP distance was greater in HOT compared to TC condition (TC 596.6 ± 242.4 vs. HOT 680.2 ± 249.1mm; p < 0.01). EMG activity of the gastrocnemius decreased in HOT compared to TC condition (TC 95.5 ± 19.8 vs. HOT 78.4 ± 22.8%mV; p = 0.02). EMG of tibialis did not change between TC and HOT (TC 83.5 ± 42.9 vs. HOT 66.1 ± 31.9%mV; p = 0.29). BP showed a greater fall in HOT compared to TC condition (∆%TC -24.5 ± 13.2 vs. ∆%HOT -33.2 ± 20.2%; p = 0.01). Heat stress causes a greater fall in blood pressure and a reduction in musculoskeletal pump activity during orthostatic onset. These effects could be potential mechanisms that underlie augmented postural instability under a heated environment.

Investigation of the role of Cremophor RH 40 and Cremophor EL in the inhibition of efflux pump of Pseudomonas aeruginosa

BackgroundThere is increasing emphasis on restoring the efficacy of existing antibiotics instead of developing new ones. ObjectivesThis study aimed to determine the role of Cremophor EL and Cremophor RH40 in the inhibition of efflux pumps in MDR Pseudomonas aeruginosa strains. MethodsEfflux pump-active MDR strains of P. aeruginosa were identified and confirmed by flow cytometry. The identified efflux-active strains were further subjected to determination of the MIC of ciprofloxacin and the synergistic role of non-ionic surfactants (Cremophor EL and Cremophor RH40) along with ciprofloxacin. ResultsOut of 30 samples, 6 strains displayed high efflux pump activity. Both Cremophor EL and Cremophor RH40 showed efflux pump inhibitory roles. A 4-fold reduction in the MIC values of ciprofloxacin was observed when Cremophor EL was used along with ciprofloxacin, while a 6-fold reduction was observed when Cremophor RH40 was used along with ciprofloxacin. Both compounds showed synergistic effects with ciprofloxacin, ticarcillin and meropenem when used in a 24-well plate efflux pump inhibitory assay. ConclusionThe inhibition of the efflux pump of MDR Pseudomonas aeruginosa by non-ionic surfactants, namely, Cremophor RH40 and Cremophor EL, provided the best strategy to restore the efficacy of ciprofloxacin.

Deletion of VPS50 protein in mouse brain impairs synaptic function and behavior

BackgroundThe VPS50 protein functions in synaptic and dense core vesicle acidification, and perturbations of VPS50 function produce behavioral changes in Caenorhabditis elegans. Patients with mutations in VPS50 show severe developmental delay and intellectual disability, characteristics that have been associated with autism spectrum disorders (ASDs). The mechanisms that link VPS50 mutations to ASD are unknown.ResultsTo examine the role of VPS50 in mammalian brain function and behavior, we used the CRISPR/Cas9 system to generate knockouts of VPS50 in both cultured murine cortical neurons and living mice. In cultured neurons, KO of VPS50 did not affect the number of synaptic vesicles but did cause mislocalization of the V-ATPase V1 domain pump and impaired synaptic activity, likely as a consequence of defects in vesicle acidification and vesicle content. In mice, mosaic KO of VPS50 in the hippocampus altered synaptic transmission and plasticity and generated robust cognitive impairments.ConclusionsWe propose that VPS50 functions as an accessory protein to aid the recruitment of the V-ATPase V1 domain to synaptic vesicles and in that way plays a crucial role in controlling synaptic vesicle acidification. Understanding the mechanisms controlling behaviors and synaptic function in ASD-associated mutations is pivotal for the development of targeted interventions, which may open new avenues for therapeutic strategies aimed at ASD and related conditions.

Computational evaluation of efflux pump homologues and lignans as potent inhibitors against multidrug-resistant Salmonella typhi.

Typhoid fever, caused by Salmonella enterica serovar typhi, presents a substantial global health threat, particularly in regions with limited healthcare infrastructure. The rise of multidrug-resistant strains of S. typhi exacerbates this challenge, severely compromising conventional treatment efficacy due to over activity of efflux pumps. In our study, a comprehensive exploration of two fundamental aspects to combat MDR in S. typhi is carried out; i.e. employing advanced bioinformatics analyses and AlphaFold AI, We successfully identified and characterised a putative homologue, ABC-TPA, reminiscent of the P-glycoprotein (P-gp) known for its role in multidrug resistance in diverse pathogens. This discovery provides a critical foundation for understanding the potential mechanisms driving antibiotic resistance in S. typhi. Furthermore, employing computational methodologies, We meticulously assessed the potential of lignans, specifically Schisandrin A, B, and C, as promising Efflux Pump Inhibitors (EPIs) against the identified P-gp homologue in S. typhi. Noteworthy findings revealed robust binding interactions of Schisandrin A and B with the target protein, indicating substantial inhibitory capabilities. In contrast, Schisandrin C exhibited instability, showing varied effectiveness among the evaluated lignans. Pharmacokinetics and toxicity predictions underscored the favourable attributes of Schisandrin A, including prolonged action duration. Furthermore, high systemic stability and demanished toxicity profile of SA and SB present their therapeutic efficacy against MDR. This comprehensive investigation not only elucidates potential therapeutic strategies against MDR strains of S. typhi but also highlights the relevance of computational approaches in identifying and evaluating promising candidates. These findings lay a robust foundation for future empirical studies to address the formidable challenges antibiotic resistance poses in this clinically significant infectious diseases.

Mechanisms of Helicobacter pylori Resistance to Antibiotics

Background: Helicobacter pylori is the most frequent human bacterial infection worldwide; it is prevalent in approximately half of the world's population. H.pylori poses a major health risk because it possesses distinct virulence factors and highly resistant to antibiotics. Multidrug resistance (MDR) and single-drug resistance are two distinct resistance profiles. Chromosome mutations influence antibiotic activity via target-mediated pathways. Inadequate drug absorption, efflux pumps activity, biofilm development, and cocci formation represent supplementary biological mechanisms by which H.pylori acquires drug resistance. The overexpression of efflux pumps genes in mutant isolates is readily detectable in post-therapy courses. Objective: Characterization of the biological and molecular mechanisms contributing to the development of antibiotic resistance in H.pylori. Conclusion: An alarming rate of drug-resistant H.pylori is on the rise. Primary and acquired resistance to clarithromycin and metronidazole has increased globally. Bacterial factors including biofilms, efflux pumps, and molecular mechanism are in association with H.pylori resistance. It is important to continue to monitor the resistance profiles of H.pylori isolates. Keywords: Helicobacter pylori, MDR, Efflux pump, Biofilm, antibiotic resistance.

Mechanisms of Helicobacter pylori Resistance to Antibiotics

Background: Helicobacter pylori is the most frequent human bacterial infection worldwide; it is prevalent in approximately half of the world's population. H.pylori poses a major health risk because it possesses distinct virulence factors and highly resistant to antibiotics. Multidrug resistance (MDR) and single-drug resistance are two distinct resistance profiles. Chromosome mutations influence antibiotic activity via target-mediated pathways. Inadequate drug absorption, efflux pumps activity, biofilm development, and cocci formation represent supplementary biological mechanisms by which H.pylori acquires drug resistance. The overexpression of efflux pumps genes in mutant isolates is readily detectable in post-therapy courses. Objective: Characterization of the biological and molecular mechanisms contributing to the development of antibiotic resistance in H.pylori. Conclusion: An alarming rate of drug-resistant H.pylori is on the rise. Primary and acquired resistance to clarithromycin and metronidazole has increased globally. Bacterial factors including biofilms, efflux pumps, and molecular mechanism are in association with H.pylori resistance. It is important to continue to monitor the resistance profiles of H.pylori isolates.

Potensi Escherichia Coli Sebagai Resistansi Antibiotik

This literature review examines the increase in antibiotic resistance in Escherichia coli, a Gram-negative bacterium that frequently causes infections in humans and animals. Although most E. coli strains are harmless, some pathogenic strains have shown resistance to various antibiotics, posing a significant challenge in the healthcare field. This study highlights the various resistance mechanisms found in E. coli, including the production of β-lactamase enzymes that can inactivate antibiotics, modification of antibiotic targets, changes in membrane permeability, and increased activity of efflux pumps that expel antibiotics from bacterial cells. Additionally, this review discusses factors contributing to the spread of resistance, such as the excessive and inappropriate use of antibiotics in medicine and livestock, and the transfer of resistance genes through mobile genetic elements. The literature highlights recent research efforts to address this issue, including the development of new antibiotics, the use of antibiotic combinations, and alternative therapies such as bacteriophages and specific enzyme inhibitors. This review emphasizes the importance of strict surveillance and prudent antibiotic use policies to prevent the further spread of antibiotic resistance in E. coli, ensuring the effectiveness of bacterial infection treatments in the future.

Resting membrane potential and intracellular [Na+] at rest, during fatigue and during recovery in rat soleus muscle fibres in situ.

Large trans-sarcolemmal ionic shifts occur with fatiguing exercise or stimulation of isolated muscles. However, it is unknown how resting membrane potential (EM) and intracellular sodium concentration ([Na+]i) change with repeated contractions in living mammals. We investigated (i) whether [Na+]i (peak, kinetics) can reveal changes of Na+-K+ pump activity during brief or fatiguing stimulation and (ii) how resting EM and [Na+]i change during fatigue and recovery of rat soleus muscle in situ. Muscles of anaesthetised rats were stimulated with brief (10s) or repeated tetani (60Hz for 200ms, every 2s, for 30s or 300s) with isometric force measured. Double-barrelled ion-sensitive microelectrodes were used to quantify resting EM and [Na+]i. Post-stimulation data were fitted using polynomials and back-extrapolated to time zero recovery. Mean pre-stimulation resting EM (layer 2-7 fibres) was -71mV (surface fibres were more depolarised), and [Na+]i was 14mM. With deeper fibres, 10s stimulation (2-150Hz) increased [Na+]i to 38-46mM whilst simultaneously causing hyperpolarisations (7.3mV for 2-90Hz). Fatiguing stimulation for 30s or 300s led to end-stimulation resting EM of -61 to -53mV, which recovered rapidly (T1/2, 8-22s). Mean end-stimulation [Na+]i increased to 86-101mM with both fatigue protocols and the [Na+]i recovery time-course (T1/2, 21-35s) showed no difference between protocols. These combined findings suggest that brief stimulation hyperpolarises the resting EM, likely via maximum Na+-induced stimulation of the Na+-K+ pump. Repeated tetani caused massive depolarisation and elevations of [Na+]i that together lower force, although they likely interact with other factors to cause fatigue. [Na+]i recovery kinetics provided no evidence of impaired Na+-K+ pump activity with fatigue. KEY POINTS: It is uncertain how resting membrane potential, intracellular sodium concentration ([Na+]i), and sodium-potassium (Na+-K+) pump activity change during repeated muscle contractions in living mammals. For rat soleus muscle fibres in situ, brief tetanic stimulation for 10s led to raised [Na+]i, anticipated to evoke maximal Na+-induced stimulation of the Na+-K+ pump causing an immediate hyperpolarisation of the sarcolemma. More prolonged stimulation with repeated tetanic contractions causes massive elevations of [Na+]i, which together with large depolarisations (via K+ disturbances) likely reduce force production. These effects occurred without impairment of Na+-K+ pump function. Together these findings suggest that rapid activation of the Na+-K+ pump occurs with brief stimulation to maintain excitability, whereas more prolonged stimulation causes rundown of the trans-sarcolemmal K+ gradient (hence depolarisation) and Na+ gradient, which in combination can impair contraction to contribute to fatigue in living mammals.

Na+/K+-ATPase: More than an Electrogenic Pump.

The sodium pump, or Na+/K+-ATPase (NKA), is an essential enzyme found in the plasma membrane of all animal cells. Its primary role is to transport sodium (Na+) and potassium (K+) ions across the cell membrane, using energy from ATP hydrolysis. This transport creates and maintains an electrochemical gradient, which is crucial for various cellular processes, including cell volume regulation, electrical excitability, and secondary active transport. Although the role of NKA as a pump was discovered and demonstrated several decades ago, it remains the subject of intense research. Current studies aim to delve deeper into several aspects of this molecular entity, such as describing its structure and mode of operation in atomic detail, understanding its molecular and functional diversity, and examining the consequences of its malfunction due to structural alterations. Additionally, researchers are investigating the effects of various substances that amplify or decrease its pumping activity. Beyond its role as a pump, growing evidence indicates that in various cell types, NKA also functions as a receptor for cardiac glycosides like ouabain. This receptor activity triggers the activation of various signaling pathways, producing significant morphological and physiological effects. In this report, we present the results of a comprehensive review of the most outstanding studies of the past five years. We highlight the progress made regarding this new concept of NKA and the various cardiac glycosides that influence it. Furthermore, we emphasize NKA's role in epithelial physiology, particularly its function as a receptor for cardiac glycosides that trigger intracellular signals regulating cell-cell contacts, proliferation, differentiation, and adhesion. We also analyze the role of NKA β-subunits as cell adhesion molecules in glia and epithelial cells.

Analysis of Groundwater Time Series With Limited Pumping Information in Unconfined Aquifer: Response Function Based on Lagging Theory

AbstractGroundwater extraction from aquifers is a common practice for human use, and variations in groundwater levels can provide valuable information on the hydrogeological properties of the aquifer. However, reliable data on pumping rates and distribution are often lacking due to unsupervised groundwater pumping activities. This study presents a new mathematical model for transfer function modeling that depicts the drawdown response caused by pumping in an unconfined aquifer system. To account for the dense and unsupervised pumping events, the uniform pumping approach was used to estimate these effects. To more accurately represent unconfined flow, the model first integrates lagging theory into a response function derived from the Boussinesq equation. The lagging theory accounts for the effects of both inertial force and capillary suction. Furthermore, the model has been used to derive both specific yield and transmissivity along with two lagging parameters simultaneously using only groundwater level information from the Choshui River region in Taiwan. The estimated results suggest that this approach provides reliable estimates of hydrogeological parameters, demonstrating its usefulness for water resource management and water budget evaluation.

Exploration of Antimicrobial Peptides in the Treatment of Gentamicin-Resistant Klebsiella pneumoniae Infection.

The emergence of multidrug-resistant Klebsiella pneumoniae (K. pneumoniae) and the decline of effective antibiotics lead to the urgent need for new antibacterial agents. The aim of this study is to investigate the therapeutic effect of antimicrobial peptides against gentamicin-resistant (RT) K. pneumoniae and to screen effective antimicrobial peptides. In this study, the RT strains were induced by gradient gentamicin, and the RT strains were selected by detecting the expression levels of efflux pump genes, porin genes, and biofilm formation genes of the strains combined with their effects on the cells. Then the effects of four antimicrobial peptides on the efflux pump activity, biofilm formation level and cell condition after infection were detected to explore the effects of antimicrobial peptides on RT strains. Finally, the RT strain was used to induce a mouse model of pneumonia, and the four antimicrobial peptides were used to treat pneumonia mice for in vivo experiments. The pathological changes in lung tissues in each group were detected to explore the antimicrobial peptide with the most significant effect on the RT strain in vivo. The results showed that the minimal inhibitory concentrations of the RT strains (strain C and strain I) were significantly higher than those of the wild-type strain, and the expression of efflux pump, porin and biofilm formation genes was significantly increased. The antimicrobial peptides could effectively inhibit the biofilm formation and efflux pump protein function of the RT strains. In addition, the antimicrobial peptides showed promising antibacterial effects both in vitro and in vivo. Our study provided a theoretical basis for the treatment of gentamicin resistant K. pneumoniae infection with antimicrobial peptides, and found that KLA was significantly superior to LL37, Magainin I, KLA and Dermaseptin (10 μg/mL in cells, 50 μg in mice).