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Mitochondrial membrane potential directly powers many critical functions of mitochondria, including ATP production, mitochondrial protein import, and metabolite transport. Its loss is a cardinal feature of aging and mitochondrial diseases, and cells closely monitor membrane potential as an indicator of mitochondrial health. Given its central importance, it is logical that cells would modulate mitochondrial membrane potential in response to demand and environmental cues, but there has been little exploration of this question. We report that loss of the Sit4 protein phosphatase in yeast increases mitochondrial membrane potential, both by inducing the electron transport chain and the phosphate starvation response. Indeed, a similarly elevated mitochondrial membrane potential is also elicited simply by phosphate starvation or by abrogation of the Pho85-dependent phosphate sensing pathway. This enhanced membrane potential is primarily driven by an unexpected activity of the ADP/ATP carrier. We also demonstrate that this connection between phosphate limitation and enhancement of mitochondrial membrane potential is observed in primary and immortalized mammalian cells as well as in Drosophila. These data suggest that mitochondrial membrane potential is subject to environmental stimuli and intracellular signaling regulation and raise the possibility for therapeutic enhancement of mitochondrial function even in defective mitochondria.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a 'highly transmissible respiratory pathogen, leading to severe multi-organ damage. However, knowledge regarding SARS-CoV-2-induced cellular alterations is limited. In this study, we report that SARS-CoV-2 aberrantly elevates mitochondrial bioenergetics and activates the EGFR-mediated cell survival signal cascade during the early stage of viral infection. SARS-CoV-2 causes an increase in mitochondrial transmembrane potential via the SARS-CoV-2 RNA-nucleocapsid cluster, thereby abnormally promoting mitochondrial elongation and the OXPHOS process, followed by enhancing ATP production. Furthermore, SARS-CoV-2 activates the EGFR signal cascade and subsequently induces mitochondrial EGFR trafficking, contributing to abnormal OXPHOS process and viral propagation. Approved EGFR inhibitors remarkably reduce SARS-CoV-2 propagation, among which vandetanib exhibits the highest antiviral efficacy. Treatment of SARS-CoV-2-infected cells with vandetanib decreases SARS-CoV-2-induced EGFR trafficking to the mitochondria and restores SARS-CoV-2-induced aberrant elevation in OXPHOS process and ATP generation, thereby resulting in the reduction of SARS-CoV-2 propagation. Furthermore, oral administration of vandetanib to SARS-CoV-2-infected hACE2 transgenic mice reduces SARS-CoV-2 propagation in lung tissue and mitigates SARS-CoV-2-induced lung inflammation. Vandetanib also exhibits potent antiviral activity against various SARS-CoV-2 variants of concern, including alpha, beta, delta and omicron, in in vitro cell culture experiments. Taken together, our findings provide novel insight into SARS-CoV-2-induced alterations in mitochondrial dynamics and EGFR trafficking during the early stage of viral infection and their roles in robust SARS-CoV-2 propagation, suggesting that EGFR is an attractive host target for combating COVID-19.

[Display omitted] • LTD occurs reliably at cortical inputs into the amygdala. • The plasticity of synaptic connections is an evolving and dynamic process. • Pyramidal neurons exhibit similar LTD despite different spiking patterns. • The LTD in the amygdala is progressively stronger upon recurrent stimuli. • Gradually stronger LTD exhibits similitude to that of LTP. The convergence of conditioned and unconditioned stimuli (CS and US) into the lateral amygdala (LA) serves as a substrate for an adequate fear response in vivo. This well-known Pavlovian paradigm modulates the synaptic plasticity of neurons, as can be proved by the long-term potentiation (LTP) phenomenon in vitro. Although there is an increasing body of evidence for the existence of LTP in the amygdala, only a few studies were able to show a reliable long-term depression (LTD) of excitation in this structure. We have used coronal brain slices and conducted patch-clamp recordings in pyramidal neurons of the lateral amygdala (LA). After obtaining a stable baseline excitatory postsynaptic current (EPSC) response at a holding potential of −70 mV, we employed a paired-pulse paradigm at 1 Hz at the same membrane potential and could observe a reliable LTD. The different durations of stimulation (ranging between 1.5–24 min) were tested first in the same neuron, but the intensity was kept constant. The latter paradigm resulted in a step-wise LTD with a gradually increasing magnitude under these conditions. [ABSTRACT FROM AUTHOR]

Background and Purpose: The AMP‐activated protein kinase (AMPK) signalling pathway is a desirable target for various cardiovascular diseases (CVD), while the involvement of AMPK‐mediated specific downstream pathways and effective interventions in hyperlipidaemia‐induced endothelial dysfunction remain largely unknown. Herein, we aim to identify an effective AMPK activator and to explore its efficacy and mechanism against endothelial dysfunction. Experimental Approach: Molecular docking technique was adopted to screen for the potent AMPK activator among 11 most common rare ginsenosides. In vivo, poloxamer 407 (P407) was used to induce acute hyperlipidaemia in C57BL/6J mice. In vitro, palmitic acid (PA) was used to induce lipid toxicity in HAEC cells. Key Results: We discovered the strongest binding of ginsenoside Rh4 to AMPKα1 and confirmed the action of Rh4 on AMPK activation. Rh4 effectively attenuated hyperlipidaemia‐related endothelial injury and oxidative stress both in vivo and in vitro and restored cell viability, mitochondrial membrane potential and mitochondrial oxygen consumption rate in HAEC cells. Mechanistically, Rh4 bound to AMPKα1 and simultaneously up‐regulated AKT/eNOS‐mediated NO release, promoted PGC‐1α‐mediated mitochondrial biogenesis and inhibited P38 MAPK/NFκB‐mediated inflammatory responses in both P407‐treated mice and PA‐treated HAEC cells. The AMPK inhibitor Compound C treatment completely abrogated the regulation of Rh4 on the above pathways and weakened the lowering effect of Rh4 on endothelial impairment markers, suggesting that the beneficial effects of Rh4 are AMPK dependent. Conclusion and Implications: Rh4 may serve as a novel AMPK activator to protect against hyperlipidaemia‐induced endothelial dysfunction, providing new insights into the prevention and treatment of endothelial injury‐associated CVD. [ABSTRACT FROM AUTHOR]

Photodynamic therapy (PDT) utilizes reactive oxygen species (ROS) for eradication of cancer cells. Its effectiveness is governed by the oxygen content, which is scarce in the hypoxic tumor microenvironment. We report herein two zinc(II) phthalocyanines substituted with two or four nitric oxide (NO)‐releasing moieties, namely ZnPc‐2NO and ZnPc‐4NO, which can suppress the mitochondrial respiration, thereby sparing more intracellular oxygen for PDT. Using HT29 human colorectal adenocarcinoma cells and A549 human lung carcinoma cells, we have demonstrated that both conjugates release NO upon interaction with the intracellular glutathione, which can reduce the cellular oxygen consumption rate and adenosine triphosphate generation and alter the mitochondrial membrane potential. They can also relieve the hypoxic status of cancer cells and decrease the expression of hypoxia‐inducible factor protein HIF‐1α. Upon light irradiation, both conjugates can generate ROS and induce cytotoxicity even under a hypoxic condition, overcoming the oxygen‐dependent nature of PDT. Interestingly, the photodynamic action of ZnPc‐2NO elicits the release of damage‐associated molecular patterns, inducing the maturation of dendritic cells and triggering an antitumor immune response. The immunogenic cell death caused by this oxygen‐economized PDT has been demonstrated through a series of in vitro and in vivo experiments. [ABSTRACT FROM AUTHOR]

Cells depend on a continuous supply of adenosine triphosphate (ATP), the universal energy currency. In mitochondria, ATP is produced by a series of redox reactions, whereby an electrochemical gradient is established across the inner mitochondrial membrane. The ATP synthase harnesses the energy of the gradient to generate ATP from adenosine diphosphate (ADP) and inorganic phosphate. We determined the structure of ATP synthase within mitochondria of the unicellular flagellate Polytomella by electron cryo-tomography and subtomogram averaging at up to 4.2-angstrom resolution, revealing six rotary positions of the central stalk, subclassified into 21 substates of the F1 head. The Polytomella ATP synthase forms helical arrays with multiple adjacent rows defining the cristae ridges. The structure of ATP synthase under native operating conditions in the presence of a membrane potential represents a pivotal step toward the analysis of membrane protein complexes in situ. [ABSTRACT FROM AUTHOR]

AbstractVenetoclax (VEN), a B-cell lymphoma 2 (BCL-2) selective inhibitor, is widely used for treating acute myeloid leukemia (AML) with promising results. However, the anti-leukemic effect of VEN in relapsed/refractory (R/R)- AML requires improvement. In this study, we observed that combining homoharringtonine (HHT) with VEN plus azacitidine resulted in a significantly higher response and better survival than VA alone in patients with R/R-AML. Basic research indicates that HHT combined with VEN has a highly synergistic effect against both resistant AML cells and primary cells with/without mesenchymal stem cell (MSC) co-culture in vivo, inhibiting proliferation and colony-forming capacity of AML cells associated with concomitant cell cycle arrest. Mechanistically, HHT sensitizes AML cells to VEN by downregulating the anti-apoptotic proteins MCL-1/BCL-xL, activating reactive oxygen species (ROS), leading to mitochondrial membrane potential loss, and attenuating fatty acid (FA) uptake. These findings adding HHT to VEN-based regimens may enhance outcomes in R/R-AML patients. [ABSTRACT FROM AUTHOR]

Repressing BET proteins' function using bromodomain inhibitors (BETi) has been shown to elicit antitumor effects by regulating the transcription of genes downstream of BRD4. We previously showed that BETi promoted cell death of triple‐negative breast cancer (TNBC) cells. Here, we proved that BETi induce altered mitochondrial dynamics fitness in TNBC cells falling in cell death. We demonstrated that BETi treatment downregulated the expression of BCL‐2, and proteins involved in mitochondrial fission and increased fused mitochondria. Impaired mitochondrial fission affected oxidative phosphorylation (OXPHOS) inducing the expression of OXPHOS‐related genes, SDHa and ATP5a, and increased cell death. Consistently, the amount of mitochondrial DNA and mitochondrial membrane potential (∆Ψm) increased in BETi‐treated cells compared to control cells. Lastly, BETi in combination with Metformin reduced cell growth. Our results indicate that mitochondrial dynamics and OXPHOS metabolism support breast cancer proliferation and represent novel BETi downstream targets in TNBC cells. [ABSTRACT FROM AUTHOR]

AbstractThis study aimed to investigate the effects and molecular mechanism of PF on high glucose (HG)-induced podocyte injury. Results found that PF increased proliferation activity, decreased apoptosis, LDH, and caspase-3 levels, and increased nephrin and podocin expression in HG-induced cells. Similarly, PF improved HG-induced mitochondrial damage, decreased Ca2+ and ROS content, alleviated oxidative stress, inhibited mPTP opening, increased mitochondrial membrane potential, and decreased the expressions of Drp1, Bak, Bax, and Cytc in cytoplasm, increased the expressions of SIRT1, PGC-1α, HSP70, HK2, and Cytc in mitochondria of podocytes. The use of mPTP agonist/blocker and SIRT1 inhibitor confirmed that PF alleviates HG-induced podocyte injury by regulating mitochondrial mPTP opening through SIRT1/PGC-1α. In addition, PF affected HK2-VDAC1 protein binding to regulate mPTP opening via the SIRT1/PGC-1α pathway. In conclusion, PF-regulated HK2-VDAC1 protein binding affected mitochondrial mPTP opening and improved HG-induced podocyte injury through the SIRT1/PGC-1α pathway. [ABSTRACT FROM AUTHOR]

Tenofovir alafenamide (TAF) is a new drug from the nucleotide reverse transcriptase inhibitor group approved for the treatment of chronic Hepatitis B in 2016. With this study, we aimed to test whether possible cellular toxicity can be reduced by controlled drug release as a result of loading with chitosan nanoparticles (CHS). We investigated the genotoxic and mitotoxic effects of 45 µM TAF-loaded CHS and TAF-only on HepG2 cells by micronucleus (MN), comet assay, determination of mtDNA quantification, mitochondrial membrane potential (ΔΨm), and ROS levels. Additionally, we compared the samples by RNAseq analyses to reveal the transcriptional responses to each regimen. In terms of genotoxic tests, although MN and comet were found higher in all experimental treatment conditions, the encapsulation of CHS reduced the genotoxicity of TAF. MtDNA level was found to be lower in the TAF treatment, whereas it was higher in CHS and CHS-TAF treatments. The TAF-loaded CHS and TAF treatments had an impaired ΔΨm value. Cellular ROS levels were higher in all treatment conditions. According to the analyses of gene expression patterns; CHS-only changed the expression of relatively few genes (187 genes), while TAF changed the expression of the 1974 genes and TAF-loaded CHS changed the expression of 734 genes. Considering the gene expression numbers, CHS encapsulation of TAF significantly reduced the number of genes that were differentially expressed by TAF-only. Overall, we observed that TAF has genotoxic and mitotoxic effects on HepG2 cells, and upon encapsulation with CHS, its genotoxic and mitotoxic effects were decreased. [ABSTRACT FROM AUTHOR]

Myocardial infarction (MI) is a significant cause of death in diabetic patients. Growing evidence suggests that mitochondrial dysfunction contributes to heart failure in diabetes. However, the molecular mechanisms of mitochondrial dysfunction mediating heart failure in diabetes are still poorly understood. We examined MRPL12 levels in right atrial appendage tissues from diabetic patients undergoing coronary artery bypass graft (CABG) surgery. Using AC-16 cells overexpressing MRPL12 under normal and hyperglycemic conditions we performed mitochondrial functional assays OXPHOS, bioenergetics, mitochondrial membrane potential, ATP production and cell death. We observed elevated MRPL12 levels in heart tissue samples from diabetic patients with ischemic heart disease compared to non-diabetic patients. Overexpression of MRPL12 under hyperglycemic conditions did not affect oxidative phosphorylation (OXPHOS) levels, cellular ATP levels, or cardiomyocyte cell death. However, notable impairment in mitochondrial membrane potential (MMP) was observed under hyperglycemic conditions, along with alterations in both basal respiration oxygen consumption rate (OCR) and maximal respiratory capacity OCR. Overall, our results suggest that MRPL12 may have a compensatory role in the diabetic myocardium with ischemic heart disease, suggesting that MRPL12 may implicate in the pathophysiology of MI in diabetes. [Display omitted] • MRPL12 levels were increased in the Right atrial appendages (RAA) tissues of human diabetic patients undergoing CABG. • MRPL12 overexpression in AC-16 cardiomyocytes under hyperglycemic conditions affects mitochondrial membrane potential and bioenergetics, however oxidative phosphorylation was not significantly altered nor were ATP production or cell death. • Increased expression of MRPL12 may act as a compensatory mechanism in diabetic cardiomyocytes. [ABSTRACT FROM AUTHOR]

Doxorubicin (Dox) use is limited by Dox-induced cardiotoxicity. TANK-blinding kinase 1 (TBK1) is an important kinase involved in the regulation of mitophagy, but the role of TBK1 in cardiomyocytes in chronic Dox-induced cardiomyopathy remains unclear. Cardiomyocyte-specific Tbk1 knockout (Tbk1 CKO) mice received Dox (6 mg/kg, injected intraperitoneally) once a week for 4 times, and cardiac assessment was performed 4 weeks after the final Dox injection. Adenoviruses encoding Tbk1 or containing shRNA targeting Tbk1 , or a TBK1 phosphorylation inhibitor were used for overexpression or knockdown of Tbk1 , or inhibit phosphorylation of TBK1 in isolated primary cardiomyocytes. Our results revealed that moderate Dox challenge decreased TBK1 phosphorylation (with no effect on TBK1 protein levels), resulting in compromised myocardial function, obvious mortality and overt interstitial fibrosis, and the effects were accentuated by Tbk1 deletion. Dox provoked mitochondrial membrane potential collapse and oxidative stress, the effects of which were exacerbated and mitigated by Tbk1 knockdown, specific inhibition of phosphorylation and overexpression, respectively. However, Tbk1 （Ser172A） overexpression did not alleviate these effects. Further scrutiny revealed that TBK1 exerted protective effects on mitochondria via SQSTM1/P62-mediated mitophagy. Tbk1 overexpression mediated cardioprotective effects on Dox-induced cardiotoxicity were cancelled off by Sqstm1/P62 knockdown. Moreover, TBK1-mitophagy-mitochondria cascade was confirmed in heart tissues from dilated cardiomyopathy patients. Taken together, our findings denoted a pivotal role of TBK1 in Dox-induced mitochondrial injury and cardiotoxicity possibly through its phosphorylation and SQSTM1/P62-mediated mitophagy. [Display omitted] • Cardiomyocyte-specific Tbk1 deletion aggravated chronic Doxorubicin-induced cardiotoxicity. • Specific inhibition of TBK1（Ser172） phosphorylation exacerbated the cardiomyocyte toxicity of Doxorubicin. • Overexpression of Tbk1 could mitigate the cardiotoxicity of Doxorubicin, while overexpression of Tbk1 (Ser172A) could not. • The TBK1-P62-mitophagy cascade may play a crucial role in the pathogenesis of Doxorubicin-induced cardiotoxicity. [ABSTRACT FROM AUTHOR]

This article discusses the functional properties of high-affinity K+ transporters (HKTs) in plants, specifically TmHKT1;4-A2 from einkorn and OsHKT2;2 from rice. The researchers used cell-free expression and artificial lipid bilayer membranes to study the transporters at the single-protein level. They found that both TmHKT1;4-A2 and OsHKT2;2 displayed channel-like gating mechanisms and unitary conductances, indicating that they can function as coordinated units. This research provides valuable insights into the mechanisms of salt tolerance in plants and could contribute to the development of crops with improved salt tolerance. [Extracted from the article]

As the aging population continues to increase, aging-related inflammation, oxidative stress, and neurodegenerative diseases have become serious global health threats. Resveratrol, a star molecule in natural polyphenols, has been widely reported to have physiological activities such as anti-aging, anti-inflammatory, antioxidant, and neuroprotection. However, its poor water solubility, rapid metabolism, low bioavailability and poor targeting ability, which limits its application. Accordingly, a brain-targeted resveratrol liposome (ANG-RES-LIP) was developed to solve these issues. Experimental results showed that ANG-RES-LIP has a uniform size distribution, good biocompatibility, and a drug encapsulation rate of over 90%. Furthermore, in vitro cell experiments showed that the modification of the targeting ligand ANG significantly increased the capability of RES to cross the BBB and neuronal uptake. Compared with free RES, ANG-RES-LIP demonstrated stronger antioxidant activity and the ability to rescue oxidatively damaged cells from apoptosis. Additionally, ANG-RES-LIP showed the ability to repair damaged neuronal mitochondrial membrane potential. In vivo experiments further demonstrated that ANG-RES-LIP improved cognitive function by reducing oxidative stress and inflammation levels in the brains of aging model mice, repairing damaged neurons and glial cells, and increasing brain-derived neurotrophic factor. In summary, this study not only provides a new method for further development and application of resveratrol but also a promising strategy for preventing and treating age-related neurodegenerative diseases. [ABSTRACT FROM AUTHOR]

Summary: In this work, deferoxamine (DFO) was injected into Tibetan sheep meat (distilled water as control) to reveal the effects of lysosomal Fe2+ on mitochondrial apoptosis and muscle tenderness. The reactive oxygen species (ROS) content, lysosomal membrane stability, cathepsins activity, mitochondrial membrane permeability transition pore (MPTP), mitochondrial membrane potential (MMP), cytochrome c (Cyt‐c) reduction level, caspase activity, myofibril fragmentation index (MFI) and shear force of Oula Tibetan sheep meat were measured at different ageing times. The results indicated that the ROS content, cathepsins activity, MPTP opening degree, lysosomal membrane stability, caspase‐3/9 activity and MFI of Tibetan sheep meat were significantly decreased after DFO treatment, whilst MMP, shear force and Cyt‐c reduction levels were noticeably increased. Furthermore, correlation analysis and principal component analysis showed that mitochondrial damage was positively correlated with meat tenderness. In sum, the study suggests that lysosomal Fe2+ may promote mitochondrial apoptosis and improve the tenderness of Tibetan sheep meat. [ABSTRACT FROM AUTHOR]

The in vitro maturation (IVM) quality of oocytes is directly related to the subsequent developmental potential of embryos and a fundamental of in vitro embryo production. However, conventional IVM methods fail to maintain the gap-junction intercellular communication (GJIC) between cumulus–oocyte complexes (COCs), which leads to insufficient oocyte maturation. Herein, we investigated the effects of three different three-dimensional (3D) culture methods on oocyte development in vitro , optimized of the alginate-hydrogel embedding method, and assessed the effects of the alginate-hydrogel embedding method on subsequent embryonic developmental potential of oocytes after IVM and parthenogenetic activation (PA). The results showed that Matrigel embedding and alginate-hydrogel embedding benefited the embryonic developmental potential of oocytes after IVM and PA. With the further optimization of alginate-hydrogel embedding, including crosslinking and decrosslinking of parameters, we established a 3D culture system that can significantly increase oocyte maturation and the blastocyst rate of embryos after PA (27.2 ± 1.5 vs 36.7 ± 2.8, P < 0.05). This 3D culture system produced oocytes with markedly increased mitochondrial intensity and membrane potential, which reduced the abnormalities of spindle formation and cortical granule distribution. The alginate-hydrogel embedding system can also remarkably enhance the GJIC between COCs. In summary, based on alginate-hydrogel embedding, we established a 3D culture system that can improve the IVM quality of porcine oocytes, possibly by enhancing GJIC. [ABSTRACT FROM AUTHOR]

In this study, we aimed to investigate cytoplasmic maturation and miRNA expression of mature oocytes cultured in porcine follicular fluid exosomes. We also examined the effect of miR-339-5p on oocyte maturation. Twenty eight differentially expressed miRNAs were detected using miRNA-seq. We then transfected cumulus oocyte complexes with miR-339-5p mimics and inhibitor during culture. The results showed that exosomes increased endoplasmic reticulum levels and the amount of lipid droplets, and decreased ROS levels, lipid droplet size, and percentage of oocytes with abnormal cortical granule distribution. Overexpressing miR-339-5p significantly decreased cumulus expansion genes, oocyte maturation-related genes, target gene proline/glutamine-rich splicing factor (SFPQ), ERK1/2 phosphorylation levels, oocyte maturation rate, blastocyst rate, and lipid droplet number, but increased lipid droplet size and the ratio of oocytes with abnormal cortical granule distribution. Inhibiting miR-339-5p reversed the decrease observed during overexpression. Mitochondrial membrane potential and ROS levels did not differ significantly between groups. In summary, exosomes promote oocyte cytoplasmic maturation and miR-339-5p regulating ERK1/2 activity through SFPQ expression, thereby elevating oocyte maturation and blastocyst formation rate in vitro. • This study examined the mechanisms behind enhanced in vitro oocyte maturation in pigs with follicular fluid exosomes. • MiRNA sequencing experiments demonstrated that miR-339-5p was differentially expressed between treatment groups. • MiR-339-5p regulated the ERK1/2 pathway through SFPQ and improved various measures of oocyte maturation. [ABSTRACT FROM AUTHOR]

Post-mitotic, non-proliferative dermal fibroblasts have crucial functions in maintenance and restoration of tissue homeostasis. They are involved in essential processes such as wound healing, pigmentation and hair growth, but also tumor development and aging-associated diseases. These processes are energetically highly demanding and error prone when mitochondrial damage occurs. However, mitochondrial function in fibroblasts and the influence of mitochondrial dysfunction on fibroblast-specific demands are still unclear. To address these questions, we created a mouse model in which accelerated cell-specific mitochondrial DNA (mtDNA) damage accumulates. We crossed mice carrying a dominant-negative mutant of the mitochondrial replicative helicase Twinkle (RosaSTOP system) with mice that express fibroblast-specific Cre Recombinase (Collagen1A2 CreERT) which can be activated by Tamoxifen (TwinkleFIBRO). Thus, we are able to induce mtDNA deletions and duplications in specific cells, a process which resembles the physiological aging process in humans, where this damage accumulates in all tissues. Upon proliferation in vitro , Tamoxifen induced Twinkle fibroblasts deplete most of their mitochondrial DNA which, although not disturbing the stoichiometry of the respiratory chain complexes, leads to reduced ROS production and mitochondrial membrane potential as well as an anti-inflammatory and anti-fibrotic profile of the cells. In Sodium Azide treated wildtype fibroblasts, without a functioning respiratory chain, we observe the opposite, a rather pro-inflammatory and pro-fibrotic signature. Upon accumulation of mitochondrial DNA mutations in vivo the TwinkleFIBRO mice are protected from fibrosis development induced by intradermal Bleomycin injections. This is due to dampened differentiation of the dermal fibroblasts into α−smooth-muscle-actin positive myofibroblasts in TwinkleFIBRO mice. We thus provide evidence for striking differences of the impact that mtDNA mutations have in contrast to blunted mitochondrial function in dermal fibroblasts and skin homeostasis. These data contribute to improved understanding of mitochondrial function and dysfunction in skin and provide mechanistic insight into potential targets to treat skin fibrosis in the future. [Display omitted] [ABSTRACT FROM AUTHOR]

Tert-butyl hydroperoxide (t-BuOOH) is an organic hydroperoxide widely used as a model compound to induce oxidative stress. It leads to a plethora of cellular damage, including lipid peroxidation, DNA double-strand breaks (DNA DSBs), and breakdown of the mitochondrial membrane potential (MMP). We could show in several cell lines that t-BuOOH induces ferroptosis, triggered by iron-dependent lipid peroxidation. We have further revealed that not only t-BuOOH-mediated ferroptosis, but also DNA DSBs and loss of MMP are prevented by cell–cell contacts. The underlying mechanisms are not known. Here, we show in murine fibroblasts and a human colon carcinoma cell line that t-BuOOH (50 or 100 µM, resp.) causes an increase in intracellular Ca2+, and that this increase is key to lipid peroxidation and ferroptosis, DNA DSB formation and dissipation of the MMP. We further demonstrate that cell–cell contacts prevent t-BuOOH-mediated raise in intracellular Ca2+. Hence, we provide novel insights into the mechanism of t-BuOOH-triggered cellular damage including ferroptosis and propose a model in which cell–cell contacts control intracellular Ca2+ levels to prevent lipid peroxidation, DNA DSB-formation and loss of MMP. Since Ca2+ is a central player of toxicity in response to oxidative stress and is involved in various cell death pathways, our observations suggest a broad protective function of cell–cell contacts against a variety of exogenous toxicants. [ABSTRACT FROM AUTHOR]

Sevoflurane (Sev) is a commonly used inhalation anaesthetic that has been shown to cause hippocampus dysfunction through multiple underlying molecular processes, including mitochondrial malfunction, oxidative stress and inflammation. Dihydromyricetin (DHM) is a 2,3‐dihydroflavonoid with various biological properties, such as anti‐inflammation and anti‐oxidative stress. The purpose of this study was to investigate the effect of DHM on Sev‐induced neuronal dysfunction. HT22 cells were incubated with 10, 20 and 30 μM of DHM for 24 h, and then stimulated with 4% Sev for 6 h. The effects and mechanism of DHM on inflammation, oxidative stress and mitochondrial dysfunction were explored in Sev‐induced HT22 cells by Cell Counting Kit‐8, flow cytometry, enzyme‐linked immunosorbent assay, reverse transcription‐quantitative polymerase chain reaction, colorimetric detections, detection of the level of reactive oxygen species (ROS), mitochondrial ROS and mitochondrial membrane potential (MMP), immunofluorescence and western blotting. Our results showed that DHM increased Sev‐induced cell viability of HT22 cells. Pretreatment with DHM attenuated apoptosis, inflammation, oxidative stress and mitochondrial dysfunction in Sev‐elicited HT22 cells by remedying the abnormality of the indicators involved in these progresses, including apoptosis rate, the cleaved‐caspase 3 expression, as well as the level of tumour necrosis factor α, interleukin (IL)‐1β, IL‐6, malondialdehyde, superoxide dismutase, catalase, ROS, mitochondrial ROS and MMP. Mechanically, pretreatment with DHM restored the Sev‐induced the expression of SIRT1/FOXO3a pathway in HT22 cells. Blocking of SIRT1 counteracted the mitigatory effect of DHM on apoptosis, inflammation, oxidative stress and mitochondrial dysfunction in Sev‐elicited HT22 cells. Collectively, pretreatment with DHM improved inflammation, oxidative stress and mitochondrial dysfunction via SIRT1/FOXO3a pathway in Sev‐induced HT22 cells. [ABSTRACT FROM AUTHOR]

Pheochromocytoma (PCC) and abdominal paraganglioma (aPGL) (together abbreviated PPGL) frequently present with an underlying genetic event in a PPGL driver gene, and additional susceptibility genes are anticipated. Here, we re-analyzed whole-exome sequencing data for PCC patients and identified two patients with rare missense variants in the calcium voltage-gated channel subunit 1H gene (CACNA1H). CACNA1H variants were also found in the clinical setting in PCC patients using targeted sequencing and from analysis of The Cancer Genome Atlas database. In total, CACNA1H variants were found in six PCC cases. Three of these were constitutional, and two are known to have functional consequences on hormone production and gene expression in primary aldosteronism and aldosteroneproducing adrenocortical adenoma. In general, PPGL exhibited reduced CACNA1H mRNA expression as compared to normal adrenal. Immunohistochemistry showed strong CACNA1H (CaV3.2) staining in adrenal medulla while PPGL typically had weak or negative staining. Reduced CACNA1H gene expression was especially pronounced in PCC compared to aPGL and in PPGL with cluster 2 kinase signaling phenotype. Furthermore, CACNA1H levels correlated with HIF1A and HIF2A. Moreover, TCGA data revealed a correlation between CACNA1H methylation density and gene expression. Expression of rCacna1h in PC12 cells induced differential protein expression profiles, determined by mass spectrometry, as well as a shift in the membrane potential where maximum calcium currents were observed, as determined by electrophysiology. The findings suggest the involvement of CACNA1H/CaV3.2 in pheochromocytoma development and establish a potential link between the etiology of adrenomedullary and adrenocortical tumor development. [ABSTRACT FROM AUTHOR]

Objective: We aimed to broaden our understanding of a potential interaction between B1R and TLR4, considering earlier studies suggesting that lipopolysaccharide (LPS) may trigger B1R stimulation. Methods: We assessed the impact of DBK and LPS on the membrane potential of thoracic aortas from C57BL/6, B1R, or TLR4 knockout mice. Additionally, we examined the staining patterns of these receptors in the thoracic aortas of C57BL/6 and in endothelial cells (HBMEC). Results: DBK does not affect the resting membrane potential of aortic rings in C57BL/6 mice, but it hyperpolarizes preparations in B1KO and TLR4KO mice. The hyperpolarization mechanism in B1KO mice involves B2R, and the TLR4KO response is independent of cytoplasmic calcium influx but relies on potassium channels. Conversely, LPS hyperpolarizes thoracic aorta rings in both C57BL/6 and B1KO mice, with the response unaffected by a B1R antagonist. Interestingly, the absence of B1R alters the LPS response to potassium channels. These activities are independent of nitric oxide synthase (NOS). While exposure to DBK and LPS does not alter B1R and TLR4 mRNA expression, treatment with these agonists increases B1R staining in endothelial cells of thoracic aortic rings and modifies the staining pattern of B1R and TLR4 in endothelial cells. Proximity ligation assay suggests a interaction between the receptors. Conclusion: Our findings provide additional support for a putative connection between B1R and TLR4 signaling. Given the involvement of these receptors and their agonists in inflammation, it suggests that drugs and therapies targeting their effects could be promising therapeutic avenues worth exploring. [ABSTRACT FROM AUTHOR]

Plasma membrane H+-ATPases (PMAs) provide an H+ electrochemical gradient to establish a proton motive force and membrane potential for the transport of various nutrients across the plasma membrane. PMAs are regulated at multiple levels, and genetic modulation of PMA expression could enhance nutrient acquisition and use efficiency in plants. PMAs are involved in the coordination of the uptake and assimilation of both nitrate and ammonium. PMAs are involved in nutrient utilization by regulating root growth, facilitating organic anion exudation, and promoting the acquisition of nutrients through H+ extrusion-mediated apoplast and rhizosphere acidification. PMAs play essential roles in mycorrhizal symbioses in plant roots for nutrient acquisition. Plasma membrane H+-ATPases (PMAs) pump H+ out of the cytoplasm by consuming ATP to generate a membrane potential and proton motive force for the transmembrane transport of nutrients into and out of plant cells. PMAs are involved in nutrient acquisition by regulating root growth, nutrient uptake, and translocation, as well as the establishment of symbiosis with arbuscular mycorrhizas. Under nutrient stresses, PMAs are activated to pump more H+ and promote organic anion excretion, thus improving nutrient availability in the rhizosphere. Herein we review recent progress in the physiological functions and the underlying molecular mechanisms of PMAs in the efficient acquisition and utilization of various nutrients in plants. We also discuss perspectives for the application of PMAs in improving crop production and quality. [ABSTRACT FROM AUTHOR]

Metal complexes based on N‐heterocyclic carbene (NHC) ligands have emerged as promising broad‐spectrum antitumor agents in bioorganometallic medicinal chemistry. In recent decades, studies on cytotoxic metal–NHC complexes have yielded numerous compounds exhibiting superior cytotoxicity compared to cisplatin. Although the molecular mechanisms of these anticancer complexes are not fully understood, some potential targets and modes of action have been identified. However, a comprehensive review of their biological mechanisms is currently absent. In general, apoptosis caused by metal–NHCs is common in tumor cells. They can cause a series of changes after entering cells, such as mitochondrial membrane potential (MMP) variation, reactive oxygen species (ROS) generation, cytochrome c (cyt c) release, endoplasmic reticulum (ER) stress, lysosome damage, and caspase activation, ultimately leading to apoptosis. Therefore, a detailed understanding of the influence of metal–NHCs on cancer cell apoptosis is crucial. In this review, we provide a comprehensive summary of recent advances in metal–NHC complexes that trigger apoptotic cell death via different apoptosis‐related targets or signaling pathways, including B‐cell lymphoma 2 (Bcl‐2 family), p53, cyt c, ER stress, lysosome damage, thioredoxin reductase (TrxR) inhibition, and so forth. We also discuss the challenges, limitations, and future directions of metal–NHC complexes to elucidate their emerging application in medicinal chemistry. [ABSTRACT FROM AUTHOR]

BACKGROUND: Owing to the nonavailability of any clear targets for molluscicides against Pomacea canaliculata, target‐based screening strategy cannot be employed. In this study, the molluscicidal effects of typical pesticides on P. canaliculata were evaluated to obtain the molluscicide target. A series of arylpyrrole compounds were synthesized based on the discovered target, and their structure–activity relationships explored. A preliminary strategy for screening molluscicides based on specific targets was also developed. RESULTS: A laboratory colony of P. canaliculata was developed, which showed no difference in sensitivity to niclosamide compared with the wild group, while exhibiting a higher stability against pesticide response. Mitochondrial adenosine triphosphate (ATP) synthase inhibitors and mitochondrial membrane potential uncouplers were identified and validated as potential targets for molluscicide screening against P. canaliculata. A series of arylpyrrole compounds were designed and synthesized. The median lethal concentration of 4‐bromo‐2‐(4‐chlorophenyl)‐5‐(trifluoromethyl)‐1H‐pyrrole‐3‐carbonitrile (Compound 102) was 10‐fold lower than that of niclosamide. CONCLUSION: New molluscicide targets were discovered and validated, and preliminary strategies were explored for pesticide screening based on these targets. Compound 102 exhibited a high molluscicidal activity and had a great potential value for exploring a molluscicide to control P. canaliculata. © 2024 Society of Chemical Industry. [ABSTRACT FROM AUTHOR]

Oral squamous cell carcinoma (OSCC) is the most common malignant cancer of the head and neck, with high morbidity and mortality, ranking as the sixth most common cancer in the world. The treatment of OSCC is mainly radiotherapy, chemotherapy and surgery, however, the prognosis of patients is still poor and the recurrence rate is high. This paper reviews the range of effects of natural medicinal plant active ingredients (NMPAIs) on OSCC cancer, including the types of NMPAIs, anti-cancer mechanisms, involved signaling pathways, and clinical trials. The NMPAIs include terpenoids, phenols, flavonoids, glycosides, alkaloids, coumarins, and volatile oils. These active ingredients inhibit proliferation, induce apoptosis and autophagy, inhibit migration and invasion of OSCC cells, and regulate cancer immunity to exert anti-cancer effects. The mechanism involves signaling pathways such as mitogen-activated protein kinase, phosphatidylinositol 3 kinase/protein kinase B, nuclear factor kappa B, miR-22/WNT1/β-catenin and Nrf2/Keap1. Clinically, NMPAIs can inhibit the growth of OSCC, and the combined drug is more effective. Natural medicinal plants are promising candidates for the treatment of OSCC. [ABSTRACT FROM AUTHOR]

Voltage‐dependent K+ (Kv) channels play an important role in restoring the membrane potential to its resting state, thereby maintaining vascular tone. In this study, native smooth muscle cells from rabbit coronary arteries were used to investigate the inhibitory effect of quetiapine, an atypical antipsychotic agent, on Kv channels. Quetiapine showed a concentration‐dependent inhibition of Kv channels, with an IC50 of 47.98 ± 9.46 μM. Although quetiapine (50 μM) did not alter the steady‐state activation curve, it caused a negative shift in the steady‐state inactivation curve. The application of 1 and 2 Hz train steps in the presence of quetiapine significantly increased the inhibition of Kv current. Moreover, the recovery time constants from inactivation were prolonged in the presence of quetiapine, suggesting that its inhibitory action on Kv channels is use (state)‐dependent. The inhibitory effects of quetiapine were not significantly affected by pretreatment with Kv1.5, Kv2.1, and Kv7 subtype inhibitors. Based on these findings, we conclude that quetiapine inhibits Kv channels in both a concentration‐ and use (state)‐dependent manner. Given the physiological significance of Kv channels, caution is advised in the use of quetiapine as an antipsychotic due to its potential side effects on cardiovascular Kv channels. The primary goal of this study was to examine the direct effect of quetiapine on vascular Kv channels. Our results indicate that quetiapine reduces Kv currents in both concentration‐ and use (state)‐dependent manners. This inhibition is characterized by a shift in the steady‐state inactivation curve, with no effect on the activation curve. [ABSTRACT FROM AUTHOR]

Beryllium sulfate (BeSO4) can cause inflammation through the mechanism, which has not been elucidated. Mitochondrial DNA (mtDNA) is a key contributor of inflammation. With mitochondrial damage, released mtDNA can bind to specific receptors (e.g., cGAS) and then activate related pathway to promote inflammatory responses. To investigate the mechanism of mtDNA in BeSO4‐induced inflammatory response in 16HBE cells, we established the BeSO4‐induced 16HBE cell inflammation model and the ethidium bromide (EB)‐induced ρ016HBE cell model to detect the mtDNA content, oxidative stress‐related markers, mitochondrial membrane potential, the expression of the cGAS‐STING pathway, and inflammation‐related factors. Our results showed that BeSO4 caused oxidative stress, decline of mitochondrial membrane potential, and the release of mtDNA into the cytoplasm of 16HBE cells. In addition, BeSO4 induced inflammation in 16HBE cells by activating the cGAS‐STING pathway. Furthermore, mtDNA deletion inhibited the expression of cGAS‐STING pathway, IL‐10, TNF‐α, and IFN‐β. This study revealed a novel mechanism of BeSO4‐induced inflammation in 16HBE cells, which contributes to the understanding of the molecular mechanism of beryllium and its compounds‐induced toxicity. In this study, the inflammation and ρ0 cells model based on 16HBE cells was established to detect the mtDNA content, mitochondrial function‐related markers, the expression of the cGAS‐STING pathway, and inflammation. We found that BeSO4 caused oxidative stress, mitochondrial membrane potential decline, and the release of mtDNA into cytoplasm. The cGAS‐STING pathway was activated in inflamed 16HBE cells. mtDNA deletion inhibited the expression of cGAS‐STING pathway and inflammation. Our results contribute to the understanding of the molecular mechanism of BeSO4‐induced toxicity. [ABSTRACT FROM AUTHOR]

The purpose of this study is to examine thymoquinone (TQ), as a bioactive compound of Nigella sativa, from a safety point of view and to evaluate the possible toxic effects on isolated mitochondria as a screening tool for drug safety and predictive toxicology. Mitochondria were isolated form different organs (brain, heart, kidney and liver) and treated with various concentrations of TQ (0–2000 µM). Mitochondrial toxicity parameters such as succinate dehydrogenases (SDH) activity, mitochondrial swelling, lipid peroxidation, reactive oxygen species (ROS) formation and mitochondrial membrane potential (MMP) collapse were analyzed. Results showed that various concentrations of TQ did not induce any toxicity or abnormalities in isolated mitochondria obtained from brain, heart, kidney and liver. We demonstrated for the first time that TQ does not cause any detrimental effect on isolated mitochondria from different organs of rat. Our data highlight that TQ does not directly interact with mitochondria and disrupt the mitochondrial functions. [ABSTRACT FROM AUTHOR]

The processing of synaptic signals in somatodendritic compartments determines neuronal computation. Although the amplification of excitatory signals by local voltage-dependent cation channels has been extensively studied, their spatiotemporal dynamics in elaborate dendritic branches remain obscure owing to technical limitations. Using fluorescent voltage imaging throughout dendritic arborizations in hippocampal pyramidal neurons, we demonstrate a unique chloride ion (Cl-)-dependent remote computation mechanism in the distal branches. Excitatory postsynaptic potentials triggered by local laser photolysis of caged glutamate spread along dendrites, with gradual amplification toward the distal end while attenuation toward the soma. Tour de force subcellular patch-clamp recordings from thin branches complemented by biophysical model simulations revealed that the asymmetric augmentation of excitation relies on tetrodotoxin-resistant sodium ion (Na+) channels and Cl- conductance accompanied by a more hyperpolarized dendritic resting potential. Together, this study reveals the cooperative voltage-dependent actions of cation and anion conductance for dendritic supralinear computation, which can locally decode the spatiotemporal context of synaptic inputs. [ABSTRACT FROM AUTHOR]

Key points Primary motor cortex (M1) network stability depends on activity of inhibitory interneurons, for which susceptibility to stress was previously demonstrated in limbic regions. Hyperexcitability in M1 following changes in the excitatory/inhibitory balance is a key pathological hallmark of amyotrophic lateral sclerosis (ALS). Using electrophysiological approaches, we assessed the impact of acute restraint stress on inhibitory interneurons excitability and global synaptic plasticity in M1 of the SOD1G93A ALS mouse model at a late pre‐symptomatic stage (10–12.5 weeks). Based on their firing type (continuous, discontinuous, with accommodation or not) and electrophysiological characteristics (resting potential, rheobase, firing frequency), interneurons from M1 slices were separated into four clusters, labelled from 1 to 4. Among them, only interneurons from the first cluster, presenting continuous firing with few accommodations, tended to show increased excitability in wild‐type (WT) and decreased excitability in SOD1G93A animals following stress. In vivo analyses of evoked field potentials showed that stress suppressed the theta burst‐induced plasticity of an excitatory component (N1) recorded in the superficial layers of M1 in WT, with no impact on an inhibitory complex (N2–P1) from the deeper layers. In SOD1G93A mice, stress did not affect N1 but suppressed the N2–P1 plasticity. These data suggest that stress can alter M1 network functioning in a different manner in WT and SOD1G93A mice, possibly through changes of inhibitory interneurons excitability and synaptic plasticity. This suggests that stress‐induced activity changes in M1 may therefore influence ALS outcomes. Disruption of the excitatory/inhibitory balance in the primary motor cortex (M1) has been linked to cortical hyperexcitability development, a key pathological hallmark of amyotrophic lateral sclerosis (ALS). Psychological stress was reported to influence excitatory/inhibitory balance in limbic regions, but very little is known about its influence on the M1 functioning under physiological or pathological conditions. Our study revealed that acute stress influences the excitatory/inhibitory balance within the M1, through changes in interneurons excitability along with network plasticity. Such changes were different in pathological (SOD1G93A ALS mouse model) vs. physiological (wild‐type) conditions. The results of our study help us to better understand how stress modulates the M1 and highlight the need to further characterize stress‐induced motor cortex changes because it may be of importance when evaluating ALS outcomes. [ABSTRACT FROM AUTHOR]

Key points Heavy metals disrupt mitochondrial function and activate the NOD‐like receptor pyrin‐containing 3 (NLRP3) inflammasome. We investigated the effect of lead (Pb)/cadmium (Cd) on mitochondrial function and NLRP3 inflammasome activation in human trophoblast under normoxic, hypoxic and pro‐inflammatory conditions. JEG‐3, BeWo and HTR‐8/SVneo cells were exposed to Pb or Cd for 24 h in the absence or presence of hypoxia or pro‐inflammatory lipopolysaccharide (LPS) or poly(I:C). Then, we evaluated cell viability, apoptosis, mitochondrial DNA copy number (mtDNAcn), mitochondrial membrane potential (ΔΨ), NLRP3 inflammasome proteins and interleukin (IL)‐1β secretion. Although our data showed that Pb, Cd, hypoxia, poly(I:C) and LPS decreased mtDNAcn in the three cell lines, the effects of these treatments on other biomarkers were different in the different cell lines. We found that hypoxia decreased ΔΨ and promoted apoptosis in JEG‐3 cells, increased ΔΨ and prevented apoptosis in BeWo cells, and did not change ΔΨ and apoptosis in HTR‐8/SVneo cells. Moreover, Pb under hypoxic conditions reduced ΔΨ and promoted apoptosis of BeWo cells. Exposure of BeWo and HTR‐8/SVneo cells to hypoxia, Pb or Cd alone upregulated the expression of NLRP3 and pro‐caspase 1 but did not activate the NLRP3 inflammasome since cleaved‐caspase 1 and IL‐1β were not increased. To conclude, Pb and Cd affected trophoblast mitochondrial function and NLRP3 proteins in trophoblast cell lines, but in a cell line‐specific way. The objective of this work was an understanding of the effect of lead (Pb) and cadmium (Cd) on mitochondrial function and NLRP3 inflammasome activation in human trophoblast cell lines under normoxic, hypoxic and pro‐inflammatory conditions. Apoptosis of JEG‐3 cells was increased by hypoxia, while in BeWo cells, apoptosis was decreased by hypoxia, and in HTR‐8/SVneo, apoptosis was not affected by hypoxic treatment. Exposure to either Pb or Cd decreased mtDNAcn in three human placental trophoblast cell lines. However, Pb under hypoxia induced a decrease of ΔΨ and promoted apoptosis of BeWo cells, but Cd did not induce a reduction in ΔΨ in the three trophoblast cell lines under any conditions. Exposure to hypoxia, Pb or Cd increased NLRP3 and pro‐caspase 1 in BeWo and HTR‐8/SVneo cells. Our findings highlight that Pb and Cd affected trophoblast mitochondrial function and NLRP3 proteins in trophoblast cell lines but in a cell line‐specific way. [ABSTRACT FROM AUTHOR]

Efforts in researching the efficient anti-tumor properties of three novel arene ruthenium(II) complexes incorporating thiophene-based aroylhydrazone ligands have been undertaken. The complexes' elemental composition was [(η6-p-cymene)Ru(L)Cl]. They were comprehensively characterized through elemental and spectroscopic analyses (FT-IR, UV-vis, NMR, and HR-MS). Single crystal X-ray diffraction studies revealed a pseudo-octahedral geometry with bidentate coordination of the ligands in a representative complex. The in vitro assessment of the complexes' cancer cell growth inhibition was conducted using the MTT assay against A549 (human lung carcinoma), HeLa (human cervical carcinoma), HuH-7 (hepatocellular carcinoma), and NIH-3T3 (mouse fibroblast non-cancerous cell line). Results indicated significant cytotoxicity across all cancer cell lines, with IC50 concentrations of complex 2 being 6.8 μM for A549, 11.6 μM for HeLa, and 9.4 μM for HuH-7, compared to cisplatin with IC50 values of 18.9 μM, 17.68 μM, and 24 μM respectively. Notably, complex 2 demonstrated particularly promising cytotoxicity against all tested cancerous cell lines. Fluorescent staining analysis such as acridine orange/ethidium bromide (AO–EB) and HOECHST 33342 revealed cell death mechanisms involving membrane disintegration and nuclear condensation following treatment with complex 2. Further studies were conducted to measure reactive oxygen species (ROS) levels using the dichlorodihydrofluorescein diacetate (DCFH-DA) assay, and mitochondrial membrane potential (MMP) was assessed using the JC-1 dye assay. These studies demonstrated that complex 2 increased ROS levels, decreased membrane potential, and promoted mitochondrial dysfunction-mediated cell death pathways. Additionally, flow cytometry analysis, utilizing dual staining of Annexin V-FITC and propidium iodide (PI), was employed to quantitatively study apoptosis induction. [ABSTRACT FROM AUTHOR]

SLC4A11 is the most abundant membrane transport protein in corneal endothelial cells. Its functional presence is necessary to support the endothelial fluid pump that draws fluid from the corneal stroma, preventing corneal edema. Several molecular actions have been proposed for SLC4A11 including H2O transport and cell adhesion. One of the most reproduced actions that SLC4A11 mediates is a H+ (or OH-) conductance that is enhanced in the presence of NH4Cl. The mechanism by which this occurs is controversial with some providing evidence in favor of NH3-H+ cotransport and others providing evidence for uncoupled H+ transport that is indirectly stimulated by the effects of NH4Cl upon intracellular pH and membrane potential. In the present study we provide new evidence and revisit previous studies, to support a model in which NH4Cl causes direct allosteric activation of SLC4A11 by means of an acidic shift in the intracellular pK (pKi) that governs the relationship between intracellular pH (pHi) and SLC4A11 H+-conductance. These findings have important implications for the assignment of a physiological role for SLC4A11. [ABSTRACT FROM AUTHOR]

Voltage-sensing phosphatase (VSP) exhibits voltage-dependent phosphatase activity toward phosphoinositides. VSP generates a specialized phosphoinositide environment in mammalian sperm flagellum. However, the voltage-sensing mechanism of VSP in spermatozoa is not yet characterized. Here, we found that VSP is activated during sperm maturation, indicating that electric signals in immature spermatozoa are essential. Using a heterologous expression system, we show the voltage-sensing property of mouse VSP (mVSP). The voltage-sensing threshold of mVSP is approximately −30 mV, which is sensitive enough to activate mVSP in immature spermatozoa. We also report several knock-in mice in which we manipulate the voltage-sensitivity or electrochemical coupling of mVSP. Notably, the V312R mutant, with a minor voltage-sensitivity change, exhibits abnormal sperm motility after, but not before, capacitation. Additionally, the V312R mutant shows a significant change in the acyl-chain profile of phosphoinositide. Our findings suggest that electrical signals during sperm maturation are crucial for establishing the optimal phosphoinositide environment in spermatozoa. Membrane potentials in living cells can function as electrical signals. Here, the authors report that spermatozoa utilize their electrical signals during the maturing stage to create appropriate membrane lipids environment that is important for sperm motility. [ABSTRACT FROM AUTHOR]

Cisplatin-induced nephrotoxicity restricts its clinical use against solid tumors. The present study elucidated the pharmacological effects of Renogrit, a plant-derived prescription medicine, using cisplatin-induced human renal proximal tubular (HK-2) cells and Caenorhabditis elegans. Quantification of phytochemicals in Renogrit was performed on HPTLC and UHPLC platforms. Renogrit was assessed in vitro in HK-2 cells post-exposure to clinically relevant concentration of cisplatin. It was observed that renoprotective properties of Renogrit against cisplatin-induced injury stem from its ability to regulate renal injury markers (KIM-1, NAG levels; NGAL mRNA expression), redox imbalance (ROS generation; GST levels), and mitochondrial dysfunction (mitochondrial membrane potential; SKN-1, HSP-60 expression). Renogrit was also found to modulate apoptosis (EGL-1 mRNA expression; protein levels of p-ERK, p-JNK, p-p38, c-PARP1), necroptosis (intracellular calcium accumulation; RIPK1, RIPK3, MLKL mRNA expression), mitophagy (lysosome population; mRNA expression of PINK1, PDR1; protein levels of p-PINK1, LC3B), and inflammation (IL-1β activity; protein levels of LXR-α). More importantly, Renogrit treatment did not hamper normal anti-proliferative effects of cisplatin as observed from cytotoxicity analysis on MCF-7, A549, SiHa, and T24 human cancer cells. Taken together, Renogrit could be a potential clinical candidate to mitigate cisplatin-induced nephrotoxicity without compromising the anti-neoplastic properties of cisplatin. [ABSTRACT FROM AUTHOR]

Background: Leigh syndrome (LS) is a common mitochondrial disease caused by mutations in both mitochondrial and nuclear genes. Isoleucyl-tRNA synthetase 2 (IARS2) encodes mitochondrial isoleucine-tRNA synthetase, and variants in IARS2 have been reported to cause LS. However, the pathogenic mechanism of IARS2 variants is still unclear. Methods: Two unrelated patients, a 4-year-old boy and a 5-year-old boy diagnosed with LS, were recruited, and detailed clinical data were collected. The DNA of the patients and their parents was isolated from the peripheral blood for the identification of pathogenic variants using next-generation sequencing and Sanger sequencing. The ClustalW program, allele frequency analysis databases (gnomAD and ExAc), and pathogenicity prediction databases (Clinvar, Mutation Taster and PolyPhen2) were used to predict the conservation and pathogenicity of the variants. The gene expression level, oxygen consumption rate (OCR), respiratory chain complex activity, cellular adenosine triphosphate (ATP) production, mitochondrial membrane potential (MMP) and mitochondrial reactive oxygen species (ROS) levels were measured in patient-derived lymphocytes and IARS2-knockdown HEK293T cells to evaluate the pathogenicity of the variants. Results: We reported 2 unrelated Chinese patients manifested with LS who carried biallelic IARS2 variants (c.1_390del and c.2450G > A from a 4-year-old boy, and c.2090G > A and c.2122G > A from a 5-year-old boy), of which c.1_390del and c.2090G > A were novel. Functional studies revealed that the patient-derived lymphocytes carrying c.1_390del and c.2450G > A variants exhibited impaired mitochondrial function due to severe mitochondrial complexes I and III deficiencies, which was also found in IARS2-knockdown HEK293T cells. The compensatory experiments in vitro cell models confirmed the pathogenicity of IARS2 variants since re-expression of wild-type IARS2 rather than mutant IARS2 could rescue complexes I and III deficiency, oxygen consumption, and cellular ATP content in IARS2 knockdown cells. Conclusion: Our results not only expand the gene mutation spectrum of LS, but also reveal for the first time the pathogenic mechanism of IARS2 variants due to a combined deficiency of mitochondrial complexes I and III, which is helpful for the clinical diagnosis of IARS2 mutation-related diseases. [ABSTRACT FROM AUTHOR]

Key points Mammalian olfactory sensory neurons (OSNs) generate an odorant‐induced response by sequentially activating two ion channels, which are in their ciliary membranes. First, a cationic, Ca2+‐permeable cyclic nucleotide‐gated channel is opened following odorant stimulation via a G protein‐coupled transduction cascade and an ensuing rise in cAMP. Second, the increase in ciliary Ca2+ opens the excitatory Ca2+‐activated Cl− channel TMEM16B, which carries most of the odorant‐induced receptor current. While the role of TMEM16B in amplifying the response has been well established, it is less understood how this secondary ion channel contributes to response kinetics and action potential generation during single as well as repeated stimulation and, on the other hand, which response properties the cyclic nucleotide‐gated (CNG) channel determines. We first demonstrate that basic membrane properties such as input resistance, resting potential and voltage‐gated currents remained unchanged in OSNs that lack TMEM16B. The CNG channel predominantly determines the response delay and adaptation during odorant exposure, while the absence of the Cl− channels shortens both the time the response requires to reach its maximum and the time to terminate after odorant stimulation. This faster response termination in Tmem16b knockout OSNs allows them, somewhat counterintuitively despite the large reduction in receptor current, to fire action potentials more reliably when stimulated repeatedly in rapid succession, a phenomenon that occurs both in isolated OSNs and in OSNs within epithelial slices. Thus, while the two olfactory ion channels act in concert to generate the overall response, each one controls specific aspects of the odorant‐induced response. Mammalian olfactory sensory neurons (OSNs) generate odorant‐induced responses by activating two ion channels sequentially in their ciliary membranes: a Na+, Ca2⁺‐permeable cyclic nucleotide‐gated (CNG) channel and the Ca2⁺‐activated Cl⁻ channel TMEM16B. The CNG channel controls response delay and adaptation during odorant exposure, while TMEM16B amplifies the response and influences the time required for the response to reach its peak and terminate. OSNs lacking TMEM16B display faster response termination, allowing them to fire action potentials more reliably during rapid repeated stimulation. The CNG and TMEM16B channels have distinct and complementary roles in shaping the kinetics and reliability of odorant‐induced responses in OSNs. [ABSTRACT FROM AUTHOR]

It is well recognized that changes in the extracellular concentration of calcium ions influence the excitability of neurons, yet what mechanism(s) mediate these effects is still a matter of debate. Using patch‐clamp recordings from rat hippocampal CA1 pyramidal neurons, we examined the contribution of G‐proteins and intracellular calcium‐dependent signaling mechanisms to changes in intrinsic excitability evoked by altering the extracellular calcium concentration from physiological (1.2 mM) to a commonly used experimental (2 mM) level. We find that the inhibitory effect on intrinsic excitability of calcium ions is mainly expressed as an increased threshold for action potential firing (with no significant effect on resting membrane potential) that is not blocked by either the G‐protein inhibitor GDPβS or the calcium chelator BAPTA. Our results therefore argue that in the concentration range studied, G‐protein coupled calcium‐sensing receptors, non‐selective cation conductances, and intracellular calcium signaling pathways are not involved in mediating the effect of extracellular calcium ions on intrinsic excitability. Analysis of the derivative of the action potential, dV/dt versus membrane potential, indicates a current shift towards more depolarized membrane potentials at the higher calcium concentration. Our results are thus consistent with a mechanism in which extracellular calcium ions act directly on the voltage‐gated sodium channels by neutralizing negative charges on the extracellular surface of these channels to modulate the threshold for action potential activation. [ABSTRACT FROM AUTHOR]

Background: While calcium is known to play a crucial role in mammalian sperm physiology, how it flows in and out of the male gamete is not completely understood. Herein, we investigated the involvement of Na+/Ca2+ exchangers (NCX) in mammalian sperm capacitation. Using the pig as an animal model, we first confirmed the presence of NCX1 and NCX2 isoforms in the sperm midpiece. Next, we partially or totally blocked Ca2+ outflux (forward transport) via NCX1/NCX2 with different concentrations of SEA0400 (2-[4-[(2,5-difluorophenyl)methoxy]phenoxy]-5-ethoxyaniline; 0, 0.5, 5 and 50 µM) and Ca2+ influx (reverse transport) with SN6 (ethyl 2-[[4-[(4-nitrophenyl)methoxy]phenyl]methyl]-1,3-thiazolidine-4-carboxylate; 0, 0.3, 3 or 30 µM). Sperm were incubated under capacitating conditions for 180 min; after 120 min, progesterone was added to induce the acrosome reaction. At 0, 60, 120, 130, and 180 min, sperm motility, membrane lipid disorder, acrosome integrity, mitochondrial membrane potential (MMP), tyrosine phosphorylation of sperm proteins, and intracellular levels of Ca2+, reactive oxygen species (ROS) and superoxides were evaluated. Results: Partial and complete blockage of Ca2+ outflux and influx via NCX induced a significant reduction of sperm motility after progesterone addition. Early alterations on sperm kinematics were also observed, the effects being more obvious in totally blocked than in partially blocked samples. Decreased sperm motility and kinematics were related to both defective tyrosine phosphorylation and mitochondrial activity, the latter being associated to diminished MMP and ROS levels. As NCX blockage did not affect the lipid disorder of plasma membrane, the impaired acrosome integrity could result from reduced tyrosine phosphorylation. Conclusions: Inhibition of outflux and influx of Ca2+ triggered similar effects, thus indicating that both forward and reverse Ca2+ transport through NCX exchangers are essential for sperm capacitation. [ABSTRACT FROM AUTHOR]

Objectives Methods Results Conclusions This study aimed to investigate the expression and biological significance of Semenogelin 1 (SEMG1), a member of the cancer‐testis antigen family, in oral squamous cell carcinoma (OSCC). Further, we explored its potential association with metabolism‐related molecules.SEMG1 expression levels in OSCC were determined through immunohistochemistry, flow cytometry, and Western blot analyses. To decipher the biological implications of SEMG1 in OSCC, the CAL27 OSCC cell line was either stably overexpressed with SEMG1 or subjected to SEMG1‐shRNA knockdown. The relationship between clinicopathological parameters and SEMG1 expression in OSCC patients was also assessed.SEMG1 was found to be overexpressed in OSCC, though its expression was not influenced by the pathological grade. The fluorescent dihydroethidium assay indicated that SEMG1 augmented reactive oxygen species production. The mitochondrial membrane potential assay suggested a significant upregulation of mitochondrial membrane potential by SEMG1. Cell cycle assessments highlighted that SEMG1 overexpression led to a notable rise in cells entering the S‐phase. Additionally, a strong correlation between SEMG1 expression and both ENO1 and PKM2 expression in OSCC was observed.The findings underscore the elevated expression of SEMG1 in OSCC and its contributory role in the tumorigenesis of OSCC patients. [ABSTRACT FROM AUTHOR]

Mitochondria are responsible for maintaining cellular energy levels, and play a major role in regulating homeostasis, which ensures physiological function from the molecular to whole animal. Sirtuin 3 (SIRT3) is the major protein deacetylase of mitochondria. SIRT3 serves as a nutrient sensor; under conditions of mild metabolic stress, SIRT3 activity is increased. Within the mitochondria, SIRT3 regulates every complex of the electron transport chain, the tricarboxylic acid (TCA) and urea cycles, as well as the mitochondria membrane potential, and other free radical scavengers. This article reviews the role of SIRT3 in regulating homeostasis, and thus physiological function. We discuss the role of SIRT3 in regulating reactive oxygen species (ROS), ATP, immunological function and mitochondria dynamics. [ABSTRACT FROM AUTHOR]

The open-channel block of N-methyl-D-aspartate receptors (NMDARs) and their calcium-dependent desensitization (CDD) represent conventional mechanisms of glutamatergic synapse regulation. In neurotrauma, neurodegeneration, and neuropathic pain the clinical benefits of cure with memantine, ketamine, Mg2+, and some tricyclic antidepressants are often attributed to NMDAR open-channel block, while possible involvement of NMDAR CDD in the therapy is not well established. Here the effects of selective high-affinity sodium-calcium exchanger (NCX) isoform 1 inhibitor, SEA0400, on NMDA-activated whole-cell currents and their block by amitriptyline, desipramine and clomipramine recorded by patchclamp technique in cortical neurons of primary culture were studied. We demonstrated that in the presence of extracellular Ca2+, 50 nM SEA0400 caused a reversible decrease of the steady-state amplitude of NMDAR currents, whereas loading neurons with BAPTA or the removal of extracellular Ca2+ abolished the effect. The decrease did not exceed 30% of the amplitude and did not depend on membrane voltage. The external Mg2+ block and 50 nM SEA0400 inhibition of currents were additive, suggesting their independent modes of action. In the presence of Ca2+ SEA0400 speeded up the decay of NMDAR currents to the steady state determined by CDD. The measured IC50 value of 27 nM for SEA0400-induced inhibition coincides with that for NCX1. Presumably, SEA0400 effects are induced by an enhancement of NMDAR CDD through the inhibition of Ca2+ extrusion by NCX1. SEA0400, in addition, at nanomolar concentrations could interfere with Ca2+-dependent effect of tricyclic antidepressants. In the presence of 50 nM SEA0400, the IC50s for NMDAR inhibition by amitriptyline and desipramine increased by about 20 folds, as the Ca2+-dependent NMDAR inhibition disappeared. This observation highlights NCX1 involvement in amitriptyline and desipramine effects on NMDARs and unmasks competitive relationships between SEA0400 and these antidepressants. Neither amitriptyline nor desipramine could affect NCX3. The open-channel block of NMDARs by these substances was not affected by SEA0400. In agreement, SEA0400 did not change the IC50 for clomipramine, which acts as a pure NMDAR open-channel blocker. Thus, NCX seems to represent a promising molecular target to treat neurological disorders, because of the ability to modulate NMDARs by decreasing the open probability through the enhancement of their CDD. [ABSTRACT FROM AUTHOR]

Objective Methods Results Significance Pharmacological activation of neuronal Kv7 channels by the antiepileptic drug retigabine (RTG; ezogabine) has been proven effective in treating partial epilepsy. However, RTG was withdrawn from the market due to the toxicity caused by its phenazinium dimer metabolites, leading to peripheral skin discoloration and retinal abnormalities. To address the undesirable metabolic properties of RTG and prevent the formation of phenazinium dimers, we made chemical modifications to RTG, resulting in a new RTG derivative, 1025c, N,N′‐{4‐[(4‐fluorobenzyl) (prop‐2‐yn‐1‐yl)amino]‐1,2‐phenylene}bis(3,3‐dimethylbutanamide).Whole‐cell recordings were used to evaluate Kv7 channel openers. Site‐directed mutagenesis and molecular docking were adopted to investigate the molecular mechanism underlying 1025c and Kv7.2 interactions. Mouse seizure models of maximal electroshock (MES), subcutaneous pentylenetetrazol (scPTZ), and PTZ‐induced kindling were utilized to test compound antiepileptic activity.The novel compound 1025c selectively activates whole‐cell Kv7.2/7.3 currents in a concentration‐dependent manner, with half‐maximal effective concentration of .91 ± .17 μmol·L−1. The 1025c compound also causes a leftward shift in Kv7.2/7.3 current activation toward a more hyperpolarized membrane potential, with a shift of the half voltage of maximal activation (ΔV1/2) of −18.6 ± 3.0 mV. Intraperitoneal administration of 1025c demonstrates dose‐dependent antiseizure activities in assays of MES, scPTZ, and PTZ‐induced kindling models. Moreover, through site‐directed mutagenesis combined with molecular docking, a key residue Trp236 has been identified as critical for 1025c‐mediated activation of Kv7.2 channels. Photostability experiments further reveal that 1025c is more photostable than RTG and is unable to dimerize.Our findings demonstrate that 1025c exhibits potent and selective activation of neuronal Kv7 channels without being metabolized to phenazinium dimers, suggesting its developmental potential as an antiseizure agent for therapy. [ABSTRACT FROM AUTHOR]

Simple Summary: Breast cancer is one of the most frequent tumors worldwide, based on the number of new cases and deaths. Unfortunately, the low selectivity of action and the rapid development of multiple drug resistances remain the main disadvantages of anticancer compounds. The search for new agents with pronounced antitumor activity is an urgent task in modern biology and medicine. We focused on the investigation of the antitumor potential of novel hybrid pyrrolidinedione–thiazolidinone derivatives. The synthesized derivatives are effective and selective agents that exhibit their antitumor effects in breast carcinoma cells via (1) inhibiting viability, proliferation, and the ability to form colonies; (2) inducing extrinsic and intrinsic apoptotic pathways; and (3) decreasing the level of proteins associated with autophagy, invasion, and metastasis. Our results indicate that synthesized derivatives are potential candidates for deeper exploration of their therapeutic efficiency. The development of new, effective agents for the treatment of breast cancer remains a high-priority task in oncology. A strategy of treatment for this pathology depends significantly on the genotype and phenotype of human breast cancer cells. We aimed to investigate the antitumor activity of new pyrrolidinedione–thiazolidinone hybrid molecules Les-6287, Les-6294, and Les-6328 towards different types of human breast cancer cells of MDA-MB-231, MCF-7, T-47D, and HCC1954 lines and murine breast cancer 4T1 cells by using the MTT, clonogenic and [3H]-Thymidine incorporation assays, flow cytometry, ELISA, and qPCR. The studied hybrids possessed toxicity towards the mentioned tumor cells, with the IC50 ranging from 1.37 to 21.85 µM. Simultaneously, these derivatives showed low toxicity towards the pseudonormal human breast epithelial cells of the MCF-10A line (IC50 > 93.01 µM). Les-6287 at 1 µM fully inhibited the formation of colonies of the MCF-7, MDA-MB-231, and HCC1954 cells, while Les-6294 and Les-6328 did that at 2.5 and 5 µM, respectively. Les-6287 suppressed DNA biosynthesis in the MCF-7, MDA-MB-231, and HCC1954 cells. At the same time, such an effect on the MCF-10A cells was significantly lower. Les-6287 induces apoptosis using extrinsic and intrinsic pathways via a decrease in the mitochondrial membrane potential, increasing the activity of caspases 3/7, 8, 9, and 10 in all immunohistochemically different human breast cancer cells. Les-6287 decreased the concentration of the metastasis- and invasion-related proteins MMP-2, MMP-9, and ICAM-1. It did not induce autophagy in treated cells. In conclusion, the results of our study suggest that the synthesized hybrid pyrrolidinedione–thiazolidinones might be promising agents for treating breast tumors of different types. [ABSTRACT FROM AUTHOR]

Depression, a complex disorder with significant treatment challenges, necessitates innovative therapeutic approaches to address its multifaceted nature and enhance treatment outcomes. The modulation of KCNQ potassium (K+) channels, pivotal regulators of neuronal excitability and neurotransmitter release, is a promising innovative therapeutic target in psychiatry. Widely expressed across various tissues, including the nervous and cardiovascular systems, KCNQ channels play a crucial role in modulating membrane potential and regulating neuronal activity. Recent preclinical evidence suggests that KCNQ channels, particularly KCNQ3, contribute to the regulation of neuronal excitability within the reward circuitry, offering a potential target for alleviating depressive symptoms, notably anhedonia. Studies using animal models demonstrate that interventions targeting KCNQ channels can restore dopaminergic firing balance and mitigate depressive symptoms. Human studies investigating the effects of KCNQ channel activators, such as ezogabine, have shown promising results in alleviating depressive symptoms and anhedonia. The aforementioned observations underscore the therapeutic potential of KCNQ channel modulation in depression management and highlight the need and justification for phase 2 and phase 3 dose-finding studies as well as studies prespecifying symptomatic targets in depression including anhedonia. • Potential antidepressant effects via modulation of dopaminergic activity and restoration of neuronal firing balance. • Significant improvements in depressive symptoms and anhedonia with KCNQ channel activators. • KCNQ channel activators present promising avenues for antidepressant treatment but require further research. [ABSTRACT FROM AUTHOR]

The Yak (Bos grunniens) is a special breed of livestock predominantly distributed in the Qinghai–Tibet Plateau of China. Intramuscular fat (IMF) content in beef cattle is a vital indicator of meat quality. In this study, RNA-Seq and Protein-Seq were respectively employed to sequence the transcriptome and proteome of the longissimus dorsi (LD) tissue from 4-year-old yaks with significant differences in IMF content under the same fattening conditions. Five overlapping genes (MYL3, ACADS, L2HGDH, IGFN1, and ENSBGRG00000000-926) were screened using combined analysis. Functional verification tests demonstrated that the key gene ACADS inhibited yak intramuscular preadipocyte (YIMA) differentiation and proliferation, promoted mitochondrial biogenesis gene expression, and increased the mitochondrial membrane potential (MMP). Furthermore, co-transfection experiments further demonstrated that interfering with ACADS reversed the effect of PPARα agonists in promoting lipid differentiation. In conclusion, ACADS potentially inhibits lipid deposition in YIAMs by regulating the PPARα signalling pathway. These findings offer insights into the molecular mechanisms underlying yak meat quality. [ABSTRACT FROM AUTHOR]

The TRESK (K2P18.1, KCNK18) background potassium channel is expressed in primary sensory neurons and has been reported to contribute to the regulation of pain sensations. In the present study, we examined the interaction of TRESK with NDFIP1 (Nedd4 family-interacting protein 1) in the Xenopus oocyte expression system by two-electrode voltage clamp and biochemical methods. We showed that the coexpression of NDFIP1 abolished the TRESK current under the condition where the other K+ channels were not affected. Mutations in the three PPxY motifs of NDFIP1, which are responsible for the interaction with the Nedd4 ubiquitin ligase, prevented a reduction in the TRESK current. Furthermore, the overexpression of a dominant-negative Nedd4 construct in the oocytes coexpressing TRESK with NDFIP1 partially reversed the down-modulating effect of the adaptor protein on the K+ current. The biochemical data were also consistent with the functional results. An interaction between epitope-tagged versions of TRESK and NDFIP1 was verified by co-immunoprecipitation experiments. The coexpression of NDFIP1 with TRESK induced the ubiquitination of the channel protein. Altogether, the results suggest that TRESK is directly controlled by and highly sensitive to the activation of the NDFIP1-Nedd4 system. The NDFIP1-mediated reduction in the TRESK component may induce depolarization, increase excitability, and attenuate the calcium dependence of the membrane potential by reducing the calcineurin-activated fraction in the ensemble background K+ current. [ABSTRACT FROM AUTHOR]

Previous studies have suggested a role for selective serotonin reuptake inhibitors (SSRIs) such as fluoxetine (Prozac®) in the treatment of dizziness and inner ear vestibular dysfunction. The potential mechanism of action within the vestibular system remains unclear; however, fluoxetine has been reported to block certain types of K+ channel in other systems. Here, we investigated the direct actions of fluoxetine on membrane currents in presynaptic hair cells and postsynaptic calyx afferents of the gerbil peripheral vestibular system using whole cell patch clamp recordings in crista slices. We explored differences in K+ currents in peripheral zone (PZ) and central zone (CZ) calyces of the crista and their response to fluoxetine application. Outward K+ currents in PZ calyces showed greater inactivation at depolarized membrane potentials compared to CZ calyces. The application of 100 μM fluoxetine notably reduced K+ currents in calyx terminals within both zones of the crista, and the remaining currents exhibited distinct traits. In PZ cells, fluoxetine inhibited a non-inactivating K+ current and revealed a rapidly activating and inactivating K+ current, which was sensitive to blocking by 4-aminopyridine. This was in contrast to CZ calyces, where low-voltage-activated and non-inactivating K+ currents persisted following application of 100 μM fluoxetine. Additionally, marked inhibition of transient inward Na+ currents by fluoxetine was observed in calyces from both crista zones. Different concentrations of fluoxetine were tested, and the EC50 values were found to be 40 µM and 32 µM for K+ and Na+ currents, respectively. In contrast, 100 μM fluoxetine had no impact on voltage-dependent K+ currents in mechanosensory type I and type II vestibular hair cells. In summary, micromolar concentrations of fluoxetine are expected to strongly reduce both Na+ and K+ conductance in afferent neurons of the peripheral vestibular system in vivo. This would lead to inhibition of action potential firing in vestibular sensory neurons and has therapeutic implications for disorders of balance. [ABSTRACT FROM AUTHOR]

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