Decreased Calcium Influx Research Articles

Ursolic acid attenuates pseudo-allergic reactions via reducing MRGPRX2-mediated mast cell degranulation

Mas-related G protein–coupled receptor X2 (MRGPRX2) is a newly identified receptor on mast cells that contribute to IgE-independent pseudo-allergy. Ursolic acid (UA), a pentacyclic triterpenoid, has been reported for its anti-allergy effects. However, the protective mechanism against pseudo-allergic reactions remains unclear. This study aims to investigate the effects of UA on pseudo-allergic reactions both in vivo and in vitro, focusing on the therapeutical mechanism underlying its effect on mast cells. In present study, UA reduced degranulation and chemokines production induced by MRGPRX2 agonists, including compound 48/80 (C48/80) and substance P (SP), in LAD2 cells in vitro. UA also alleviated C48/80 and SP-induced systemic anaphylaxis and passive cutaneous anaphylaxis (PCA) in vivo. Furthermore, UA demonstrated strong binding affinity to the MRGPRX2 protein, leading to a decrease in calcium influx in both LAD2 cells and MRGPRX2-HEK293 cells stimulated with C48/80 and SP. Moreover, UA effectively suppressed phosphorylation levels within phospholipase C-γ (PLCγ) pathway and nuclear factor kappa-B (NF-κB) pathway of MRGPRX2 downstream proteins. Our findings indicated that UA exerts an attenuating effect in pseudo-allergic reactions by suppressing MRGPRX2-mediated mast cell activation, targeting PLCγ pathway and NF-κB pathway. These results suggest that UA may serve as a promising therapeutic agent for MRGPRX2-dependent pseudo-allergic reactions.

Astaxanthin inhibits apoptosis in a cell model of tauopathy by attenuating endoplasmic reticulum stress and unfolded protein response

The accumulation of misfolded proteins is a common pathological characteristic shared by many neurodegenerative diseases including Alzheimer’s disease and Parkinson’s disease. The disruption of proteostasis triggers endoplasmic reticulum (ER) stress, during which the unfolded protein response (UPR) is initiated by the activation of protein kinase R-like ER kinase (PERK), inositol-requiring enzyme 1 (IRE1) and activating transcription factor 6 (ATF6). These three branches of UPR signals act in concert to reduce the levels of abnormal proteins and restore ER homeostasis. However, the overactivation of UPR impairs cell function and induces apoptosis, which has been implicated in neurodegeneration. Astaxanthin is a xanthophyll carotenoid which has been shown to have neuroprotective effects in both cell and animal models; however, its effects on ER stress and UPR induced by disrupted proteostasis remain unclear. In this study, the effects of astaxanthin on ER stress and cytotoxicity were investigated in N2a cells stably expressing the pro-aggregant tau repeat domain carrying FTDP-17 mutation ΔK280 (Tau4RDΔK280). The results demonstrated that astaxanthin significantly inhibited Tau4RDΔK280-induced loss of cell viability and apoptosis, attenuating Tau4RDΔK280-induced caspase-3 activation and decrease of Bcl-2. Further studies revealed that astaxanthin treatment alleviated Tau4RDΔK280-induced ER stress and suppressed the activation of PERK, IRE1 and ATF6 signaling pathways. These findings suggested that astaxanthin might inhibit Tau4RDΔK280-induced cytotoxicity by attenuating UPR and ER stress. In addition, astaxanthin treatment resulted in a great reduction in the production of intracellular reactive oxygen species and a significant decrease in calcium influx induced by Tau4RDΔK280, which also contributed to the protective effects of astaxanthin against Tau4RDΔK280-induced cytotoxicity.

ITPRIPL1 binds CD3ε to impede T cell activation and enable tumor immune evasion

Cancer immunotherapy has transformed treatment possibilities, but its effectiveness differs significantly among patients, indicating the presence of alternative pathways for immune evasion. Here, we show that ITPRIPL1 functions as an inhibitory ligand of CD3ε, and its expression inhibits Tcells in the tumor microenvironment. The binding of ITPRIPL1 extracellular domain to CD3ε on Tcells significantly decreased calcium influx and ZAP70 phosphorylation, impeding initial Tcell activation. Treatment with a neutralizing antibody against ITPRIPL1 restrained tumor growth and promoted Tcell infiltration in mouse models across various solid tumor types. The antibody targeting canine ITPRIPL1 exhibited notable therapeutic efficacy against naturally occurring tumors in pet clinics. These findings highlight the role of ITPRIPL1 (or CD3L1, CD3ε ligand 1) in impeding Tcell activation during the critical "signal one" phase. This discovery positions ITPRIPL1 as a promising therapeutic target against multiple tumor types.

An inactivating human TRPC6 channel mutation without focal segmental glomerulosclerosis

Transient receptor potential cation channel-6 (TRPC6) gene mutations cause familial focal segmental glomerulosclerosis (FSGS), which is inherited as an autosomal dominant disease. In patients with TRPC6-related FSGS, all mutations map to the N- or C-terminal TRPC6 protein domains. Thus far, the majority of TRPC6 mutations are missense resulting in increased or decreased calcium influx; however, the fundamental molecular mechanisms causing cell injury and kidney pathology are unclear. We report a novel heterozygous TRPC6 mutation (V691Kfs*) in a large kindred with no signs of FSGS despite a largely truncated TRPC6 protein. We studied the molecular effects of V691Kfs* TRPC6 mutant using the tridimensional cryo-EM structure of the tetrameric TRPC6 protein. The results indicated that V691 is localized at the pore-forming transmembrane region affecting the ion conduction pathway, and predicted that V691Kfs* causes closure of the ion-conducting pathway leading to channel inactivation. We assessed the impact of V691Kfs* and two previously reported TRPC6 disease mutants (P112Q and G757D) on calcium influx in cells. Our data show that the V691Kfs* fully inactivated the TRCP6 channel-specific calcium influx consistent with a complete loss-of-function phenotype. Furthermore, the V691Kfs* truncation exerted a dominant negative effect on the full-length TRPC6 proteins. In conclusion, the V691Kfs* non-functional truncated TRPC6 is not sufficient to cause FSGS. Our data corroborate recently characterized TRPC6 loss-of-function and gain-of-function mutants suggesting that one defective TRPC6 gene copy is not sufficient to cause FSGS. We underscore the importance of increased rather than reduced calcium influx through TRPC6 for podocyte cell death.

Alpha-Mangostin: A Potent Inhibitor of TRPV3 and Pro-Inflammatory Cytokine Secretion in Keratinocytes.

The TRPV3 calcium ion channel is vital for maintaining skin health and has been associated with various skin-related disorders. Since TRPV3 is involved in the development of skin inflammation, inhibiting TRPV3 could be a potential treatment strategy. Alpha-mangostin isolated from Garcinia mangostana L. extract exhibits diverse positive effects on skin health; however, the underlying mechanisms remain obscure. This study investigated the TRPV3-inhibitory properties of alpha-mangostin on TRPV3 hyperactive mutants associated with Olmsted syndrome and its impact on TRPV3-induced cytokine secretion and cell death. Our findings demonstrate that alpha-mangostin effectively inhibits TRPV3, with an IC50 of 0.077 ± 0.013 μM, showing inhibitory effects on both wild-type and mutant TRPV3. TRPV3 inhibition with alpha-mangostin decreased calcium influx and cytokine release, protecting cells from TRPV3-induced death. These results indicate that alpha-mangostin reduced inflammation in TRPV3-activated skin keratinocytes, suggesting that alpha-mangostin could be potentially used for improving inflammatory skin conditions such as dermatitis.

Sophorolipid inhibits histamine-induced itch by decreasing PLC/IP3R signaling pathway activation and modulating TRPV1 activity

Biosurfactants are attracting much interest due to their potential application as therapeutic agents in the medical and cosmetic field. Previous studies have demonstrated that biosurfactant such as sophorolipid (SL) exhibits immunomodulatory effects. In this article, we found the potential of sophorolipid for inhibiting histamine-induced itch and preliminarily explored its molecular basis. First, behavioral tests indicated that SL can remit the histamine-induced scratching behaviors of mice. Second, SL can suppress the the calcium influx induced by histamine, HTMT and VUF8430 in HaCaT cells. RT-PCR analysis showed that the histamine-induced upregulation of mRNA levels of phospholipase Cγ1, 1,4,5-trisphosphate receptor (IP3R), and protein kinase Cα can be inhibted by SL, suggesting that SL may impede the PLC/IP3R signaling pathway activated by histamine. In further tests, the capsaicin-induced calcium influx can also be inhibited by SL. The immunofluorescence and molecular docking analysis indicated that SL acts as an inhibitor of transient receptor potential vanilloid-1 (TRPV1) activation to decrease calcium influx against stimuli. In summary, these results revealed that SL may inhibit histamine-induced itch by decreasing PLC/IP3R signaling pathway activation and modulating TRPV1 activity. This paper indicates that SL can be a useful treatment for histamine-dependent itch.

UCP2 Deficiency Renders Th1/2 Pathogenic Phenotype to CD4+ T cells

Abstract Uncoupling protein 2 (UCP2) is an anion transporter in the mitochondrial matrix that controls ROS production, calcium influx, and C4 metabolites. It is regarded as a metabolic regulator. A lack of UCP2 has led to an extreme clinical score with higher oxidative stress and inflammation in autoimmune illness, even though elevated UCP2 has been found in many tumors and is known to be a potential target for tumor therapy. Uncertainty still exists regarding UCP2’s function in T cells. A significant rise in the Th2/Th1 ratio was seen in this work using CD4 cells from UCP2−/− mice. These cells exhibited an improved Th2 phenotype with decreased calcium influx, elevated iNOS expression, and elevated urea metabolism. CD4 cells were treated with NOC-18, a source of nitric oxide, or Genipin to demonstrate nitric oxide-mediated Th2 polarization (inhibitor for UCP2). There were more cytotoxic markers and Th2/Th1 double positive cells. The B16/F10 tumor cells were controlled in vitro in a tumor killing assay utilizing genipin-treated CD4 cells, according to the findings. However, in UCP2−/− mice, the severity of EAE worsened, and heterogenic Th2 cells were infiltrated into central nerves system. Our findings imply that the loss of UCP2 in CD4 increases Th2 polarization with a cytotoxic phenotype and may be relevant to various illnesses. R01 CA250458, R01 CA236379

STUDY OF THE EFFECTS OF VINPOCETINE ON INDICATORS OF ACID-ALKALINE STATE FOLLOWING THE ISCHEMIC BRAIN DAMAGE

Vinpocetine is a multipurpose medicine with several pharmacological targets. It is known for its neuroprotective properties. Its mechanism of action in brain pathology is due to a selective inhibitor of phosphodiesterase type 1, inhibition of voltage-gated sodium channels, and a decrease in calcium influx into neurons. Vinpocetine combines high clinical efficacy and safety of use. Therefore it successfully competes in the pharmaceutical market with an ever-increasing number of new medicines. The purpose of the work was to experimentally study the effects of vinpocetine in cats at a dose of 5 mg/kg on the indicators of the acid-base state (ABS) of arterial and venous blood in the simulation of total cerebral ischemia. The effect of vinpocetine on acid-base status indicators should be considered as the most important aspect of its action, contributing to the neuroprotective effect. The paper discusses possible molecular targets and mechanisms for implementing the protective effect of vinpocetine in ischemia, considering its favorable effect on ABS. These new findings may point to increased use of vinpocetine in human cardiovascular diseases accompanied by ABS deterioration.

Broader functions of TIR domains in Arabidopsis immunity

TIR domains are NAD-degrading enzymes that function during immune signaling in prokaryotes, plants, and animals. In plants, most TIR domains are incorporated into intracellular immune receptors termed TNLs. In Arabidopsis, TIR-derived small molecules bind and activate EDS1 heterodimers, which in turn activate RNLs, a class of cation channel-forming immune receptors. RNL activation drives cytoplasmic Ca2+ influx, transcriptional reprogramming, pathogen resistance, and host cell death. We screened for mutants that suppress an RNL activation mimic allele and identified a TNL, SADR1. Despite being required for the function of an autoactivated RNL, SADR1 is not required for defense signaling triggered by other tested TNLs. SADR1 is required for defense signaling initiated by some transmembrane pattern recognition receptors and contributes to the unbridled spread of cell death in lesion simulating disease 1. Together with RNLs, SADR1 regulates defense gene expression at infection site borders, likely in a non-cell autonomous manner. RNL mutants that cannot sustain this pattern of gene expression are unable to prevent disease spread beyond localized infection sites, suggesting that this pattern corresponds to a pathogen containment mechanism. SADR1 potentiates RNL-driven immune signaling not only through the activation of EDS1 but also partially independently of EDS1. We studied EDS1-independent TIR function using nicotinamide, an NADase inhibitor. Nicotinamide decreased defense induction from transmembrane pattern recognition receptors and decreased calcium influx, pathogen growth restriction, and host cell death following intracellular immune receptor activation. We demonstrate that TIR domains can potentiate calcium influx and defense and are thus broadly required for Arabidopsis immunity.

Detrimental effects of simulated microgravity on mast cell homeostasis and function.

Exposure to microgravity causes significant alterations in astronauts' immune systems during spaceflight; however, it is unknown whether microgravity affects mast cell homeostasis and activation. Here we show that microgravity negatively regulates the survival and effector function of mast cells. Murine bone marrow-derived mast cells (BMMCs) were cultured with IL-3 in a rotary cell culture system (RCCS) that generates a simulated microgravity (SMG) environment. BMMCs exposed to SMG showed enhanced apoptosis along with the downregulation of Bcl-2, and reduced proliferation compared to Earth's gravity (1G) controls. The reduction in survival and proliferation caused by SMG exposure was recovered by stem cell factor. In addition, SMG impaired mast cell degranulation and cytokine secretion. BMMCs pre-exposed to SMG showed decreased release of β-hexosaminidase, interleukin-6 (IL-6), and tumor necrosis factor-α (TNF-α) upon stimulation with phorbol 12-myristate-13-acetate (PMA) plus calcium ionophore ionomycin, which correlated with decreased calcium influx. These findings provide new insights into microgravity-mediated alterations of mast cell phenotypes, contributing to the understanding of immune system dysfunction for further space medicine research.

Acute, sub-chronic and chronic exposures to TiO2 and Ag nanoparticles differentially affects neuronal function in vitro

In vivo toxicokinetic studies provide evidence for the translocation and accumulation of nanoparticles (NP) in the brain, thereby causing concern for adverse health effects, particularly for effects following chronic exposure. To date, only few studies investigated the effects of NP exposure on neuronal function in vitro, primarily focusing on short-term effects. The aim of this study was therefore to investigate the effects of two common types of NP, titanium dioxide NP (TiO2NP) and silver NP (AgNP), on neuronal function following acute (0.5 h), sub-chronic (24 h and 48 h) and chronic (14 days) exposure in vitro. Effects of NP exposure on intracellular calcium homeostasis, spontaneous neuronal (network) activity and neuronal network morphology were investigated in rat primary cortical cells using respectively, single-cell microscopy calcium imaging, micro-electrode array (MEA) recordings and immunohistochemistry. Our data demonstrate that high doses of AgNP (≥ 30 µg/mL) decrease calcium influx after 24 h exposure, although neuronal activity is not affected following acute and sub-chronic exposure. However, chronic exposure to non-cytotoxic doses of AgNP (1–10 µg/mL) potently decreases spontaneous neuronal (network) activity, without affecting network morphology and viability. Exposure to higher doses (≥ 30 µg/mL) affects network morphology and is also associated with cytotoxicity. In contrast, acute and sub-chronic exposure to TiO2NP is without effects, whereas chronic exposure only modestly reduces neuronal function without affecting morphology. Our combined findings indicate that TiO2NP exposure is of limited hazard for neuronal function whereas AgNP, in particularly during chronic exposure, has profound effects on neuronal (network) function and morphology.

The Utility of Ecmo and Intraaortic Balloon Pumps in Lipophilic Beta Blocker and Bupropion Overdose

Patient: Female, 21 year old Final Diagnosis: Cardiogenic shock due to ingestion Symptoms: Encephalopathy, seizures Medication: Propranolol, Bupropion Clinical Procedure: ECMO, Intraaortic Balloon Pump Specialty: Critical Care, Toxicology Background Combined massive ingestion of Beta Adrenergic Blocking Medications (BBs), Calcium Channel Blockers (CCBs), and behavioral modulators can be dangerous due to their cardiotoxicity leading to hypotension and severe refractory cardiogenic shock. Beta blockers can cause serious myocardial depression by decreasing calcium influx into cells. Some beta blockers exhibit sodium channel blockade, while others cross the blood brain barrier due to their lipophilicity causing CNS toxicity. Behavioral modulators, specifically Bupropion, cause significant cardiac depression as well by acting as a potential cardiac gap junctions blocker and therefore decreasing intercellular communication. Case Report We present a case of combined massive drug ingestion leading to severe refractory cardiogenic shock where invasive mechanical support was implemented. Patient was a 21-year-old female who presented after polyingestion in a suicide attempt. Patient took a large amount of propranolol and bupropion. She initially presented in cardiogenic shock, and seizing. She was intubated for airway protection, started multiple vasopressors, and high - dose insulin. Despite multiple therapies, she continued to deteriorate and her ejection fraction continued to worsen, at which point she was cannulated for Extracorporeal Membrane Oxygenation (ECMO) and an Intraaortic Balloon Pump (IABP) for refractory cardiogenic shock. She was in the intensive care unit for multiple weeks. Her cardiac function improved and she was able to be weaned off the mechanical support devices and vasopressors. She was discharged to a rehab facility with a good neurological outcome. Conclusion The use of ECMO contributed to the overall outcome of our patient. ECMO has a direct impact on drug pharmacokinetics. ECMO can be used to alter drug levels based on altering clearance as well as direct extraction from the circuit. Based on these results, implementation of early ECMO should be considered in patients with massive overdoses in refractory cardiogenic shock.

Sparsely Distributed, Pre-synaptic Kv3 K+ Channels Control Spontaneous Firing and Cross-Unit Synchrony via the Regulation of Synaptic Noise in an Auditory Brainstem Circuit.

Spontaneous subthreshold activity in the central nervous system is fundamental to information processing and transmission, as it amplifies and optimizes sub-threshold signals, thereby improving action potential initiation and maintaining reliable firing. This form of spontaneous activity, which is frequently considered noise, is particularly important at auditory synapses where acoustic information is encoded by rapid and temporally precise firing rates. In contrast, when present in excess, this form of noise becomes detrimental to acoustic information as it contributes to the generation and maintenance of auditory disorders such as tinnitus. The most prominent contribution to subthreshold noise is spontaneous synaptic transmission (synaptic noise). Although numerous studies have examined the role of synaptic noise on single cell excitability, little is known about its pre-synaptic modulation owing in part to the difficulties of combining noise modulation with monitoring synaptic release. Here we study synaptic noise in the auditory brainstem dorsal cochlear nucleus (DCN) of mice and show that pharmacological potentiation of Kv3 K+ currents reduces the level of synaptic bombardment onto DCN principal fusiform cells. Using a transgenic mouse line (SyG37) expressing SyGCaMP2-mCherry, a calcium sensor that targets pre-synaptic terminals, we show that positive Kv3 K+ current modulation decreases calcium influx in a fifth of pre-synaptic boutons. Furthermore, while maintaining rapid and precise spike timing, positive Kv3 K+ current modulation increases the synchronization of local circuit neurons by reducing spontaneous activity. In conclusion, our study identifies a unique pre-synaptic mechanism which reduces synaptic noise at auditory synapses and contributes to the coherent activation of neurons in a local auditory brainstem circuit. This form of modulation highlights a new therapeutic target, namely the pre-synaptic bouton, for ameliorating the effects of hearing disorders which are dependent on aberrant spontaneous activity within the central auditory system.

Arctigenin Exerts Neuroprotective Effect by Ameliorating Cortical Activities in Experimental Autoimmune Encephalomyelitis In Vivo.

Multiple sclerosis (MS) is a chronic disease in the central nervous system (CNS), characterized by inflammatory cells that invade into the brain and the spinal cord. Among a bulk of different MS models, the most widely used and best understood rodent model is experimental autoimmune encephalomyelitis (EAE). Arctigenin, a botanical extract from Arctium lappa, is reported to exhibit pharmacological properties, including anti-inflammation and neuroprotection. However, the effects of arctigenin on neural activity attacked by inflammation in MS are still unclear. Here, we use two-photon calcium imaging to observe the activity of somatosensory cortex neurons in awake EAE mice in vivo and found added hyperactive cells, calcium influx, network connectivity, and synchronization, mainly at preclinical stage of EAE model. Besides, more silent cells and decreased calcium influx and reduced network synchronization accompanied by a compensatory rise in functional connectivity are found at the remission stage. Arctigenin treatment not only restricts inordinate individually neural spiking, calcium influx, and network activity at preclinical stage but also restores neuronal activity and communication at remission stage. In addition, we confirm that the frequency of AMPA receptor-mediated spontaneous excitatory postsynaptic current (sEPSC) is also increased at preclinical stage and can be blunted by arctigenin. These findings suggest that excitotoxicity characterized by calcium influx is involved in EAE at preclinical stage. What is more, arctigenin exerts neuroprotective effect by limiting hyperactivity at preclinical stage and ameliorates EAE symptoms, indicating that arctigenin could be a potential therapeutic drug for neuroprotection in MS-related neuropsychological disorders.

Homosynaptic frequency-dependent depression by release site inactivation at neonatal hippocampal synapses in the stratum lacunosum-moleculare.

When activated at low frequencies (0.1-1 Hz), second postnatal week synapses onto the most distal part of the apical dendritic tree (stratum lacunosum-moleculare) of rat hippocampal CA1 pyramidal cells display a frequency-dependent synaptic depression not observed for the more proximal (stratum radiatum) synapses. Depression in this frequency range is thought of as a possible contributor to behavioural habituation. In fact, in contrast to the proximal synapses, the distal synapses provide more direct sensory information from the entorhinal cortex as well as from thalamic nuclei. The use of antagonists showed that the activation of GABAA , GABAB , NMDA, mGlu, kainate, adenosine, or endocannabinoid receptors was not directly involved in the depression, indicating it to be intrinsic to the synapses themselves. While the depression affected paired-pulse plasticity in a manner indicating a decrease in vesicle release probability, the depression could not be explained by a stimulus-dependent decrease in calcium influx. Despite affecting the synaptic response evoked by brief high-frequency stimulation (10 impulses, 20 Hz) in a manner indicating vesicle depletion, the depression was unaffected by large variations in release probability. The depression was found not only to affect the synaptic transmission at low frequencies (0.1-1 Hz) but also to contribute to the depression evolving during brief high-frequency stimulation (10 impulses, 20 Hz). We propose that a release-independent process directly inactivating release sites with a fast onset (ms) and long duration (up to 20 s) underlies this synaptic depression.

Astrocytic mitochondria in adult mouse brain slices show spontaneous calcium influx events with unique properties

Astrocytes govern critical aspects of brain function via spontaneous calcium signals in their soma and processes. A significant proportion of these spontaneous astrocytic calcium events are associated with mitochondria, however, the extent, sources, or kinetics of astrocytic mitochondrial calcium influx have not been studied in the adult mouse brain. To measure calcium influx into astrocytic mitochondria in situ, we generated an adeno-associated virus (AAV) with the astrocyte-specific GfaABC1D promoter driving expression of the genetically encoded calcium indicator, GCaMP6f tagged to mito7, a mitochondrial matrix targeted signal sequence. Using this construct, we observed AAV-mediated expression of GCaMP6f in adult mouse astrocytic mitochondria that co-localized with MitoTracker deep red (MTDR) in the dorsolateral striatum (DLS) and in the hippocampal stratum radiatum (HPC). Astrocytic mitochondria co-labeled with MTDR and GCaMP6f displayed robust, spontaneous calcium influx events in situ, with subcellular differences in calcium influx kinetics between somatic, branch, and branchlet mitochondria, and inter-regional differences between mitochondria in DLS and HPC astrocytes. Calcium influx into astrocytic mitochondria was strongly dependent on endoplasmic reticulum calcium stores, but did not require the mitochondrial calcium uniporter, MCU. Exposure to either glutamate, D1 or D2 dopamine receptor agonists increased calcium influx in some mitochondria, while simultaneously decreasing calcium influx in other mitochondria from the same astrocyte. These findings show that astrocytic mitochondria possess unique properties with regard to their subcellular morphology, mechanisms of calcium influx, and responses to neurotransmitter receptor agonists. Our results have important implications for understanding the role of astrocytic mitochondria during pathological processes.

A computational study of suppression of sharp wave ripple complexes by controlling calcium and gap junctions in pyramidal cells

ABSTRACT The hippocampus plays a key role in memory formation and learning. According to the concept of active systems memory consolidation, transiently stored memory traces are transferred from the hippocampus into the neocortex for permanent storage. This phenomenon relies on hippocampal network oscillations, particularly sharp wave ripples [SPW-Rs). In this process prior saved data in the hippocampus may be reactivated. Recent investigations reveal that several neurotransmitters and neuromodulators including norepinephrine, acetylcholine, serotonin, etc., suppress SPW-Rs activity in rodents’ hippocampal slices. This suppression of SPW-Rs may depend on various presynaptic and postsynaptic parameters including decrease in calcium influx, hyperpolarization/depolarization and alteration in gap junctions’ function in pyramidal cells. In this study, we demonstrate the impact of calcium influx and gap junctions on pyramidal cells for the modulation of SPW-Rs in a computational model of CA1. We used,SPW-Rs model with some modifications. SPW-Rs are simulated with gradual reduction of calcium and with decreasing conductance through gap junctions in PCs. Both, with calcium reduction as well as with conductance reduction through gap junctions, SPW-Rs are suppressed. Both effects add up synergistically in combination.

Poloxamer 188 Protects Isolated Adult Mouse Cardiomyocytes from Reoxygenation Injury.

Reperfusion injury is a complex pathological event involving processes that can lead to further disruption of the cell membrane and function following an ischemic event. Return of blood flow allows for the needed reperfusion; however, for a period of time before remaining viable cells stabilize, reperfusion results in additional cellular injury. In cardiomyocytes, loss of membrane integrity allows abnormal influx of extracellular calcium, leading to hyper‐contracture and cell death. Methods to improve the membrane integrity of cardiomyocytes overwhelmed by pathological disruptions, such as reperfusion injury, are needed to prevent cell death, because of the myocardium's limited ability to regenerate. Research has shown administration of the copolymer P(oloxamer) 188 before ischemia/reperfusion can protect cardiomyocytes through membrane stabilization. This study sought to determine whether the administration of P188 at the beginning of the clinically more relevant time of reperfusion after ischemia will attenuate any additional damage to cardiomyocytes by stabilizing membrane integrity to allow the cells to maintain function. Using an in‐vitro cardiomyocyte model subjected to hypoxia/reoxygenation to simulate ischemia/reperfusion injury, we show that reoxygenation significantly potentiates the injury caused by hypoxia itself. P188, with its unique combination of hydrophobic and hydrophilic chemical properties, and only delivered at the beginning of reoxygenation, dose‐dependently protected cardiomyocytes from injury due to reoxygenation by repairing cell membranes, decreasing calcium influx, and maintaining cellular morphology. Our study also shows the hydrophobic portion of P188 is necessary for the stabilization of cell membrane integrity in providing protection to cardiomyocytes against reoxygenation injury.

Neferine, a bisbenzylisoquinoline alkaloid of Nelumbo nucifera, inhibits glutamate release in rat cerebrocortical nerve terminals through 5-HT1A receptors

Neferine, a bisbenzylisoquinoline alkaloid present in Nelumbo nucifera, has been reported to exhibit neuroprotective effects. Because reduced glutamatergic transmission through inhibition of glutamate release has been proposed as a mechanism of neuroprotection, we investigated whether and how neferine inhibits glutamate release in the nerve terminals of the cerebral cortex of rats. The results demonstrated that neferine inhibits the glutamate release that is evoked by the potassium channel blocker 4-aminopyridine, doing so in a dose-dependent manner. This effect was prevented by removing extracellular calcium and blocking vesicular transporters or N- and P/Q-type calcium channels but not by blocking glutamate transporters. Neferine decreased the 4-aminopyridine-stimulated elevation in intrasynaptosomal calcium concentration; however, it had no effect on the synaptosomal membrane potential. The inhibition of glutamate release by neferine was also eliminated by the selective 5-hydroxytryptamine 1A (5HT1A) receptor antagonist WAY100635, Gi/o protein inhibitor pertussis toxin, adenylyl cyclase inhibitor MDL12330A, and protein kinase A inhibitor H89. Moreover, immunocytochemical analysis revealed the presence of 5-HT1A receptor proteins in the vesicular transporter of glutamate type 1 positive synaptosomes. The molecular docking study also demonstrated that neferine exhibited the highest binding affinity with 5-HT1A receptors (Autodock scores for 5-HA1A = −11.4 kcal/mol). Collectively, these results suggested that neferine activates 5-HT1A receptors in cortical synaptosomes, which decreases calcium influx and glutamate release through the activation of Gi/o protein and the inhibition of adenylyl cyclase/cAMP/protein kinase A cascade.

ORAI1 and ORAI2 modulate murine neutrophil calcium signaling, cellular activation, and host defense.

Calcium signals are initiated in immune cells by the process of store-operated calcium entry (SOCE), where receptor activation triggers transient calcium release from the endoplasmic reticulum, followed by opening of plasma-membrane calcium-release activated calcium (CRAC) channels. ORAI1, ORAI2, and ORAI3 are known to comprise the CRAC channel; however, the contributions of individual isoforms to neutrophil function are not well understood. Here, we show that loss of ORAI1 partially decreases calcium influx, while loss of both ORAI1 and ORAI2 completely abolishes SOCE. In other immune-cell types, loss of ORAI2 enhances SOCE. In contrast, we find that ORAI2-deficient neutrophils display decreased calcium influx, which is correlated with measurable differences in the regulation of neutrophil membrane potential via KCa3.1. Decreased SOCE in ORAI1-, ORAI2-, and ORAI1/2-deficient neutrophils impairs multiple neutrophil functions, including phagocytosis, degranulation, leukotriene, and reactive oxygen species (ROS) production, rendering ORAI1/2-deficient mice highly susceptible to staphylococcal infection. This study demonstrates that ORAI1 and ORAI2 are the primary components of the neutrophil CRAC channel and identifies subpopulations of neutrophils where cell-membrane potential functions as a rheostat to modulate the SOCE response. These findings have implications for mechanisms that modulate neutrophil function during infection, acute and chronic inflammatory conditions, and cancer.