Calcium Signaling In Neurons Research Articles

Alterations in store-operated calcium entry in neurodegenerative pathologies: history, facts, perspectives

Neurodegenerative disorders, along with cardiovascular and oncological pathologies, are one of the most actual issues facing modern medicine. Therefore, the study of the molecular mechanisms of their pathogenesis and the search for new drug targets is highly demanded. Neuronal calcium signalling has attracted close attention, as altered calcium homeostasis has been demonstrated in the pathogenesis of various neurodegenerative diseases. In this review, we focus on one of the most ubiquitous and important pathways for calcium uptake: store-operated calcium entry. Here we describe studies demonstrating disturbances in store-operated calcium entry in various neurodegenerative pathologies, including Alzheimer’s, Parkinson’s and Huntington’s diseases. Also, we analyse the molecular determinants underlying these disturbances and propose ways for pharmacological correction of altered calcium signaling. The information summarized in the review will allow us to consider store-operated calcium channels as promising targets for the drug development in order to treat neurodegenerative pathologies and outline further promising directions for the investigation.

GABAergic neurons in the central amygdala promote emergence from isoflurane anesthesia in mice.

Recent evidence indicates that general anesthesia and sleep-wake behavior share some overlapping neural substrates. GABAergic neurons in the central amygdala (CeA) have a high firing rate during wakefulness and play a role in regulating arousal-related behaviors. The objective of this study is to investigate whether CeA GABAergic neurons participate in the regulation of isoflurane general anesthesia and uncover the underlying neural circuitry. Fiber photometry recording was used to determine the changes in calcium signals of CeA GABAergic neurons during isoflurane anesthesia in Vgat-Cre mice. Chemogenetic and optogenetic approaches were used to manipulate the activity of CeA GABAergic neurons, and a righting reflex test was used to determine the induction and emergence from isoflurane anesthesia. Cortical electroencephalogram (EEG) recording was used to assess the changes in EEG spectral power and burst-suppression ratio during 0.8% and 1.4% isoflurane anesthesia, respectively. Both male and female mice were used in this study. The calcium signals of CeA GABAergic neurons decreased during the induction of isoflurane anesthesia and was restored during the emergence. Chemogenetic activation of CeA GABAergic neurons delayed induction time (mean ± SD, vehicle vs. clozapine-N-oxide: 58.75±5.42 s vs. 67.63±5.01 s; n=8, P=0.0017) and shortened emergence time (385.50±66.26 s vs. 214.60±40.21 s; n=8, P=0.0017) from isoflurane anesthesia. Optogenetic activation of CeA GABAergic neurons produced a similar effect. Furthermore, optogenetic activation decreased EEG delta power (Pre-stim vs. Stim: 46.63%±4.40% vs. 34.16%±6.47%; n=8, P=0.0195) and burst-suppression ratio (83.39%±5.15% vs. 52.60%±12.98%; n=8, P=0.0002). Moreover, optogenetic stimulation of terminals of CeA GABAergic neurons in the basal forebrain (BF) also promoted cortical activation and accelerated behavioral emergence from isoflurane anesthesia. Our results suggest that CeA GABAergic neurons play a role in general anesthesia regulation, which facilitates behavioral and cortical emergence from isoflurane anesthesia through the GABAergic CeA-BF pathway.

A Non-Invasive and DNA-free Approach to Upregulate Mammalian Voltage-Gated Calcium Channels and Neuronal Calcium Signaling via Terahertz Stimulation.

Mammalian voltage-gated calcium channels (CaV) play critical roles in cardiac excitability, synaptic transmission, and gene transcription. Dysfunctions in CaV are implicated in a variety of cardiac and neurodevelopmental disorders. Current pharmacological approaches to enhance CaV activity are limited by off-target effects, drug metabolism issues, cytotoxicity, and imprecise modulation. Additionally, genetically-encoded channel activators and optogenetic tools are restricted by gene delivery challenges and biosafety concerns. Here a novel terahertz (THz) wave-based method to upregulate CaV1.2, a key subtype of CaV, and boost CaV1-mediated Ca2+ signaling in neurons without introducing exogenous DNA is presented. Using molecular dynamics simulations, it is shown that 42.5 THz (7.05µm, 1418cm-1) waves enhance Ca2+ conductance in CaV1.2 by resonating with the stretching mode of the -COO- group in the selectivity filter. Electrophysiological recordings and Ca2+ imaging confirm that these waves rapidly, reversibly, and non-thermally increase calcium influx of CaV1.2 in HEK293 cells and induce acute Ca2+ signals in neurons. Furthermore, this irradiation upregulates critical CaV1 signals, including CREB phosphorylation and c-Fos expression, in vitro and in vivo, without raising significant biosafety risks. This DNA-free, non-invasive approach offers a promising approach for modulating CaV gating and Ca2+ signaling and treating diseases characterized by deficits in CaV functions.

Infrared neuroglial modulation of spinal locomotor networks

Infrared neural stimulation (INS) emerges as a promising tool for stimulating the nervous system by its high spatial precision and absence of the use of exogenous agents into the tissue, which led to the first successful proof of concept in human brain. While neural networks have been the focal point of INS research, this technique is also non cell type specific as it triggers activity in non electrically excitable cells. Despite increasing interest, there remains to demonstrate well defined simultaneous astrocytic and neuronal signals in response to INS. Using calcium imaging, we show that INS has the capacity to initiate calcium signaling in both astrocytes and neurons simultaneously from the rostral lumbar spinal cord, each exhibiting distinct temporal and amplitude characteristics. Importantly, the mechanism underlying infrared-induced neuronal and astrocytic calcium signaling differ, with neuronal activity relying on sodium channels, whereas induced astrocytic signaling is predominantly influenced by extracellular calcium and TRPV4 channels. Furthermore, our findings demonstrate the frequency shift of neuronal calcium oscillations through infrared stimulation. By deepening our understanding in INS fundamentals, this technique holds great promise for advancing neuroscience, deepening our understanding of pathologies, and potentially paving the way for future clinical applications.

Expanded ATXN1 alters transcription and calcium signaling in SCA1 human motor neurons differentiated from induced pluripotent stem cells

Spinocerebellar ataxia type 1 (SCA1) is a dominantly inherited and lethal neurodegenerative disease caused by the abnormal expansion of CAG repeats in the ATAXIN-1 (ATXN1) gene. Pathological studies identified dysfunction and loss of motor neurons (MNs) in the brain stem and spinal cord, which are thought to contribute to premature lethality by affecting the swallowing and breathing of SCA1 patients. However, the molecular and cellular mechanisms of MN pathogenesis remain unknown.To study SCA1 pathogenesis in human MNs, we differentiated induced pluripotent stem cells (iPSCs) derived from SCA1 patients and their unaffected siblings into MNs. We examined proliferation of progenitor cells, neurite outgrowth, spontaneous and glutamate-induced calcium activity of SCA1 MNs to investigate cellular mechanisms of pathogenesis. RNA sequencing was then used to identify transcriptional alterations in iPSC-derived MN progenitors (pMNs) and MNs which could underlie functional changes in SCA1 MNs. We found significantly decreased spontaneous and evoked calcium activity and identified dysregulation of genes regulating calcium signaling in SCA1 MNs. These results indicate that expanded ATXN1 causes dysfunctional calcium signaling in human MNs.

Navigating the Fractional Calcium Dynamics of Orai Mechanism in Polar Dimensions.

Calcium plays a crucial role as a second messenger in neuronal signal transduction pathways. The influx of calcium ions through various physicochemical gating channels activates neuronal calcium signaling. The Endoplasmic Reticulum (ER) is a significant intracellular structure that sequesters calcium and controls signaling through SERCA, IPR, and leak channel mechanisms. Disruption of calcium dynamics can trigger intrinsic dyshomeostasis, cell damage, and apoptosis. The present study articulates a Caputo fractional time derivative in the polar coordinate dimensions to investigate the role of nonlocal calcium-free ions in the neuron through the Orai channel, and ER fluxes, incorporating various physiological parameters. The solution was obtained through the hybrid integral transform technique for analytical form. The closed form was generated using Green's function in terms of Mainardi and Wright's functions. Our simulation uncovered the calcium concentration bandwidth of interaction with different neuronal parameters. Parameters and calcium ion synergy show normal and Alzheimer's disease-impacted interaction through different illustrations. Our simulation reveals that S100B and BAPTA have significant calcium-controlling behavior.

Calcium homeostasis restoration in pyramidal neurons through micrometer-scale wireless electrical stimulation in spinal cord injured mice

Spinal Cord Injury (SCI) is a significant neurological disorder that can result in severe motor and cognitive impairments. Neuronal regeneration and functional recovery are critical aspects of SCI treatment, with calcium signaling being a crucial indicator of neuronal excitability. In this study, we utilized a murine model to investigate the effects of targeted wireless electrical stimulation (ES) on neuronal activity following SCI. After establishing a complete SCI model in normal mice, flexible electrodes were implanted, and targeted wireless ES was administered to the injury site. We employed fiber-optic photometric in vivo calcium imaging to monitor calcium signals in pyramidal neurons within the CA3 region of the hippocampus and the M1 region of the primary motor cortex. The experimental results demonstrated a significant reduction in calcium signals in CA3 and M1 pyramidal neurons following SCI (reduced by 76 % and 59 %, in peak respectively). However, the application of targeted wireless ES led to a marked increase in calcium signals in these neurons (increased by 118 % and 69 %, in peak respectively), indicating a recovery of calcium activity. These observations suggest that wireless ES has a positive modulatory effect on the excitability of pyramidal neurons post-SCI. Understanding these mechanisms is crucial for developing therapeutic strategies aimed at enhancing neuronal recovery and functional restoration following spinal cord injuries.

The activity of cholinergic neurons in the basal forebrain interferes with anesthesia-arousal process of propofol

Previous research has demonstrated that basal forebrain (BF) regulates arousal during propofol anesthesia. However, as the BF comprises cholinergic neurons alongside two other types of neurons, the specific role of cholinergic neurons has not been definitively elucidated. In our study, calcium signal imaging was utilized to monitor the real-time activities of cholinergic neurons in the BF during propofol anesthesia. Additionally, we selectively stimulated these neurons to investigate EEG and behavioral responses during propofol anesthesia. Furthermore, we specifically lesioned cholinergic neurons in the BF to investigate the sensitivity to propofol and the induction time. The results revealed that propofol suppressed calcium signals of cholinergic neurons within the BF following intraperitoneal injection. Notably, upon recovery of the righting reflex, the calcium signals partially recovered. Spectral analysis of the EEG elucidated that optical stimulation of cholinergic neurons led to a decrease in δ power underlie propofol anesthesia. Conversely, depletion of cholinergic neurons in the BF enhanced sensitivity to propofol and shortened the induction time. These findings clarify the role of cholinergic neurons in the anesthesia-arousal process, as well as the depth and the sensitivity of propofol anesthesia.

Glioblastoma disrupts cortical network activity at multiple spatial and temporal scales.

The emergence of glioblastoma in cortical tissue initiates early and persistent neural hyperexcitability with signs ranging from mild cognitive impairment to convulsive seizures. The influence of peritumoral synaptic density, expansion dynamics, and spatial contours of excess glutamate upon higher order neuronal network modularity is unknown. We combined cellular and widefield imaging of calcium and glutamate fluorescent reporters in two glioblastoma mouse models with distinct synaptic microenvironments and infiltration profiles. Functional metrics of neural ensembles are dysregulated during tumor invasion depending on the stage of malignant progression and tumor cell proximity. Neural activity is differentially modulated during periods of accelerated and inhibited tumor expansion. Abnormal glutamate accumulation precedes and outpaces the spatial extent of baseline neuronal calcium signaling, indicating these processes are uncoupled in tumor cortex. Distinctive excitability homeostasis patterns and functional connectivity of local and remote neuronal populations support the promise of precision genetic diagnosis and management of this devastating brain disease.

Mechanisms of synapse-to-nucleus calcium signalling in striatal neurons and impairments in Huntington's disease.

Huntington's disease (HD) is a monogenic disorder with autosomal dominant inheritance. In HD patients, neurons in the striatum and cortex degenerate, leading to motor, psychiatric and cognitive disorders. Dysregulated synaptic function and calcium handling are common in many neurodegenerative diseases, including HD. N-methyl-D-aspartate (NMDA) receptor function is enhanced in HD at extrasynaptic sites, altering the balance of calcium-dependent neuronal survival versus death signalling pathways. Endoplasmic reticulum (ER) calcium handling is also abnormal in HD. The ER, which is continuous with the nuclear envelope, is purportedly involved in nuclear calcium signalling; based on this, we hypothesised that nuclear calcium signalling is altered in HD. We explored this hypothesis with calcium imaging techniques, including simultaneous epifluorescent imaging of cytosolic and nuclear calcium using jRCaMP1b and GCaMP3 sensors, respectively, in striatal spiny projection neurons in cortical-striatal co-cultures from the YAC128 mouse model of HD. Our data show contributions from a variety of calcium channels to nuclear calcium signalling. NMDA receptors (NMDARs) play an essential role in initiating action potential-dependent calcium signalling to the nucleus, and ryanodine receptors (RyR) contribute to both cytosolic and nuclear calcium signals. Unlike previous reports in glutamatergic hippocampal and cortical neurons, we found that in GABAergic striatal neurons, L-type voltage-gated calcium channels (CaV) contribute to cytosolic, but not nuclear calcium signalling. Calcium imaging also suggests impairments in nuclear calcium signalling in HD striatal neurons, where spontaneous action potential-dependent calcium transients in the nucleus were smaller in YAC128 striatal neurons compared to those of wild-type (WT). Our results elucidate mechanisms involved in action potential-dependent nuclear calcium signalling in GABAergic striatal neurons, and have revealed a clear deficit in this signalling in HD.

Simulation of nonlinear system dynamics of calcium and dopamine signaling in neurons

Simulation of nonlinear system dynamics of calcium and dopamine signaling in neurons

Increased Neuronal Nitric Oxide Synthase in Alzheimer's Disease Mediates Spontaneous Calcium Signaling and Divergent Glutamatergic Calcium Responses.

Nitrosative stress is a feature of Alzheimer's disease (AD). Aims: We aimed to identify the cause underpinning increased nitric oxide (NO) in neurons and the impact of NO on neuronal function in AD. Results: We analyzed neuronal nitric oxide synthase (nNOS) protein levels in postmortem tissue and induced pluripotent stem cell (iPSC)-derived neurons from Alzheimer's patients and controls by immunohistochemistry and Western blots. Furthermore, we assessed the impact of modulating nNOS function or NO levels on neuronal glutamatergic signaling using calcium imaging. We show that nNOS protein levels are increased in early and severely affected brain regions of AD postmortem tissue, but not late and mildly affected regions, or cognitively normal individuals. The increased nNOS phenotype was also present in iPSC-derived neurons from late-onset Alzheimer's disease (LOAD) patients compared with controls, along with increased levels of nitrite, a stable marker of NO. Innovation: We observed a divergent functional impact of NO that included strengthening the calcium response in control neurons, while dysregulating calcium signaling and altering the amplitude and kinetics of the calcium responses to glutamate in the AD neurons. Pharmacological scavenging of NO or inhibition of nNOS prevented aberrant spontaneous calcium signaling in AD neurons. Conclusion: Together these data identify increases in nNOS protein in AD. Functional data suggest that NO modulation of glutamatergic calcium signaling is neuroprotective under nonpathogenic conditions, with increased nNOS and NO contributing to dysregulated spontaneous calcium signaling in AD neurons.

Enhanced fear memory after social defeat in mice is dependent on interleukin-1 receptor signaling in glutamatergic neurons.

Chronic stress is associated with increased anxiety, cognitive deficits, and post-traumatic stress disorder. Repeated social defeat (RSD) in mice causes long-term stress-sensitization associated with increased microglia activation, monocyte accumulation, and enhanced interleukin (IL)-1 signaling in endothelia and neurons. With stress-sensitization, mice have amplified neuronal, immune, and behavioral responses to acute stress 24 days later. This is clinically relevant as it shares key aspects with post-traumatic stress disorder. The mechanisms underlying stress-sensitization are unclear, but enhanced fear memory may be critical. The purpose of this study was to determine the influence of microglia and IL-1R1 signaling in neurons in the development of sensitization and increased fear memory after RSD. Here, RSD accelerated fear acquisition, delayed fear extinction, and increased cued-based freezing at 0.5 day. The enhancement in contextual fear memory after RSD persisted 24 days later. Next, microglia were depleted with a CSF1R antagonist prior to RSD and several parameters were assessed. Microglia depletion blocked monocyte recruitment to the brain. Nonetheless, neuronal reactivity (pCREB) and IL-1β RNA expression in the hippocampus and enhanced fear memory after RSD were microglial-independent. Because IL-1β RNA was prominent in the hippocampus after RSD even with microglia depletion, IL-1R1 mediated signaling in glutamatergic neurons was assessed using neuronal Vglut2+/IL-1R1-/- mice. RSD-induced neuronal reactivity (pCREB) in the hippocampus and enhancement in fear memory were dependent on neuronal IL-1R1 signaling. Furthermore, single-nuclei RNA sequencing (snRNAseq) showed that RSD influenced transcription in specific hippocampal neurons (DG neurons, CA2/3, CA1 neurons) associated with glutamate signaling, inflammation and synaptic plasticity, which were neuronal IL-1R1-dependent. Furthermore, snRNAseq data provided evidence that RSD increased CREB, BDNF, and calcium signaling in DG neurons in an IL-1R1-dependent manner. Collectively, increased IL-1R1-mediated signaling (monocytes/microglia independent) in glutamatergic neurons after RSD enhanced neuronal reactivity and fear memory.

Comparing tonic and phasic dendritic calcium in cholinergic pedunculopontine neurons and dopaminergic substantia nigra neurons.

Several brainstem nuclei degenerate in Parkinson's disease (PD). In addition to the well-characterized dopaminergic neurons of the substantia nigra pars compacta (SNc), the cholinergic neurons of the pedunculopontine nucleus (PPN) also degenerate in PD. One leading hypothesis of selective vulnerability is that pacemaking activity and the activation of low-threshold L-type calcium current are major contributors to tonic calcium load and cellular stress in SNc dopaminergic neurons. However, it is not yet clear whether the vulnerable PPN cholinergic neurons share this property. Therefore, we used two-photon dendritic calcium imaging and whole-cell electrophysiology to evaluate the role of L-type calcium channels in tonic and phasic dendritic calcium signals in PPN and SNc neurons. In addition, we investigated N- and P/Q-type calcium channel regulation of firing properties and dendritic calcium in PPN neurons. We found that blocking L-type channels reduces tonic firing rate and dendritic calcium levels in SNc neurons. By contrast, the tonic calcium load in PPN neurons did not depend on L-, N- or P/Q-type channels. However, we found that blocking either L-type (with nifedipine) or N- and P/Q-type (with omega-conotoxin MVIIC) channels reducesphasic calcium influx in PPN dendrites. Together, these findings show that L-type calcium channels play different roles in the activity of SNc and PPN neurons, and suggest that low-threshold L-type channels are not responsible for tonic calcium levels in PPN cholinergic neurons and are therefore not likely to be a source of selective vulnerability in these cells.

Altered neuronal group 1 metabotropic glutamate receptor- and endoplasmic reticulum-mediated Ca2+ signaling in two rodent models of Alzheimer’s disease

Calcium mobilization from the endoplasmic reticulum (ER) induced by, for example, IP3 receptor (IP3R) stimulation, and its subsequent crosstalk with extracellular Ca2+ influx mediated through voltage-gated calcium channels (VGCCs) and neuronal store-operated calcium entry (nSOCE), is essential for normal neuronal signaling and cellular homeostasis. However, several studies suggest that chronic calcium dysregulation may play a key role in the onset and/or progression of neurodegenerative conditions, particularly Alzheimer’s disease (AD). Here, using early postnatal hippocampal tissue from two transgenic murine models of AD, we provide further evidence that not only are crucial calcium signaling pathways dysregulated, but also that such dysregulation occurs at very early stages of development.Utilizing epifluorescence calcium imaging, we investigated ER-, nSOCE- and VGCC-mediated calcium signaling in cultured primary hippocampal neurons from two transgenic rodent models of AD: 3xTg-AD mice (PS1M146V/APPSWE/TauP301L) and TgF344-AD rats (APPSWE/PS1ΔE9) between 2 and 9 days old.Our results reveal that, in comparison to control hippocampal neurons, those from 3xTg-AD mice possessed significantly greater basal ER calcium levels, as measured by larger responses to I-mGluR-mediated ER Ca2+ mobilization (amplitude; 4 (0–19) vs 21(12–36) a.u., non-Tg vs 3xTg-AD; median difference (95 % Cl) = 14 a.u. (11–18); p = 0.004)) but reduced nSOCE (15 (4–22) vs 8(5–11) a.u., non-Tg vs 3xTg-AD; median difference (95 % Cl) = -7 a.u. (-3– −10 a.u.); p < 0.0001). Furthermore, unlike non-Tg neurons, where depolarization enhanced the amplitude, duration and area under the curve (A.U.C.) of I-mGluR-evoked ER-mediated calcium signals when compared with basal conditions, this was not apparent in 3xTg-AD neurons. Whilst the amplitude of depolarization-enhanced I-mGluR-evoked ER-mediated calcium signals from both non-Tg F344 and TgF344-AD neurons was significantly enhanced relative to basal conditions, the A.U.C. and duration of responses were enhanced significantly upon depolarization in non-Tg F344, but not in TgF344-AD, neurons. Overall, the nature of basal I-mGluR-mediated calcium responses did not differ significantly between non-Tg F344 and TgF344-AD neurons.In summary, our results characterizing ER- and nSOCE-mediated calcium signaling in neurons demonstrate that ER Ca2+ dyshomeostasis is an early and potentially pathogenic event in familial AD.

TRPM4 blocking antibody reduces neuronal excitotoxicity by specifically inhibiting glutamate-induced calcium influx under chronic hypoxia

Excitotoxicity arises from unusually excessive activation of excitatory amino acid receptors such as glutamate receptors. Following an energy crisis, excitotoxicity is a major cause for neuronal death in neurological disorders. Many glutamate antagonists have been examined for their efficacy in mitigating excitotoxicity, but failed to generate beneficial outcome due to their side effects on healthy neurons where glutamate receptors are also blocked. In this study, we found that during chronic hypoxia there is upregulation and activation of a nonselective cation channel TRPM4 that contributes to the depolarized neuronal membrane potential and enhanced glutamate-induced calcium entry. TRPM4 is involved in modulating neuronal membrane excitability and calcium signaling, with a complex and multifaceted role in the brain. Here, we inhibited TRPM4 using a newly developed blocking antibody M4P, which could repolarize the resting membrane potential and ameliorate calcium influx upon glutamate stimulation. Importantly, M4P did not affect the functions of healthy neurons as the activity of TRPM4 channel is not upregulated under normoxia. Using a rat model of chronic hypoxia with both common carotid arteries occluded, we found that M4P treatment could reduce apoptosis in the neurons within the hippocampus, attenuate long-term potentiation impairment and improve the functions of learning and memory in this rat model. With specificity to hypoxic neurons, TRPM4 blocking antibody can be a novel way of controlling excitotoxicity with minimal side effects that are common among direct blockers of glutamate receptors.

Comparison of the calcium signaling alterations in GABA-ergic medium spiny neurons produced from iPSCs of different origins

Disease models based on induced pluripotent stem cells (iPSCs) are in high demand because of their physiological adequacy and well-reproducibility of the pathological phenotype. Nowadays, the most common approach to generate iPSCs is the reprogramming of somatic cells using vectors based on lentivirus or Sendai virus. We have previously shown impairments of calcium signaling including store-operated calcium entry in Huntington's disease-specific iPSCs-based GABA-ergic medium spiny neurons. However, different approaches for iPSCs generation make it difficult to compare the models since the mechanism of reprogramming may influence the electrophysiological properties of the terminally differentiated neurons. Here, we have studied the features of calcium homeostasis in GABA-ergic medium spiny neurons differentiated from iPSCs obtained from fibroblasts of the same donor using different methods. Our data demonstrated that there were no significant differences neither in calcium influx through the store-operated channels, nor in the levels of proteins activating this type of calcium entry in neurons differentiated from iPSCs generated with lenti- and Sendai viruses-based approaches. We also found no differences in voltage-gated calcium entry for these neurons. Thus, we clearly showed that various methods of cell reprogramming result in similar deregulations in neuronal calcium signaling which substantiates the ability to combine the experimental data on functional studies of ion channels in models based on iPSCs obtained by different methods and expands the prospects for the use of biobanking.

IMPA1 dependent regulation of phosphatidylinositol 4,5-bisphosphate and calcium signalling by lithium.

Lithium (Li) is widely used as a mood stabilizer to treat bipolar affective disorder. However, the molecular targets of Li that underpin its therapeutic effect remain unresolved. Inositol monophosphatase (IMPA1) is an enzyme involved in phosphatidylinositol 4,5-bisphosphate (PIP2) resynthesis after PLC signaling. In vitro, Li inhibits IMPA1, but the relevance of this inhibition within neural cells remains unknown. Here, we report that treatment with therapeutic concentrations of Li reduces receptor-activated calcium release from intracellular stores and delays PIP2 resynthesis. These effects of Li are abrogated in IMPA1 deleted cells. We also observed that in human forebrain cortical neurons, treatment with Li reduced neuronal excitability and calcium signals. After Li treatment of human cortical neurons, transcriptome analyses revealed down-regulation of signaling by glutamate, a key excitatory neurotransmitter in the human brain. Collectively, our findings suggest that inhibition of IMPA1 by Li reduces receptor-activated PLC signaling and neuronal excitability.

Biocompatible neuronal stimulant-embedded poly(γ-benzyl-L-glutamate) peptide-based scaffolds promote differentiation, growth and functional maturation of human induced pluripotent stem cell-derived retinal ganglion cells

BackgroundTo culture human induced pluripotent stem cells(hiPSCs)- derived retinal ganglion cells (RGCs) with appropriate structure and functionality is critical for cell therapy of optic nerve degeneration. In this study, we aimed to assess the effectiveness of three biocompatible scaffolds -poly(γ-benzyl-L-glutamate) (PBG)-based three-dimensional fibrous scaffolds in promoting the structural and functional maturation of hiPSC-derived RGCs. MethodsThe study investigated three scaffolds (1) PBG, (2) PBG with glutamic acid (PBGA20), and (3) PBGA20 with sodium salt (PBGA20-Na). The biocompatibility was evaluated in vivo with C57BL/6 mice. Neurite outgrowth was assessed using immunostaining for axon markers, while fluorescence-based Ca2+ imaging and patch-clamp technique were employed to measure neuronal calcium signaling and electrophysiological activity, respectively. Significant findingsAll three PBG-based scaffolds exhibited excellent biocompatibility in vivo and effectively supported the directional outgrowth of neurites from hiPSC-derived RGCs while the PBGA20-Na group demonstrated the most favorable outcomes. The presence of glutamic acid in PBGA20 and PBGA20-Na scaffolds led to an increase in calcium signaling within hiPSC-derived RGCs. HiPSC-derived RGCs cultured on PBGA20 and PBGA20-Na scaffolds exhibited an earlier onset of electrophysiological activities with lower thresholds. These promising results provide a foundation for potential sheet transplantation of hiPSC-derived RGCs as cell therapy for optic nerve regeneration.

NeuroActivityToolkit-Toolbox for Quantitative Analysis of Miniature Fluorescent Microscopy Data.

The visualization of neuronal activity in vivo is an urgent task in modern neuroscience. It allows neurobiologists to obtain a large amount of information about neuronal network architecture and connections between neurons. The miniscope technique might help to determine changes that occurred in the network due to external stimuli and various conditions: processes of learning, stress, epileptic seizures and neurodegenerative diseases. Furthermore, using the miniscope method, functional changes in the early stages of such disorders could be detected. The miniscope has become a modern approach for recording hundreds to thousands of neurons simultaneously in a certain brain area of a freely behaving animal. Nevertheless, the analysis and interpretation of the large recorded data is still a nontrivial task. There are a few well-working algorithms for miniscope data preprocessing and calcium trace extraction. However, software for further high-level quantitative analysis of neuronal calcium signals is not publicly available. NeuroActivityToolkit is a toolbox that provides diverse statistical metrics calculation, reflecting the neuronal network properties such as the number of neuronal activations per minute, amount of simultaneously co-active neurons, etc. In addition, the module for analyzing neuronal pairwise correlations is implemented. Moreover, one can visualize and characterize neuronal network states and detect changes in 2D coordinates using PCA analysis. This toolbox, which is deposited in a public software repository, is accompanied by a detailed tutorial and is highly valuable for the statistical interpretation of miniscope data in a wide range of experimental tasks.