Regulation Of Membrane Potential Research Articles

Neuronostatin regulates neuronal function and energetic metabolism in Alzheimer's disease in a GPR107-dependent manner

Alzheimer's disease (AD) is the most prevalent neurodegenerative disease, which is characterized by the accumulation and aggregation of amyloid in brain. Neuronostatin (NST) is an endogenous peptide hormone that participates in many fundamental neuronal processes. However, the metabolism and function of NST in neurons of AD mice are not known. In this study, by combining the structural analyses, primary cultures, knockout cells, and various assessments, the behavior, histopathology, brain-wide expression and cellular signaling pathways in the APP/PS1 mice were investigated. It was found that NST directly bound to GPR107, which was primarily expressed in neurons. NST modulated the neuronal survivability and neurite outgrowth induced by Aβ via GPR107 in neurons. Intracerebroventricular (i.c.v.) administration of NST attenuated learning and memory abilities, reduced the synaptic protein levels of hippocampus, but improved amyloid plaques in the cortex and hippocampus of APP/PS1 mice. NST modulated glucose metabolism of hypothalamus-hippocampus-cortex axis in APP/PS1 mice and decreased ATP levels via the regulation of reactive oxygen species (ROS) and mitochondrial membrane potential (MMP) in response to Aβ, suppressed energetic metabolism, and mitochondrial function in neurons via GPR107/protein kinase A (PKA) signaling pathway. In summary, our findings suggest that NST regulates neuronal function and brain energetic metabolism in AD mice via the GPR107/PKA signaling pathway, which can be a promising target for the treatment of AD.

Expression and Function of Long Non-coding RNA in Endemic Cretinism.

Endemic cretinism (EC) is one of the most severe iodine deficiency disorders, leading to typical symptoms such as neurodevelopmental impairments or mental deficits. In addition to environmental factors, the pathogenesis of its genetic contribution remains unclear. The study revealed the differential expression profiles of long non-coding RNA(lncRNA) and messenger RNA(mRNA) based on high-throughput RNA-seq. GO and KEGG analyses were used to annotate the function and pathway of differentially expressed (DE) mRNA and co-expressed mRNA. The protein-protein interaction(PPI) network was established. The expression levels of three lncRNAs and six mRNAs were validated by quantitative real-time PCR analysis (qRT-PCR) and subjected to correlation analysis.Compared to controls, a total of 864 lncRNAs and 393 mRNAs were differentially expressed. The PPI network had 149 nodes and 238 edges, and three key protein-coding genes were observed.Levels of LINC01220 and target mRNA IDO1 were statistically elevated in EC patients.Differentially expressed lncRNA may be a new potential player in EC. LINC01220 and IDO1 might interact with each other to participate in EC. The biological process of regulation of postsynaptic membrane potential and the Rap1 signaling pathway might exert a regulating role in the pathophysiological process of EC. Our findings could provide more theoretical and experimental evidence for investigating the pathophysiological mechanisms of EC.

Ion Channels Remodeling in the Regulation of Vascular Hyporesponsiveness During Shock.

Shock is characterized with vascular hyporesponsiveness to vasoconstrictors, thereby to cause refractory hypotension, insufficient tissue perfusion, and multiple organ dysfunction. The vascular hyporeactivity persisted even though norepinephrine and fluid resuscitation were administrated, it is of critical importance to find new potential target. Ion channels are crucial in the regulation of cell membrane potential and affect vasoconstriction and vasodilation. It has been demonstrated that many types of ion channels including K+ channels, Ca2+ permeable channels, and Na+ channels exist in vascular smooth muscle cells and endothelial cells, contributing to the regulation of vascular homeostasis and vasomotor function. An increasing number of studies suggested that the structural and functional alterations of ion channels located in arteries contribute to vascular hyporesponsiveness during shock, but the underlying mechanisms remained to be fully clarified. Therefore, the expression and functional changes in ion channels in arteries associated with shock are reviewed, to pave the way for further exploring the potential of ion channel-targeted compounds in treating refractory hypotension in shock.

P-172 Mitochondria are dysregulated in poor quality oocytes and can be targeted in vitro to alter oocyte quality

Abstract Study question Does mitochondrial gene expression differ between good and poor quality oocytes and can mitochondria be targeted in vitro to alter oocyte quality? Summary answer Mitochondrial-related genes are dysregulated in poor quality oocytes and mitochondrial inhibitors can be used during IVM to alter oocyte quality. What is known already Good oocyte quality is essential for successful pregnancy yet we still have a poor understanding of the optimal composition of a good egg. Mitochondrial number and activity are important for oocyte quality and during maturation the ultrastructure of the mitochondria changes to a hooded or ring form. We have observed differences in the timing of this morphological change between good and poor quality oocytes in a sheep model. Oligomycin treatment of oocytes decreases the proportion of hooded mitochondria and increases mitochondrial membrane potential (MMP) indicating that hooded mitochondria may form during maturation to reduce mitochondrial activity prior to fertilisation. Study design, size, duration Gene expression differences were compared between poorer quality peripubertal lamb oocytes and adult oocytes harvested during estrus from pools of ∼80 abattoir-derived ovaries on three separate dates for each age. Denuded oocytes (GV stage) were frozen in three pools of ∼20 per age and total RNA isolated. Sheep oocytes (∼30 per group) were treated with oligomycin, FCCP or vehicle control in final 2 hours of IVM on five separate dates, fertilised and cultured. Participants/materials, setting, methods RNA sequencing cleaned reads were mapped using HISAT2 on sheep genome version ARS-UI_Ramb_v2. Count matrix were generated using featureCount and differential gene expression was performed using DESeq2. After normalisation, Wald test was used to define genes differentially expressed. Genes were considered significantly altered with a FDR < 5% after Benjamini-Hochberg correction. Mean cleavage rate was compared to vehicle control in five replicate experiments using one-way ANOVA with Benjamini, Krieger and Yekutieli FDR < 5%. Main results and the role of chance Oligomycin (ATP synthase inhibitor) decreased cleavage rate compared to control (P < 0.001) while FCCP (electron transport chain uncoupler) had no effect. There were 163 genes up-regulated and 194 genes down-regulated in lamb GV oocytes compared to adult GV stage (FDR <5%). Metascape pathway analysis of the up-regulated genes in lamb oocytes showed the most enriched pathway was Thermogenesis which included these genes: ACOT9, AJUBA, ATP5ME, ATP5MF, CALML4, CREB3L3, CRPPA, DNAJC15, DPM3, FMC1, LDHA, NDUFA12, NDUFAF2, NDUFS5, NT5C, PHGDH, POLR2G, PPARG, PRPS1L1, SDHAF3, SLIRP, TIMM8B and UBB. Six of these up-regulated genes were in the top 20 most highly expressed genes. Thermogenesis is the process by which the mitochondrial electron transport chain becomes uncoupled to dissipate heat and may also control production of free radicals by mitochondria and their response to oxidants. These genes and other mitochondrial related genes that were up-regulated in lamb GV oocytes (CKS2, DCN, PCP4L1, SLC51B) are involved in formation of the electron transport chain complexes, oxidative phosphorylation, ATP synthesis, mitochondrial organization and regulation of membrane potential. Together these results indicate that mitochondrial function is dysregulated in poorer quality oocytes and that treatment with mitochondrial inhibitors during IVM can alter oocyte quality. Limitations, reasons for caution This study was carried out in sheep oocytes and results would need to be confirmed in human oocytes if possible. Further research is needed to determine if decreasing MMP can improve oocyte quality and embryo development. Wider implications of the findings This animal model study indicates that mitochondria may be targeted during IVM to alter oocyte quality and this knowledge could be applied to develop ways of increasing human IVM and ART success rates. Trial registration number not applicable

O-197 Correlation of intracellular pH of non-normozoospermic men with fertilization rates in assisted reproduction procedures (Pilot study)

Abstract Study question Does measuring qualitative changes in the sperm pHi could predict the fertilizing potential of a non-normozoospermic patients under fertility treatment? Summary answer The increase of pHi correlates with fertilization rates. What is known already The intracellular pHi (pHi) is of great importance for a wide variety of sperm physiological processes, including its motility. The pHi regulates several key sperm proteins, such as the Ca2+ channel, CatSper (sperm cation channel) which is involved in sperm hyperactivation and the K+ channel SLO3.The latter participates in the regulation of membrane potential during capacitation. Interestingly, human sperm pHi has been found to positively correlate with both hyperactivated motility and in vitro fertilization (IVF) success in normozoospermic patients. Study design, size, duration This prospective study employed unidentified semen samples from 62 non-normozoospermic men (patients, age 38 ± 4 years old) undergoing fertility treatment from January to November 2023 at CITMER Reproductive Medicine, Mexico City. Frozen semen samples and procedures with less than 4 oocytes were excluded. The pHi of sperm samples was evaluated by time-lapse flow cytometry. For comparison, the pHi of 13 normozoospermic men (donors, age 28 + 6 years old) were also analyzed. Participants/materials, setting, methods Sperm were separated by density gradient and incubated in capacitating HTF media to be used for conventional IVF or ICSI. Cells were stained with vitality (propidium iodide) and pHi (BCECF-AM) sensitive fluorescent dyes. Fluorescence changes were evaluated using an AccuriC6 flow cytometer. For each pHi recording, 10 mM NH4Cl were used as an alkalinization positive control and for data normalization. Basal levels of sperm pHi and its correlation with fertilization rates, were performed using R. Main results and the role of chance The changes in pHi were recorded as changes in the BCECF fluorescence. The increase in fluorescence is proportional to an increase in pHi and vice versa. Interestingly, we observed that the increase in pHi in sperm used for ICSI but not for IVF procedures, which were paired with oocytes from female donors, are positively and significantly correlated with higher fertilization rates (R = 0.68; *p=0.02). This seems to be the same for ICSI treatment pairing sperm with oocyte from the female partner, although the correlation is not yet statistically significant (R = 0.44; p = 0.12). We did not find significant differences in the basal pHi values between sperm from donors and patients; however, we observed that sperm from patients, had higher pHi values (0.57 + 0.15 a.u.f) compared to sperm from donors (0.48 + 0.24 a.u.f). Our results suggest that sperm pHi may be an important regulator for the process of fusion and not only for fertilization. Qualitative sperm pHi values can contribute to predicting fertilization success in patients undergoing ICSI treatment. Limitations, reasons for caution The preliminary nature of the results implies a necessity to increase the number of analyzed semen samples. Physiological responses may vary even among the same patient or cell’s donor. Our population-based analysis could be underestimating such single cell variations. Wider implications of the findings Current protocols for sperm analysis fail to successfully predict the fertilizing capacity of semen samplest.Measuring pHi in capacitated sperm, could be a promising tool to help clinicians choose the best fertilization technique for each couple. This protocol is easy to perform in a laboratory that has a flow cytometer. Trial registration number Not applicable

Anoctamin-like protein 1 regulates repolarization in Paramecium behavioral responses.

Paramecium exhibits responsive behavior to environmental changes, moving either closer to or further away from stimuli. Electrophysiological experiments have revealed that these behavioral responses are controlled by membrane potentials. Anoctamin, a Ca2+-activated Cl- channel, is involved in the regulation of membrane potential in mammals. However, it remains uncertain whether Cl- channels like anoctamin regulate Paramecium behavior. Herein, replacement of external Cl- ions with acetate ion and application of Cl- channel blocker niflumic acid (NFA, 0.1 μM) increased spontaneous avoiding reactions (sARs). Hence, we hypothesized that anoctamin is involved in the stabilization of membrane potential fluctuation. Paramecium cells in which the anoctamin-like protein 1 gene was knocked down displayed frequent sARs in the culture medium without external stimulation. Treatment of anoctamin-like protein 1-knockdown cells with the Ca2+ chelator BAPTA or Ca-channel blocker nicardipine reversed the increase in sARs. Electrophysiological experiments revealed extension of membrane depolarization when positive currents were applied to anoctamin-like protein 1-knockdown cells. We concluded that anoctamin-like protein 1 works as a Cl-channel and stabilizes the membrane potential oscillation, reducing sARs.

C-di-AMP determines the hierarchical organization of bacterial RCK proteins

In bacteria, intracellular K+ is involved in the regulation of membrane potential, cytosolic pH, and cell turgor as well as in spore germination, environmental adaptation, cell-to-cell communication in biofilms, antibiotic sensitivity, and infectivity. The second messenger cyclic-di-AMP (c-di-AMP) has a central role in modulating the intracellular K+ concentration in many bacterial species, controlling transcription and function of K+ channels and transporters. However, our understanding of how this regulatory network responds to c-di-AMP remains poor. We used the RCK (Regulator of Conductance of K+) proteins that control the activity of Ktr channels in Bacillus subtilis as a model system to analyze the regulatory function of c-di-AMP with a combination of in vivo and in vitro functional and structural characterization. We determined that the two RCK proteins (KtrA and KtrC) are neither physiologically redundant or functionally equivalent. KtrC is the physiologically dominant RCK protein in the regulation of Ktr channel activity. In explaining this hierarchical organization, we found that, unlike KtrA, KtrC is very sensitive to c-di-AMP inactivation and lack of c-di-AMP regulation results in RCK protein toxicity, most likely due to unregulated K+ flux. We also found that KtrC can assemble with KtrA, conferring c-di-AMP regulation to the functional KtrA/KtrC heteromers and potentially compensating KtrA toxicity. Altogether, we propose that the central role of c-di-AMP in the control of the K+ machinery, by modulating protein levels through gene transcription and by regulating protein activity, has determined the evolutionary selection of KtrC as the dominant RCK protein, shaping the hierarchical organization of regulatory components of the K+ machinery.

Mechanism of noise induced hidden hearing loss based on proteomics

Objective: To explore the mechanism of noise-induced hidden hearing loss by proteomics. Methods: In October 2022, 64 SPF male C57BL/6J mice were divided into control group and noise exposure group with 32 mice in each group according to random sampling method. The noise exposure group was exposed to 100 dB sound pressure level, 2000-16000 Hz broadband noise for 2 h, and the mouse hidden hearing loss model was established. Auditory brainstem response (ABR) was used to test the change of hearing threshold of mice on the 7th day after noise exposure, the damage of basal membrane hair cells was observed by immunofluorescence, and the differentially expressed proteins in the inner ear of mice in each group were identified and analyzed by 4D-Label-free quantitative proteomics, and verified by Western blotting. The results were statistically analyzed by ANOVA and t test. Results: On the 7th day after noise exposure, there was no significant difference in hearing threshold between the control group and the noise exposure group at click and 8000 Hz acoustic stimulation (P>0.05) . The hearing threshold in the noise exposure group was significantly higher than that in the control group under 16000 Hz acoustic stimulation (P<0.05) . Confocal immunofluorescence showed that the basal membrane hair cells of cochlear tissue in noise exposure group were arranged neatly, but the relative expression of C-terminal binding protein 2 antibody of presynaptic membrane in middle gyrus and basal gyrus was significantly lower than that in control group (P<0.05) . GO enrichment analysis showed that the functions of differentially expressed proteins were mainly concentrated in membrane potential regulation, ligand-gated channel activity, and ligand-gated ion channel activity. KEGG pathway enrichment analysis showed that differentially expressed proteins were significantly enriched in phosphatidylinositol 3 kinase-protein kinase B (PI3K-Akt) signaling pathway, NOD-like receptor signaling pathway, calcium signaling pathway, etc. Western blotting showed that the expression of inositol 1, 4, 5-trisphosphate receptor 3 (Itpr3) was increased and the expression of solute carrier family 38 member 2 (Slc38a2) was decreased in the noise exposure group (P<0.05) . Conclusion: Through proteomic analysis, screening and verification of the differential expression proteins Itpr3 and Slc38a2 in the constructed mouse noise-induced hidden hearing loss model, the glutaminergic synaptic related pathways represented by Itpr3 and Slc38a2 may be involved in the occurrence of hidden hearing loss.

Identification and characterization of circular RNAs expression profiles in obstructive sleep apnea-induced liver injury.

Circular RNAs (circRNAs) have exhibited microRNA sponge activity, related to many important biological processes. Our study attempted to explore the comprehensive changes of circRNAs expression pattern in Obstructive sleep apnea (OSA)-induced liver injury and provide a global perspective of differentially expressed circRNAs (DECs). Then, RT-qPCR was used to confirm the microarray data. Further, gene ontology (GO) and KEGG pathway analysis were performed to annotate the DECs. Finally, the circRNA-miRNA-mRNA interaction network was established to predicted the target genes and target miRNAs of DECs for a stepwise bioinformatics analysis. We revealed a total of eighty DECs. In the meantime, six circRNAs were randomly validated by RT-qPCR. Among these circRNAs, mmu_circRNA_000469, 37851, 38959, 38983, 31665 were up-regulated in both microarray and qRT-PCR tissues, while mmu_circRNA_27565 was down-regulated. GO analysis revealed that circRNAs-target genes were largely related to liver function process such as carboxylic acid metabolic process and negative regulation of mitochondrial membrane potential. Meanwhile, KEGG analysis found that there were 13 pathways related to these circRNAs- target genes. And the most enriched pathway was Natural killer cell mediated cytotoxicity, which strongly suggests that immune responses may be important for the process of OSA-induced liver injury. In addition, four significant DECs (mmu_circRNA_000469, 38959, 38983, 27565) and their target mRNA and target miRNAs were further selected to establish the regulation network. Our study revealed that circRNAs may play a crucial role in OSA-induced liver injury and thus mmu_circRNA_000469, 38959, 38983, 27565 may serve as biomarkers of biological process of OSA-induced liver injury.

Network Pharmacology and Experimental Validation of Jinwei Decoction for Enhancement of Glucocorticoid Anti-Inflammatory Effect in COPD through miR-155-5p

Background: Jinwei decoction can enhance the anti-inflammatory effect of glucocorticoid (GC) on chronic obstructive pulmonary disease (COPD) by restoring the activity of human histone deacetylase-2 (HDAC2). However the upstream mechanism of Jinwei decoction on HDAC2 expression is not clear. Objective: To explore the target of Jinwei decoction to enhance the anti-inflammatory effect of GC on COPD through microRNA155-5p (miR-155-5p) by network pharmacology and experimental verification. Methods: The TCMSP database was used to screen active ingredients and target genes of Jinwei decoction, and miRWalk2.0 was used to predict downstream target genes of miR-155-5p. COPD-related genes were identified by searching GeneCards, Grugbank and OMIM databases; Venny 2.1 was used to screen intersection genes; Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways of intersection genes were analyzed by R software. Protein-protein interactions (PPIs) were analyzed by Cytoscape 3.7.2 software to identify core genes. Finally, interactions between main compounds and potential targets were verified by molecular docking. A COPD cell model was established by 5% cigarette smoke extract (CSE)- induced bronchial epithelial cell (BEAS-2B), and the results of network pharmacology were verified by in vitro experiments. Results: Two hundred thirty-one active ingredients, 352 Jinwei decoction drug targets, 5949 miR-155-5p target genes, 8286 COPD target genes, and 127 intersection genes were identified. Twelve core proteins of PPI networks may be involved. GO enrichment analysis showed that regulation of membrane potential， response to steroid hormone, and histone modification were involved; KEGG pathway enrichment analysis concentrated in the PI3K-Akt, mitogen-activated protein kinase (MAPK), HIF-1, and other signaling pathways. The molecular docking results showed that quercetin, luteolin and stigmasterol have higher affinity with PTGS2, HIF1A and AKT1. The results of cell experiments revealed that Jinwei decoction not only enhances the anti- inflammatory effect of GC in the COPD cell model but also reverses the high expression of miR-155-5p、PI3k、Akt, and low expression of HDAC2, thereby inhibiting the inflammatory response of COPD. result: Two hundred thirty-one active ingredients, 352 Jinwei decoction drug targets, 5949 miR-155-5p target genes, 8286 COPD target genes, and 127 intersection gene were identified. Twelve core proteins of PPI networks may be involved. GO enrichment analysis showed that regulation of membrane potential，response to steroid hormone, and histone modification were involved; KEGG pathway enrichment analysis concentrated in the PI3K-Akt, mitogen-activated protein kinase (MAPK), HIF-1, and other signaling pathways. The molecular docking results showed that quercetin, luteolin and stigmasterol have higher affinity with PTGS2, HIF1A and AKT1. The results of cell experiments revealed that Jinwei decoction not only enhances the anti-inflammatory effect of GC in the COPD cell model, but also reversed the high expression of miR-155-5p、PI3k、Akt, and low expression of HDAC2, thereby inhibiting the inflammatory response of COPD. Conclusion: Jinwei decoction can regulate HDAC2 activity and enhance the anti-inflammatory effect of GC on COPD by modulating miR-155-5p. Its mechanism of action may be related to its effect on the PI3K-Akt through miR-155-5p.

Challenges in the Therapeutic Targeting of KCa Channels: From Basic Physiology to Clinical Applications.

Calcium-activated potassium (KCa) channels are ubiquitously expressed throughout the body and are able to regulate membrane potential and intracellular calcium concentrations, thereby playing key roles in cellular physiology and signal transmission. Consequently, it is unsurprising that KCa channels have been implicated in various diseases, making them potential targets for pharmaceutical interventions. Over the past two decades, numerous studies have been conducted to develop KCa channel-targeting drugs, including those for disorders of the central and peripheral nervous, cardiovascular, and urinary systems and for cancer. In this review, we synthesize recent findings regarding the structure and activating mechanisms of KCa channels. We also discuss the role of KCa channel modulators in therapeutic medicine. Finally, we identify the major reasons behind the delay in bringing these modulators to the pharmaceutical market and propose new strategies to promote their application.

Identification and Validation of Synapse-related Hub Genes after Spinal Cord Injury by Bioinformatics Analysis.

Spinal cord injury (SCI) is a neurological disease with high morbidity and mortality. Previous studies have shown that abnormally expressed synapse-related genes are closely related to the occurrence and development of SCI. However, little is known about the interaction of these aberrantly expressed genes and the molecular mechanisms that play a role in the injury response. Therefore, deeply exploring the correlation between synapse-related genes and functional recovery after spinal cord injury and the molecular regulation mechanism is of great significance. First, we selected the function GSE45006 dataset to construct three clinically meaningful gene modules by hierarchical clustering analysis in 4 normal samples and 20 SCI samples. Subsequently, we performed functional and pathway enrichment analyses of key modules. The results showed that related module genes were significantly enriched in synaptic structures and functions, such as the regulation of synaptic membranes and membrane potential. A protein-protein interaction network (PPI) was constructed to identify 10 hub genes of SCI, and the results showed that Snap25, Cplx1, Stxbp1, Syt1, Rims1, Rab3a, Syn2, Syn1, Cask, Lin7b were most associated with SCI. Finally, these hub genes were further verified by quantitative real-time fluorescence polymerase chain reaction (qRT-PCR) in the spinal cord tissues of the blank group and SCI rats, and it was found that the expression of these hub genes was significantly decreased in the spinal cord injury compared with the blank group (P ≤ 0.05). These results suggest that the structure and function of synapses play an important role after spinal cord injury. Our study helps to understand the underlying pathogenesis of SCI patients further and identify new targets for SCI treatment.

A computational model predicts sex-specific responses to calcium channel blockers in mammalian mesenteric vascular smooth muscle

The function of the smooth muscle cells lining the walls of mammalian systemic arteries and arterioles is to regulate the diameter of the vessels to control blood flow and blood pressure. Here, we describe an in silico model, which we call the ‘Hernandez–Hernandez model’, of electrical and Ca2+ signaling in arterial myocytes based on new experimental data indicating sex-specific differences in male and female arterial myocytes from murine resistance arteries. The model suggests the fundamental ionic mechanisms underlying membrane potential and intracellular Ca2+ signaling during the development of myogenic tone in arterial blood vessels. Although experimental data suggest that KV1.5 channel currents have similar amplitudes, kinetics, and voltage dependencies in male and female myocytes, simulations suggest that the KV1.5 current is the dominant current regulating membrane potential in male myocytes. In female cells, which have larger KV2.1 channel expression and longer time constants for activation than male myocytes, predictions from simulated female myocytes suggest that KV2.1 plays a primary role in the control of membrane potential. Over the physiological range of membrane potentials, the gating of a small number of voltage-gated K+ channels and L-type Ca2+ channels are predicted to drive sex-specific differences in intracellular Ca2+ and excitability. We also show that in an idealized computational model of a vessel, female arterial smooth muscle exhibits heightened sensitivity to commonly used Ca2+ channel blockers compared to male. In summary, we present a new model framework to investigate the potential sex-specific impact of antihypertensive drugs.

A computational model predicts sex-specific responses to calcium channel blockers in mammalian mesenteric vascular smooth muscle.

The function of the smooth muscle cells lining the walls of mammalian systemic arteries and arterioles is to regulate the diameter of the vessels to control blood flow and blood pressure. Here, we describe an in silico model, which we call the 'Hernandez-Hernandez model', of electrical and Ca2+ signaling in arterial myocytes based on new experimental data indicating sex-specific differences in male and female arterial myocytes from murine resistance arteries. The model suggests the fundamental ionic mechanisms underlying membrane potential and intracellular Ca2+ signaling during the development of myogenic tone in arterial blood vessels. Although experimental data suggest that KV1.5 channel currents have similar amplitudes, kinetics, and voltage dependencies in male and female myocytes, simulations suggest that the KV1.5 current is the dominant current regulating membrane potential in male myocytes. In female cells, which have larger KV2.1 channel expression and longer time constants for activation than male myocytes, predictions from simulated female myocytes suggest that KV2.1 plays a primary role in the control of membrane potential. Over the physiological range of membrane potentials, the gating of a small number of voltage-gated K+ channels and L-type Ca2+ channels are predicted to drive sex-specific differences in intracellular Ca2+ and excitability. We also show that in an idealized computational model of a vessel, female arterial smooth muscle exhibits heightened sensitivity to commonly used Ca2+ channel blockers compared to male. In summary, we present a new model framework to investigate the potential sex-specific impact of antihypertensive drugs.

NALCN Channels Are Not Major targets of Gα o or Gα q Modulation in the C. elegans Egg-Laying Behavior Circuit.

Sodium leak channels (NALCN) are regulators of cell membrane potential. Previous studies in mammalian neurons and C. elegans have shown that Gα q and Gα o signaling antagonistically modulates NALCN activity to regulate neuron excitability and neurotransmitter release for behavior. Here, we test whether NALCNs mediate the effects of Gα q and/or Gα o signaling in the C. elegans egg-laying circuit. We find that while gain-of-function NALCN mutants exhibit hyperactive egg-laying behavior, NALCNs are not required for the effects of Gα q or Gα o signaling for egg laying. These results show that NALCNs are not major effectors of G-protein signaling for C. elegans egg-laying behavior.

A Novel Tri-Hydroxy-Methylated Chalcone Isolated from Chromolaena tacotana with Anti-Cancer Potential Targeting Pro-Survival Proteins.

Chromolaena tacotana (Klatt) R. M. King and H. Rob (Ch. tacotana) contains bioactive flavonoids that may have antioxidant and/or anti-cancer properties. This study investigated the potential anti-cancer properties of a newly identified chalcone isolated from the inflorescences of the plant Chromolaena tacotana (Klatt) R. M. King and H. Rob (Ch. tacotana). The chalcone structure was determined using HPLC/MS (QTOF), UV, and NMR spectroscopy. The compound cytotoxicity and selectivity were evaluated on prostate, cervical, and breast cancer cell lines using the MTT assay. Apoptosis and autophagy induction were assessed through flow cytometry by detecting annexin V/7-AAD, active Casp3/7, and LC3B proteins. These results were supported by Western blot analysis. Mitochondrial effects on membrane potential, as well as levels of pro- and anti-apoptotic proteins were analyzed using flow cytometry, fluorescent microscopy, and Western blot analysis specifically on a triple-negative breast cancer (TNBC) cell line. Furthermore, molecular docking (MD) and molecular dynamics (MD) simulations were performed to evaluate the interaction between the compounds and pro-survival proteins. The compound identified as 2',3,4-trihydroxy-4',6'-dimethoxy chalcone inhibited the cancer cell line proliferation and induced apoptosis and autophagy. MDA-MB-231, a TNBC cell line, exhibited the highest sensitivity to the compound with good selectivity. This activity was associated with the regulation of mitochondrial membrane potential, activation of the pro-apoptotic proteins, and reduction of anti-apoptotic proteins, thereby triggering the intrinsic apoptotic pathway. The chalcone consistently interacted with anti-apoptotic proteins, particularly the Bcl-2 protein, throughout the simulation period. However, there was a noticeable conformational shift observed with the negative autophagy regulator mTOR protein. Future studies should focus on the molecular mechanisms underlying the anti-cancer potential of the new chalcone and other flavonoids from Ch. tacotana, particularly against predominant cancer cell types.

Gray matter volume abnormalities in obsessive-compulsive disorder correlate with molecular and transcriptional profiles

Neuroimaging studies have consistently established altered brain structure in obsessive-compulsive disorder (OCD). However, the molecular and genetic mechanisms underlying structural brain abnormalities remain unclear. In this study, we aimed to investigate altered gray matter volume and its underlying molecular and genetic mechanisms in patients with OCD. Gray matter morphological abnormalities measured with voxel based morphometry analysis were identified in patients with OCD in comparison to sex- and age-matched healthy controls (HCs). Spatial correlations between gray matter morphological abnormalities and neurotransmitter maps were calculated to identify neurotransmitters relating to structural abnormalities. Structural abnormalities related genes were identified by conducting transcriptome-neuroimaging spatial correlations. Compared with HCs, patients with OCD demonstrated significant morphological abnormalities in distributed brain areas, including gray matter atrophy in the anterior cingulate and increased gray matter volume in the thalamus, caudate and precentral and postcentral gyrus. The morphological abnormalities were significantly associated with dopamine synthesis capacity and expression profiles of 1110 genes enriched for trans-synaptic signaling, regulation of membrane potential, modulation of chemical synaptic transmission, brain development, synapse organization and regulation of neurotransmitter levels. These results elucidate the molecular and transcriptional basis of altered gray matter morphology and build linking between molecular, transcriptional and neuroimaging information facilitating an integrative understanding of OCD.

Physiological functions and pathological involvement of ion channel trafficking in the vasculature.

A variety of ion channels regulate membrane potential and calcium influx in arterial smooth muscle and endothelial cells to modify vascular functions, including contractility. The current (I) generated by a population of ion channels is equally dependent upon their number (N), open probability (Po) and single channel current (i), such that I=N.PO .i. A conventional view had been that ion channels traffic to the plasma membrane in a passive manner, resulting in a static surface population. It was also considered that channels assemble with auxiliary subunits prior to anterograde trafficking of the multimeric complex to the plasma membrane. Recent studies have demonstrated that physiological stimuli can regulate the surface abundance (N) of several different ion channels in arterial smooth muscle and endothelial cells to control arterial contractility. Physiological stimuli can also regulate the number of auxiliary subunits present in the plasma membrane to modify the biophysical properties, regulatory mechanisms and physiological functions of some ion channels. Furthermore, ion channel trafficking becomes dysfunctional in the vasculature during hypertension, which negatively impacts the regulation of contractility. The temporal kinetics of ion channel and auxiliary subunit trafficking can also vary depending on the signalling mechanisms and proteins involved. This review will summarize recent work that has uncovered the mechanisms, functions and pathological modifications of ion channel trafficking in arterial smooth muscle and endothelial cells.

Inhibition of voltage-dependent K+ channels in rabbit coronary arterial smooth muscle cells by the atypical antipsychotic agent sertindole.

The regulation of membrane potential and the contractility of vascular smooth muscle cells (VSMCs) by voltage-dependent K+ (Kv) potassium channels are well-established. In this study, native VSMCs from rabbit coronary arteries were used to investigate the inhibitory effect of sertindole, an atypical antipsychotic agent, on Kv channels. Sertindole induced dose-dependent inhibition of Kv channels, with an IC50 of 3.13 ± 0.72 μM. Although sertindole did not cause a change in the steady-state activation curve, it did lead to a negative shift in the steady-state inactivation curve. The application of 1- or 2-Hz train pulses failed to alter the sertindole-induced inhibition of Kv channels, suggesting use-independent effects of the drug. The inhibitory response to sertindole was significantly diminished by pretreatment with a Kv1.5 inhibitor but not by Kv2.1 and Kv7 subtype inhibitors. These findings demonstrate the sertindole dose-dependent and use-independent inhibition of vascular Kv channels (mainly the Kv1.5 subtype) through a mechanism that involves altering steady-state inactivation curves. Therefore, the use of sertindole as an antipsychotic drug may have adverse effects on the cardiovascular system.

DMT1 differentially regulates mitochondrial complex activities to reduce glutathione loss and mitigate ferroptosis.

Mitochondria are vital for energy production and redox homeostasis, yet knowledge of relevant mechanisms remains limited. Here, through a genome-wide CRISPR-Cas9 knockout screening, we have identified DMT1 as a major regulator of mitochondria membrane potential. Our findings demonstrate that DMT1 deficiency increases the activity of mitochondrial complex I and reduces that of complex III. Enhanced complex I activity leads to increased NAD+ production, which activates IDH2 by promoting its deacetylation via SIRT3. This results in higher levels of NADPH and GSH, which improve antioxidant capacity during Erastin-induced ferroptosis. Meanwhile, loss of complex III activity impairs mitochondrial biogenesis and promotes mitophagy, contributing to suppression of ferroptosis. Thus, DMT1 differentially regulates activities of mitochondrial complex I and III to cooperatly suppress Erastin-induced ferroptosis. Furthermore, NMN, an alternative method of increasing mitochondrial NAD+, exhibits similar protective effects against ferroptosis by boosting GSH in a manner similar to DMT1 deficiency, shedding a light on potential therapeutic strategy for ferroptosis-related pathologies.