Neurodegenerative diseases pose a significant challenge to modern medicine. Despite significant advances in neurology, current therapeutic approaches often prove insufficient to treat such disorders. This study investigates the neuroprotective effect of extracellular vesicles derived from glial derivates of human-induced pluripotent stem cells. The extracellular vesicle’s cargo was characterised by proteomic analysis. The neuroprotective effect was assessed using a model of glutamate excitotoxicity performed on a primary culture of cortical neuroglial cells. The viability of cells was estimated using the MTT test and morphometric analyses. A comprehensive methodology was applied to investigate intracellular mechanisms, integrating assessments of intracellular calcium concentrations, mitochondrial membrane potential, and targeted inhibition of the PI3K-Akt pathway. Transcriptomic analysis of neuroglial cultures was used to validate the role of obtained mechanisms of extracellular vesicle’s neuroprotective effect. The obtaining results demonstrated the improvement of neuronal survival by reducing intracellular calcium levels and stabilising mitochondrial membrane potential under glutamate-induced excitotoxicity via PI3K-Akt signalling pathway activation. Moreover, the vesicles contained proteins that contribute to preventing apoptotic processes, activating regeneration of the nervous system, and modulating calcium ion transport and are associated with redox processes. Further transcriptomic analyses of neuroglial cultures treated with EVs showed an up-regulation of genes associated with regeneration, inhibition of calcium ion transport, regulation of membrane depolarisation, and negative regulation of apoptotic pathways.

Duchenne muscular dystrophy (DMD) is a severe X-linked genetic disorder caused by an array of mutations in the dystrophin gene, with the most commonly mutated regions being exons 48-55. One of the several existing approaches to treat DMD is gene therapy, based on alternative splicing and mutant exon skipping. Testing of such therapy requires animal models that carry mutations homologous to those found in human patients. Here, we report the generation of two genetically modified mouse lines, named "insT" and "insG", with distinct mutations at the same position in exon 51 that lead to a frameshift, presumably causing protein truncation. Hemizygous males of both lines exhibit classical signs of muscular dystrophy in all muscle tissues except for the cardiac tissue. However, pathological changes are more pronounced in one of the lines. Membrane localization of the protein is reduced to the point of absence in one of the lines. Moreover, an increase in full-length isoform mRNA was detected in diaphragms of insG line mice. Although further work is needed to qualify these mutations as sole origins of dissimilarity, both genetically modified mouse lines are suitable models of DMD and can be used to test gene therapy based on alternative splicing.

The study purpose: To validate by polysomnography (PSG) tools the efficacy of deep abdominal breathing (AB) as a technique improving daytime nap in healthy subjects. Materials and methods: 43 healthy subjects participated in the study, of whom 22 were included into intervention group and 21 into control group. In the intervention group, nap PSGs were recorded for 30 min after performing AB for 15 minutes. In the control group, similar PSGs were recorded after 15 min of wakefulness. To assess the nap quality, standard sleep characteristics (latency, etc.) were determined from the subjects’ hypnograms. In the intervention group total sleep time was significantly longer and activation index was significantly lower than in control group, while sleep latency did not differ significantly. In addition, the electroencephalogram (EEG) spectrum power ratio in alpha (8–13 Hz) and theta (4–8 Hz) frequency bands was analyzed. Linear regression model of alpha/theta power ratio time series was constructed within the framework of statistical analysis. It was concluded based on comparison of coefficients of this model along with the time domain sleep characteristics, that AB exercise preceding daytime nap activates physiological mechanisms accelerating fall–asleep process and making sleep more stable. This finding may be useful in the development of non-invasive approaches to insomnia treatment.

Extracellular vesicles (EVs) have recently entered the group of modern neurotherapeutic agents for Alzheimer disease treatment. Glutamate excitotoxicity remains the most common and damaging consequence of this neurodegeneration. In this work, we investigate the neuroprotective mechanisms of a new type of EVs derived from human glial progenitor cells (hGPCs) using proteomic and inhibitor assays in the glutamate excitotoxicity model. EVs were obtained by ultracentrifugation from conditioned medium of hGPCs. Glutamate excitotoxicity model was established using rat pups (P0) primary cultures of cortical neurons. To perform proteomic analysis of EV-GPCs was used an Ultimate 3000 Nano LC System chromatography system interfaced with a Q Exactive HF mass spectrometer. The proteins peak list was generated and analyzed by MASCOT using the UniProtKB database. Proteins systematization and categorization was performed using the String 8.0 database. To confirm the activation of founded PI3K-Akt signaling pathway a selective inhibitor AS605240 of PI3Kγ subunit was used. In the model of glutamate excitotoxicity, a 37.5% decrease in the viable neurons number was observed with the addition of glutamate compared to the control group. The presence of EV-hGPCs (3 μg/mL) significantly increased survival rates by 20%, whereas addition of EV-GPCs (10 μg/mL) increased survivability to control values. According to the bioinformatic analysis, the most represented protein groups of the signaling pathways (KEGG database) in EVs was the "PI3K-Akt signaling pathway" (51 proteins). To confirm that EVs can activate this pathway in cells, inhibitor assay was performed using AS605240 (1 μg/mL). Its addition completely offset the drug’s neuroprotective effect. The obtained results indicate that EV-hGPCs have neuroprotective effect on the glutamate excitotoxicity model. This is due to the proteins they contain that activate the PI3K-Akt signaling pathway. Funding : The work was financially supported by RSF grant № 23-15-00362 “Study of mechanisms of the therapeutic action of extracellular vesicles derived from human glial progenitor cells on the model of Alzheimer's disease”

Currently, stem cells technology is an effective tool in regenerative medicine. Cell therapy is based on the use of stem/progenitor cells to repair or replace damaged tissues or organs. This approach can be used to treat various diseases, such as cardiovascular, neurological diseases, and injuries of various origins. The mechanisms of cell therapy therapeutic action are based on the integration of the graft into the damaged tissue (replacement effect) and the ability of cells to secrete biologically active molecules such as cytokines, growth factors and other signaling molecules that promote regeneration (paracrine effect). However, cell transplantation has a number of limitations due to cell transportation complexity and immune rejection. A potentially more effective therapy is using only paracrine factors released by stem cells. Secreted factors can positively affect the damaged tissue: promote forming new blood vessels, stimulate cell proliferation, and reduce inflammation and apoptosis. In this work, we have studied the anti-inflammatory and neuroprotective effects of proteins with a molecular weight below 100 kDa secreted by glial progenitor cells obtained from human induced pluripotent stem cells. Proteins secreted by glial progenitor cells exerted anti-inflammatory effects in a primary glial culture model of LPS-induced inflammation by reducing nitric oxide (NO) production through inhibition of inducible NO synthase (iNOS). At the same time, added secreted proteins neutralized the effect of glutamate, increasing the number of viable neurons to control values. This effect is a result of decreased level of intracellular calcium, which, at elevated concentrations, triggers apoptotic death of neurons. In addition, secreted proteins reduce mitochondrial depolarization caused by glutamate excitotoxicity and help maintain higher NADH levels. This therapy can be successfully introduced into clinical practice after additional preclinical studies, increasing the effectiveness of rehabilitation of patients with neurological diseases.

OBJECTIVE:To test the hypothesis of the difference between 3 means of sleep latency (SL) during falling asleep: accompanied by audio stimulus embedded with binaural beats (BB); after listening to suggestive body relaxation instructions; accompanied by audio stimulus embedded with BB after listening to suggestive body relaxation instructions (that is the combination of 1 and 2). MATERIAL AND METHODS:For the purpose of the study, a special Android application was developed and installed on the subjects' individual smartphones. The application assumed screen tapping test to control for fall-asleep process. The data of 63 subjects presented with the 3 types of sound stimuli mentioned above in a counterbalanced scheme were analyzed. RESULTS:Statistical analysis confirmed the initial hypothesis about the dependence of LS on the type of sound stimulus (p<0.05). Pairwise SL comparison showed reliable difference between stimuli (3) - 1149±113 s, and (1) - 1469±89 s (p<0.01). SL for the stimulus (2) had an intermediate value of 1269±112 s (difference from (1) at a trend level). CONCLUSION:The use of background sound embedded with BBs enhances the effect of suggestive instructions to improve sleep. But it is the suggestion as a psychotherapeutic technique that is determinant.

The scratch test is used as an experimental in vitro model of mechanical damage to primary neuronal cultures to study the mechanisms of cell death in damaged areas. The involvement of NMDA receptors in processes leading to delayed neuronal death, due to calcium dysregulation and synchronous mitochondrial depolarization, has been previously demonstrated. In this study, we explored the neuroregenerative potential of Pro-Gly-Pro (PGP)—an endogenous regulatory peptide with neuroprotective and anti-inflammatory properties and a mild chemoattractant effect. Mechanical injury to the primary neuroglial culture in the form of a scratch caused acute disruption of calcium homeostasis and mitochondrial functions. This was accompanied by neuronal death alongside changes in the profile of neuronal markers (BDNF, NSE and GFAP). In another series of experiments, under subtoxic doses of glutamate (Glu, 33 μM), delayed changes in [Ca2+]i and ΔΨm, i.e., several days after scratch application, were more pronounced in cells in damaged neuroglial cultures. The percentage of cells that restored the initial level of [Ca2+]i (p < 0.05) and the rate of recovery of ΔΨm (p < 0.01) were decreased compared with undamaged cells. Prophylactic application of PGP (100 μM, once) prevented the increase in [Ca2+]i and the sharp drop in mitochondrial potential [ΔΨm] at the time of scratching. Treatment with PGP (30 μM, three or six days) reduced the delayed Glu-induced disturbances in calcium homeostasis and cell death. In the post-glutamate period, the surviving neurons more effectively restored the initial levels of [Ca2+]i (p < 0.001) and Ψm (p < 0.0001). PGP also increased intracellular levels of BDNF and reduced extracellular NSE. In the context of the peptide’s therapeutic effect, the recovery of the damaged neuronal network occurred faster due to reduced astrogliosis and increased migration of neurons to the scratch area. Thus, the peptide PGP has a neuroprotective effect, increasing the survival of neuroglial cells after mechanical trauma in vitro by reducing cellular calcium overload and preventing mitochondrial dysfunction. Additionally, the tripeptide limits the post-traumatic consequences of mechanical damage: it reduces astrogliosis and promotes neuronal regeneration.

The multifunctional promyelocytic leukemia protein (PML) is involved in the regulation of various cellular processes in both physiological and pathological conditions. Specifically, PML is one of the inositol-1,4,5-trisphosphate receptors (IP3Rs) activity regulators and can influence Ca2+ transport from the endoplasmic reticulum (ER) to mitochondria. In this work, the effects of PML knockout on calcium homeostasis in the cytosol, ER, and mitochondria of HeLa cells were studied upon stimulation with histamine, which induces Ca2+ mobilization from the ER via IP3Rs. We utilized calcium indicators with different subcellular localizations, including synthetic dyes Fura-2 (cytosolic), Xrhod-5F (mitochondrial), and protein sensor R-CEPIAer (ER), as well as mitochondrial potential-sensitive probes Rh123 and TMRM. Our results show that PML knockout induced changes in HeLa cell and mitochondrial morphology, slightly decreased basal and integral Ca2+ levels, enhanced mitochondrial Ca2+ uptake from the cytoplasm, and maintained residual mitochondrial potential after depolarization. Additionally, it reduced the Ca2+ pool in ER membranes not associated with histamine receptor activation and, consequently, IP3Rs. These findings suggest that changes in calcium ion transport due to PML knockout in HeLa cells affect mitochondrial activity.

Hydrogen sulfide (H2S), which under physiological conditions exists in cells mainly in the form of the HS– anion, is considered as a gaseous transmitter of inter- and intracellular signals along with nitrogen monoxide and carbon monoxide. Analysis of the dynamics of H2S content in living cells is impossible without the creation of sensitive and specific probes. The group of K.H. Ahn synthesized several acedan-based compounds, which in the presence of H2S attached a sulfhydryl group, forming fluorescent carbocyclic compounds. According to the spectral characteristics and reaction rate with H2S, the optimal substance was P3, which forms the carbocyclic compound csP3 with the same large Stokes shift as P3 (approx. 130 nm) and has a brighter fluorescence. In this work, we tested the suitability of csP3 for recording changes in H2S in solutions simulating the minimum salt composition of the intracellular medium, as well as in cells of primary neuronal culture from the rat cerebral cortex. It was found that the fluorescence intensity of csP3, which was formed when Na2S (H2S donor, 100 and 300 µM) was added to the P3 solution, differed for solutions corresponding in salt composition to the extracellular medium and cytosol. In both cases, fluorescence increased in the presence of bicarbonate (NaHCO3, 10 mM). A decrease in the polarity of solutions due to the addition of dimethyl sulfoxide (30

Objective. To test the hypothesis that there are differences between three mean values of sleep latency (SL) when falling asleep: when accompanied by audio stimuli based on binaural beats (BB), after listening to suggestive instructions on how to relax the body, and accompanied by BB-based audio stimuli after listening to suggestive instructions for relaxing the body (i.e., 1 and 2 combined). Materials and methods. A special app was written for the Android operating system for the purpose of conducting this study and was installed on subjects’ individual smartphones. The app included a screen touch test as a control for falling asleep. Data were analyzed from 63 subjects who were presented with the three types of sound stimuli noted above in a counterbalanced design. Results. Statistical analysis confirmed the hypothesis noted above that the SL depended on the type of sound stimulus (p < 0.05). Pairwise comparison of SL showed significant differences between stimuli (3) (1149 ± 113 sec) and (1) (1469 ± 89 sec) (p < 0.01). The SL for stimulus (2) had an intermediate value of 1269 ± 112 sec (differing from stimulus (1) at the level of a trend). Conclusion. The use of a sound background consisting of BB enhances the effect of suggestive instructions in improving sleep. However, the decisive role belongs to suggestion as a psychotherapeutic technique.

OBJECTIVE:The aim of this study was to test on a large group of subjects the hypothesis that sleep latency (SL) does not depend on the nature of low-frequency beats embedded into monotonous sound stimulus supplied through the fall-asleep process. Specifically, it does not depend on whether those beats are monaural (MB) or binaural (BB).MATERIAL AND METHODS:A special application for Android OS was developed and installed on the individual smartphones of 221 subjects for the purpose of the study. Three attempts were performed with each of them using 3 different kinds of monotonous sound supplied according to counterbalanced design. Three kinds of sound were identical in pitch but differed in the beat presence and type: BB, MB or sham (sound without beats).RESULTS:Repeated measures analysis of variance (rANOVA) revealed no significant statistical effect of stimulus type on SL (p=0.21). A pairwise comparison of SL for different stimulation conditions showed the null hypothesis significance level adjusted according to multiple comparison correction to be p=1.0. Thus, in this experiment SL did not significantly depend on the monotonous sound stimulus type: MB, BB, or sham.CONCLUSION:The software application developed is useful as universal platform to assess at home conditions the impact of various external factors on fall-asleep process.

Актуальность. При исследованиях внутриклеточного сигналинга и межнейрональной передачи сигнала в мозге в норме и при патологии все большее применение находят трансгенные животные, экспрессирующие в нейронах флуоресцентный Са2+-сенсор. Вместе с тем, пока недостаточно исследовано, насколько изменения кальциевого гомеостаза в интактном мозге соотносятся с гораздо более подробно изученными изменениями этого важнейшего параметра в нейроглиальных культурах, служащих модельными системами живого мозга. Целью работы было на модели ишемического инсульта выяснить в какой степени изменения внутриклеточной концентрации Са2+ ([Ca2+]) в мозге трансгенной мыши, измеренные с помощью флуоресцентного белкового сенсора GCaMP6f, соотносятся с изменениями [Ca2+] в первичных нейроглиальных культурах из кортекса этих животных. Методы. Методом широкопольной оптической нейровизуализации (ШОН) измерены изменения концентрации свободного Са2+ в цитозоле нейронов ([Ca2+]c) головного мозга мышей. Измерения проводили перед и после фотоиндуцированного инсульта в сенсомоторной зоне коры. Изменения [Ca2+]c отслеживали по флуоресценции GCaMP6f, экспрессируемого в нейронах кортекса. На первичных нейроглиальных культурах из коры головного мозга мышей той же линии проверено влияние эксайтотоксических доз глутамата (Glu) на [Ca2+]c и на изменения средней концентрации свободного Са2+ в цито- и нуклеоплазме ([Ca2+]i). Измерения [Ca2+]i выполнены методом флуоресцентной микроскопии с использованием синтетических Са2+- индикаторов Fura-2 и Fura-FF. В культивируемых нейронах дополнительно к измерениям кальциевого гомеостаза выполнены измерения внутриклеточного рН (pHi), митохондриального потенциала (ΔΨm) и эндогенной флуоресценции NADH. Результаты. Фотоиндуцированная ишемия вызывает сильный рост [Ca2+]c в зоне облучения ~1,1 мм2 (n=9). В течение сутокобласть высокой [Ca2+]c расширяется до ~6 мм2, но к 7-м сут практически возвращается к размерам необратимого повреждения. В нейроглиальных культурах из коры головного мозга мышей этой же линии кинетика изменений [Ca2+]c, индуцированных Glu, напоминает кинетику [Ca2+]i, однако [Ca2+]c имеет значительно меньшую амплитуду при развитии отсроченной кальциевой дисрегуляции (ОКД). Сопоставление изменений [Ca2+] и pHi показывает, что различия могут быть обусловлены тушением флуоресценции GCaMP6f при закислении цитозоля в результате эксайтотоксического действия Glu. Заключение. Сопоставление сигналов экспрессируемого нейронами Са2+-сенсора GCaMP6f в мозге и синтетических Са2+-индикаторов в первичных нейроглиальных культурах, полученных из животных той же линии, показывает, что феномен ОКД, впервые обнаруженный в культурах, вероятно, реализуется и в нейронах целого мозга при инсульте. Вместе с тем, необходимо учитывать, что относительные изменения [Ca2+]c на разных стадиях развития ишемического повреждения и последующего восстановления мозга после фотоиндуцированного инсульта, могут быть искажены за счет влияния pHi на флуоресценцию белкового сенсора. Relevance. In studies of brain intracellular and intercellular signaling in normal and pathological conditions, transgenic animals expressing a fluorescent Ca2+ sensor in neurons are increasingly used. Calcium homeostasis was studied in detail in primary neuroglial cultures, which serve as model systems of the living brain. But how change in calcium homeostasis during ischemic conditions in the intact brain correlate with experiments on cell cultures has been poorly studied so far. The aim of this work was to compare ischemia-driven changes in the intracellular concentration of Ca2+ ([Ca2+]) in vivo and in vitro: in the brain of transgenic mice, using the GCaMP6f fluorescent protein sensor, and in the neuroglial cell culture, obtained from the cortex of these animals. Methods. Changes in the cytosolic concentration of free Ca2+ ([Ca2+]c) in the neurons was measured by wide-field optical imaging (WFOI). Measurements were taken before and after photothrombotic stroke performed in the sensorimotor cortex. Changes in [Ca2+]c were monitored by the fluorescence of GCaMP6f expressed in cortical neurons. In primary neuroglial cultures from the cerebral cortex glutamate (Glu) in excitotoxic doses was used to model ischemic injury. Measurements of averaged Ca2+ concentration in the cyto- and nucleoplasm ([Ca2+]i) were carried out by fluorescence microscopy using synthetic Ca2+ indicators Fura-2 and Fura-FF. In addition, measurements of intracellular pH (pHi) and mitochondrial potential (ΔΨm) were carried out in vitro. Results. Photothrombotic stroke caused a strong increase in [Ca2+]c in the illuminated zone ~1.1 mm2 (n=9). During the 24 hours, the area with high [Ca2+]c expands to ~6 mm2, but by the 7th day it almost returns to the size of the primary damage. In neuroglial cultures from the cerebral cortex of the same mice strain, the [Ca2+]c kinetics measured by GCaMP6f resembles the [Ca2+]i kinetics, but [Ca2+]c has a significantly lower amplitude during the development of delayed calcium deregulation (DCD). Comparison of changes in [Ca2+]i and pHi shows that the differences may be due to the quenching of GCaMP6f fluorescence due to cytosol acidification as a result of the excitotoxic Glu action. Conclusion. Comparison of measurements by the genetically-encoded GCaMP6f Ca2+ sensor in vivo and by the synthetic Ca2+ indicators in primary neuroglial cultures shows that the DCD phenomenon, first discovered in cultures, is probably realized in intact brain neurons in stroke. It should be taken into account that the relative changes in [Ca2+]c can be distorted due to the effect of pHi on the fluorescence of the protein sensor at different stages of ischemia development and subsequent brain recovery after a photothrombotic stroke.

Objectives . To test the hypothesis that sleep latency (SL) does not depend on the nature (monaural (MB) or binaural (BB)) of the low-frequency beats embedded in a monotonous sound stimulus presented during the process of falling asleep in a large group of subjects. Materials and methods . An Android app was written for the study and installed on 221 subjects’ smartphones. Each subject took part in three experiments, with three different types of monotonous sound presented following a counterbalanced scheme; sounds were identical in pitch but differed in terms of the presence and type of beats: BB, MB, and control (without beats) sounds were used. Results . Repeat measures analysis of variance (rANOVA) showed no statistically significant effect of stimulus type on SL ( p = 0.21). Pairwise comparison of SL for different stimulation conditions showed a significance level of the null hypothesis, corrected for multiple comparisons, of p = 1.0. Thus, SL was not significantly dependent on the type of monotonous sound stimulus used in these experiments: MB, BB, or control. Conclusions . The software app developed here is useful as a universal platform for assessing the impact of various external factors on the process of falling asleep at home.

“Binaural beats” (BB) is a type of sound stimulus being studiedas non-invasive tools to promote sleep. BB is a sensation occuringwhen two monotonous sounds of slightly different pitch are suppliedto the listener’s right and left ear separately. This type of stimulationhas the advantage to be perceived even at very low sound volumesessentially bordering the hearing threshold, so such a stimuluscreates little disturbance to sleep. However, the patterns of itseffect are difficult to study due to the weakness of electric brainwaveresponse. The purpose of present paper is to check applicabilityof the brainwave entrainment hypothesis to auditory stimuli withembedded BB delivered during short-term human nap. The results demonstratevariability of auditory steady state response (ASSR) depending onBB frequency of the stimulus as well as on sleep stage that is not envisagedby the above hypothesis. It should be taken into account when predictingthe effect of BB-based noninvasive sleep aids. Furthermore, theanalysis of ASSR spectra allows the hypothesis to be put that thehuman brain falling asleep is a self-adjusting system in relationto incoming sound stimuli, for it strengthens the elements of auditoryresponse which contribute to deepening of naturally evolving sleep.

Objective. To test the hypothesis that music with a binaural beat (BB) effect can increase activation of the parasympathetic compartment of the autonomic nervous system (PANS) as daytime sleep gets deeper. Materials and methods. Comparison parameters were the power of the high-frequency components of the spectrum of variability of subjects’ heart rate computed in sequentia l 2-min periods during 20-min naps. Parameters were compared on going to sleep on the background of music with the BB effect (stimulus) and going to sleep in the quiet (control). Results and conclusions. Statistical comparison demonstrated a higher level of activation of the PANS on going to sleep on the background of the stimulus as compared with the control. This is consistent with conclusions in other reports on the positive influence of sound stimuli with the BB effect on the PANS.

Glutamate excitotoxicity is involved in the pathogenesis of many disorders, including stroke, traumatic brain injury, and Alzheimer’s disease, for which central insulin resistance is a comorbid condition. Neurotoxicity of glutamate (Glu) is primarily associated with hyperactivation of the ionotropic N-methyl-D-aspartate receptors (NMDARs), causing a sustained increase in intracellular free calcium concentration ([Ca2+]i) and synchronous mitochondrial depolarization and an increase in intracellular superoxide anion radical (O2–•) production. Recently, we found that insulin protects neurons against excitotoxicity by decreasing the delayed calcium deregulation (DCD). However, the role of insulin in O2–• production in excitotoxicity still needs to be clarified. The present study aims to investigate insulin’s effects on glutamate-evoked O2–• generation and DCD using the fluorescent indicators dihydroethidium, MitoSOX Red, and Fura-FF in cortical neurons. We found a linear correlation between [Ca2+]i and [O2–•] in primary cultures of the rat neuron exposed to Glu, with insulin significantly reducing the production of intracellular and mitochondrial O2–• in the primary cultures of the rat neuron. MK 801, an inhibitor of NMDAR-gated Ca2+ influx, completely abrogated the glutamate effects in both the presence and absence of insulin. In experiments in sister cultures, insulin diminished neuronal death and O2 consumption rate (OCR).

It is considered that glutamate excitotoxicity may be a major factor in the pathological death of neurons and mediate the development of neurodegenerative diseases in humans. Here, we show that isoliquiritigenin (ILG) at a concentration of 0.5–5 µM protects primary neuroglial cell culture from glutamate-induced death (glutamate 100 µM). ILG (1 µM) prevented a sharp increase in [Ca2+]i and a decrease in mitochondrial potential (ΔΨm). With the background action of ILG (1–5 µM), there was an increase in oxygen consumption rate (OCR) in response to glutamate, as well as in reserve respiration. The neuroprotective effect of ILG (5 µM) was accompanied by an increase in non-mitochondrial respiration. The results show that ILG can protect cortical neurons from death by preventing the development of calcium deregulation and limiting mitochondrial dysfunction caused by a high dose of glutamate. We hypothesize that ILG will be useful in drug development for the prevention or treatment of neurodegenerative diseases accompanied by glutamate excitotoxicity.