Ion Channels and Ionotropic Receptors in Astrocytes: Physiological Functions and Alterations in Alzheimer's Disease and Glioblastoma.

The human brain is composed of nearly one hundred billion neurons and an equal number of glial cells, including macroglia, i.e., astrocytes and oligodendrocytes, and microglia, the resident immune cells of the brain. In the last few decades, compelling evidence has revealed that glial cells are far more active and complex than previously thought. In particular, astrocytes, the most abundant glial cell population, not only take part in brain development, metabolism, and defense against pathogens and insults, but they also affect sensory, motor, and cognitive functions by constantly modulating synaptic activity. Not surprisingly, astrocytes are actively involved in neurodegenerative diseases (NDs) and other neurological disorders like brain tumors, in which they rapidly become reactive and mediate neuroinflammation. Reactive astrocytes acquire or lose specific functions that differently modulate disease progression and symptoms, including cognitive impairments. Astrocytes express several types of ion channels, including K+, Na+, and Ca2+ channels, transient receptor potential channels (TRP), aquaporins, mechanoreceptors, and anion channels, whose properties and functions are only partially understood, particularly in small processes that contact synapses. In addition, astrocytes express ionotropic receptors for several neurotransmitters. Here, we provide an extensive and up-to-date review of the roles of ion channels and ionotropic receptors in astrocyte physiology and pathology. As examples of two different brain pathologies, we focus on Alzheimer's disease (AD), one of the most diffuse neurodegenerative disorders, and glioblastoma (GBM), the most common brain tumor. Understanding how ion channels and ionotropic receptors in astrocytes participate in NDs and tumors is necessary for developing new therapeutic tools for these increasingly common neurological conditions.

Accurate Multiscale Simulation of Frictional Interfaces by Quantum Mechanics/Green's Function Molecular Dynamics.

Understanding frictional phenomena is a fascinating fundamental problem with huge potential impact on energy saving. Such an understanding requires monitoring what happens at the sliding buried interface, which is almost inaccessible by experiments. Simulations represent powerful tools in this context, yet a methodological step forward is needed to fully capture the multiscale nature of the frictional phenomena. Here, we present a multiscale approach based on linked ab initio and Green's function molecular dynamics, which is above the state-of-the-art techniques used in computational tribology as it allows for a realistic description of both the interfacial chemistry and energy dissipation due to bulk phonons in nonequilibrium conditions. By considering a technologically relevant system composed of two diamond surfaces with different degrees of passivation, we show that the presented method can be used not only for monitoring in real-time tribolochemical phenomena such as the tribologically induced surface graphitization and passivation effects but also for estimating realistic friction coefficients. This opens the way to in silico experiments of tribology to test materials to reduce friction prior to that in real labs.

TribChem: A Software for the First-Principles, High-Throughput Study of Solid Interfaces and Their Tribological Properties.

High-throughput first-principles calculations, based on solving the quantum mechanical many-body problem for hundreds of materials in parallel, have been successfully applied to advance many materials-based technologies, from batteries to hydrogen storage. However, this approach has not yet been adopted to systematically study solid-solid interfaces and their tribological properties. To this aim, we developed TribChem, an advanced software program based on the FireWorks platform, which is here presented and released. TribChem is constructed in a modular way, allowing for the separate calculation of bulk, surface, and interface properties. At present, the calculated interfacial properties include adhesion, shear strength, and charge redistribution. Further properties can be easily added due to the general structure of the main workflow. TribChem contains a high-level interface class to store/retrieve results from its own database and connect to public databases.

High-Throughput First-Principles Prediction of Interfacial Adhesion Energies in Metal-on-Metal Contacts.

Adhesion energy, a measure of the strength by which two surfaces bind together, ultimately dictates the mechanical behavior and failure of interfaces. As natural and artificial solid interfaces are ubiquitous, adhesion energy represents a key quantity in a variety of fields ranging from geology to nanotechnology. Because of intrinsic difficulties in the simulation of systems where two different lattices are matched, and despite their importance, no systematic, accurate first-principles determination of heterostructure adhesion energy is available. We have developed robust, automatic high-throughput workflow able to fill this gap by systematically searching for the optimal interface geometry and accurately determining adhesion energies. We apply it here for the first time to perform the screening of around a hundred metallic heterostructures relevant for technological applications. This allows us to populate a database of accurate values, which can be used as input parameters for macroscopic models. Moreover, it allows us to benchmark commonly used, empirical relations that link adhesion energies to the surface energies of its constituent and to improve their predictivity employing only quantities that are easily measurable or computable.

Modulation of NMDA receptor activity by CR4056, an imidazoline-2 receptor ligand with analgesic properties.

CR4056 is an imidazoline-2 receptor ligand having potent analgesic activity and synergistic effect with opioids. Very recently it has been found that CR4056 can revert the cognitive impairment in animal models of Alzheimer's disease (AD). Since several lines of evidence highlight the importance of NMDAR modulators in nociceptive signaling and in AD progression, we considered as important to investigate the effects of CR4056 on NMDAR activity. In primary culture of cortical neurons, application of NMDA and glycine elicits a current that is decreased in a dose-dependent fashion by CR4056 (IC50 5.3 ± 0.1 µM). CR4056 antagonism is reversible, not competitive and voltage-independent and it is not blocked by pertussis toxin. CR4056 interacts with the co-agonist glycine site in a competitive way, indeed high glycine concentrations diminish its effect. Fibroblasts expressing different recombinant NMDA receptors are differently modulated by CR4056: the potency and the efficacy of the compound are higher in GluN1- GluN2B than in GluN1-GluN2A containing receptors. In lamina II neurons of spinal cord slices, single stimulation of afferent fibers evokes an NMDA-mediated current that is inhibited by 10 µM CR4056. Repetitive stimulation of the dorsal root at high frequency and high intensity produces a firing activity that is significatively depressed by CR4056. Taken together, our results broad the understanding of the molecular mechanisms of CR4056 analgesic activity, involving the modulation of NMDAR activity. Therefore, we propose that the analgesic action of CR4056 and the neuroprotective effects in AD models may be mediated also by NMDAR inhibition.

Astrocytes mediate long-lasting synaptic regulation of ventral tegmental area dopamine neurons.

The plasticity of glutamatergic transmission in the ventral tegmental area (VTA) represents a fundamental mechanism in the modulation of dopamine neuron burst firing and phasic dopamine release at target regions. These processes encode basic behavioral responses, including locomotor activity, learning and motivated behaviors. Here we describe a hitherto unidentified mechanism of long-term synaptic plasticity in mouse VTA. We found that the burst firing in individual dopamine neurons induces a long-lasting potentiation of excitatory synapses on adjacent dopamine neurons that crucially depends on Ca2+ elevations in astrocytes, mediated by endocannabinoid CB1 and dopamine D2 receptors co-localized at the same astrocytic process, and activation of pre-synaptic metabotropic glutamate receptors. Consistent with these findings, selective in vivo activation of astrocytes increases the burst firing of dopamine neurons in the VTA and induces locomotor hyperactivity. Astrocytes play, therefore, a key role in the modulation of VTA dopamine neuron functional activity.

Corrigendum: GABA tonic currents and glial cells are altered during epileptogenesis in a mouse model of Dravet syndrome.

[This corrects the article DOI: 10.3389/fncel.2022.919493.].

GABA tonic currents and glial cells are altered during epileptogenesis in a mouse model of Dravet syndrome.

Dravet Syndrome (DS) is a rare autosomic encephalopathy with epilepsy linked to Nav1.1 channel mutations and defective GABAergic signaling. Effective therapies for this syndrome are lacking, urging a better comprehension of the mechanisms involved. In a recognized mouse model of DS, we studied GABA tonic current, a form of inhibition largely neglected in DS, in brain slices from developing mice before spontaneous seizures are reported. In neurons from the temporal cortex (TeCx) and CA1 region, GABA tonic current was reduced in DS mice compared to controls, while in the entorhinal cortex (ECx) it was not affected. In this region however allopregnanonole potentiation of GABA tonic current was reduced in DS mice, suggesting altered extrasynaptic GABAA subunits. Using THIP as a selective agonist, we found reduced δ subunit mediated tonic currents in ECx of DS mice. Unexpectedly in the dentate gyrus (DG), a region with high δ subunit expression, THIP-evoked currents in DS mice were larger than in controls. An immunofluorescence study confirmed that δ subunit expression was reduced in ECx and increased in DG of DS mice. Finally, considering the importance of neuroinflammation in epilepsy and neurodevelopmental disorders, we evaluated classical markers of glia activation. Our results show that DS mice have increased Iba1 reactivity and GFAP expression in both ECx and DG, compared to controls. Altogether we report that before spontaneous seizures, DS mice develop significant alterations of GABA tonic currents and glial cell activation. Understanding all the mechanisms involved in these alterations during disease maturation and progression may unveil new therapeutic targets.

Dysbindin-1A modulation of astrocytic dopamine and basal ganglia dependent behaviors relevant to schizophrenia.

The mechanisms underlying the dichotomic cortical/basal ganglia dopaminergic abnormalities in schizophrenia are unclear. Astrocytes are important non-neuronal modulators of brain circuits, but their role in dopaminergic system remains poorly explored. Microarray analyses, immunohistochemistry, and two-photon laser scanning microscopy revealed that Dys1 hypofunction increases the reactivity of astrocytes, which express only the Dys1A isoform. Notably, behavioral and electrochemical assessments in mice selectively lacking the Dys1A isoform unraveled a more prominent impact of Dys1A in behavioral and dopaminergic/D2 alterations related to basal ganglia, but not cortical functioning. Ex vivo electron microscopy and protein expression analyses indicated that selective Dys1A disruption might alter intracellular trafficking in astrocytes, but not in neurons. In agreement, Dys1A disruption only in astrocytes resulted in decreased motivation and sensorimotor gating deficits, increased astrocytic dopamine D2 receptors and decreased dopaminergic tone within basal ganglia. These processes might have clinical relevance because the caudate, but not the cortex, of patients with schizophrenia shows a reduction of the Dys1A isoform. Therefore, we started to show a hitherto unknown role for the Dys1A isoform in astrocytic-related modulation of basal ganglia behavioral and dopaminergic phenotypes, with relevance to schizophrenia.

Calcium Signals in Astrocyte Microdomains, a Decade of Great Advances.

The glial cells astrocytes have long been recognized as important neuron-supporting elements in brain development, homeostasis, and metabolism. After the discovery that the reciprocal communication between astrocytes and neurons is a fundamental mechanism in the modulation of neuronal synaptic communication, over the last two decades astrocytes became a hot topic in neuroscience research. Crucial to their functional interactions with neurons are the cytosolic Ca2+ elevations that mediate gliotransmission. Large attention has been posed to the so-called Ca2+microdomains, dynamic Ca2+ changes spatially restricted to fine astrocytic processes including perisynaptic astrocytic processes (PAPs). With presynaptic terminals and postsynaptic neuronal membranes, PAPs compose the tripartite synapse. The distinct spatial-temporal features and functional roles of astrocyte microdomain Ca2+ activity remain poorly defined. However, thanks to the development of genetically encoded Ca2+ indicators (GECIs), advanced microscopy techniques, and innovative analytical approaches, Ca2+ transients in astrocyte microdomains were recently studied in unprecedented detail. These events have been observed to occur much more frequently (∼50-100-fold) and dynamically than somatic Ca2+ elevations with mechanisms that likely involve both IP3-dependent and -independent pathways. Further progress aimed to clarify the complex, dynamic machinery responsible for astrocytic Ca2+ activity at microdomains is a crucial step in our understanding of the astrocyte role in brain function and may also reveal astrocytes as novel therapeutic targets for different brain diseases. Here, we review the most recent studies that improve our mechanistic understanding of the essential features of astrocyte Ca2+ microdomains.

Experimental and Ab Initio Characterization of Mononuclear Molybdenum Dithiocarbamates in Lubricant Mixtures.

Molybdenum dithiocarbamates (MoDTCs) are a class of lubricant additives widely employed in automotives. Most of the studies concerning MoDTC take into account the dimeric structures because of their industrial relevance, with the mononuclear compounds usually neglected, because isolating and characterizing subgroups of MoDTC molecules are generally difficult. However, the byproducts of the synthesis of MoDTC can impact the friction reduction performance at metallic interfaces, and the effect of mononuclear MoDTC (mMoDTC) compounds in the lubrication has not been considered yet in the literature. In this study, we consider for the first time the impurities of MoDTC consisting of mononuclear compounds and combine experimental and computational techniques to elucidate the interaction of these impurities with binuclear MoDTC in commercial formulations. We present a preliminary strategy to separate a commercial MoDTC product in chemically different fractions. These fractions present different tribological behaviors depending on the relative amount of mononuclear and binuclear complexes. The calculations indicate that the dissociation mechanism of mMoDTC is similar to the one observed for the dimeric structures. However, the different chemical properties of mMoDTC impact the kinetics for the formation of the beneficial molybdenum disulfide (MoS2) layers, as shown by the tribological experiments. These results help to understand the functionality of MoDTC lubricant additives, providing new insights into the complex synergy between the different chemical structures.

Altered Cl- homeostasis hinders forebrain GABAergic interneuron migration in a mouse model of intellectual disability.

Impairments of inhibitory circuits are at the basis of most, if not all, cognitive deficits. The impact of OPHN1, a gene associate with intellectual disability (ID), on inhibitory neurons remains elusive. We addressed this issue by analyzing the postnatal migration of inhibitory interneurons derived from the subventricular zone in a validated mouse model of ID (OPHN1-/y mice). We found that the speed and directionality of migrating neuroblasts were deeply perturbed in OPHN1-/y mice. The significant reduction in speed was due to altered chloride (Cl-) homeostasis, while the overactivation of the OPHN1 downstream signaling pathway, RhoA kinase (ROCK), caused abnormalities in the directionality of the neuroblast progression in mutants. Blocking the cation-Cl- cotransporter KCC2 almost completely rescued the migration speed while proper directionality was restored upon ROCK inhibition. Our data unveil a strong impact of OPHN1 on GABAergic inhibitory interneurons and identify putative targets for successful therapeutic approaches.

Cellular and molecular mechanisms of new onset seizure generation.

New onset epilepsy and seizures are common neurological disorders in aged people, second only to stroke and dementia. They are frequently related to other pathological conditions including stroke, trauma, tumors and neurological diseases whereas in about one-third of cases the origin is unknown. Besides the origin, the cellular and molecular events that suddenly trigger seizures are poorly defined. Using an acute model of seizure generation that better resembles new onset seizures, we studied GABAergic interneurons and astrocytes during seizure generation. We found that seizures are preceded by a GABAergic rhythmic hyperactivity that synchronizes pyramidal neurons by inducing a rebound spiking that favors seizures' onset. Furthermore, the intense activity in GABAergic interneurons evokes Ca2+ elevations in astrocytes that, by releasing glutamate, further excite neuronal network. Elucidating the cellular and molecular events that generate seizures may reveal new targets for treatment of new onset seizures and epilepsy.

High-throughput screening of the static friction and ideal cleavage strength of solid interfaces.

We present a comprehensive ab initio, high-throughput study of the frictional and cleavage strengths of interfaces of elemental crystals with different orientations. It is based on the detailed analysis of the adhesion energy as a function of lateral, γ(x, y), and perpendicular displacements, γ(z), with respect to the considered interface plane. We use the large amount of computed data to derive fundamental insight into the relation of the ideal strength of an interface plane with its adhesion. Moreover, the ratio between the frictional and cleavage strengths is provided as good indicator for the material failure mode - dislocation propagation versus crack nucleation. All raw and curated data are made available to be used as input parameters for continuum mechanic models, benchmarks, or further analysis.

Correction to: Optogenetic Interneuron Stimulation and Calcium Imaging in Astrocytes.

Figures 1 and 2 were inadvertently switched during the production and this has been corrected so the figures appear in the proper order.

Optogenetic Interneuron Stimulation and Calcium Imaging in Astrocytes.

In brain networks, neurons are constantly involved in a dynamic interaction with the other cell populations and, particularly, with astrocytes, the most abundant glial cells in the brain. Astrocytes respond to neurotransmitters with Ca2+ elevations which represent a key event in the modulation of local brain circuits played by these glial cells. Due to technical limitations, the study of Ca2+ signal dynamics in astrocytes has focused for decades almost exclusively on somatic and perisomatic regions. Accordingly, Ca2+ signal in astrocytic fine protrusions, which are in close contact with the synapse, has been poorly investigated. Over the last years, the diffusion of novel tools such as the viral vector gene delivery of genetically encoded Ca2+ indicators (GECI), the optogenetics, and multiphoton laser scanning microscopy has boosted significantly our capability to study astrocytic Ca2+ signals in the different subcellular compartments. Here we report a protocol that combines these techniques to study astrocyte Ca2+ signaling in response to somatostatin (SST)-expressing interneurons, one of the main classes of GABAergic inhibitory interneurons.

Interneuron-specific signaling evokes distinctive somatostatin-mediated responses in adult cortical astrocytes.

The signaling diversity of GABAergic interneurons to post-synaptic neurons is crucial to generate the functional heterogeneity that characterizes brain circuits. Whether this diversity applies to other brain cells, such as the glial cells astrocytes, remains unexplored. Using optogenetics and two-photon functional imaging in the adult mouse neocortex, we here reveal that parvalbumin- and somatostatin-expressing interneurons, two key interneuron classes in the brain, differentially signal to astrocytes inducing weak and robust GABAB receptor-mediated Ca2+ elevations, respectively. Furthermore, the astrocyte response depresses upon parvalbumin interneuron repetitive stimulations and potentiates upon somatostatin interneuron repetitive stimulations, revealing a distinguished astrocyte plasticity. Remarkably, the potentiated response crucially depends on the neuropeptide somatostatin, released by somatostatin interneurons, which activates somatostatin receptors at astrocytic processes. Our study unveils, in the living brain, a hitherto unidentified signaling specificity between interneuron subtypes and astrocytes opening a new perspective into the role of astrocytes as non-neuronal components of inhibitory circuits.

Interneuronal Network Activity at the Onset of Seizure-Like Events in Entorhinal Cortex Slices.

The onset of focal seizures in humans and in different animal models of focal epilepsy correlates with reduction of neuronal firing and enhanced interneuronal network activity. Whether this phenomenon contributes to seizure generation is still unclear. We used the in vitro entorhinal cortex slices bathed in 4-aminopirydine (4-AP) as an experimental paradigm model to evaluate the correlation between interneuronal GABAergic network activity and seizure-like events. Epileptiform discharges were recorded in layer V-VI pyramidal neurons and fast-spiking interneurons in slices from male and female mice and in the isolated female guinea pig brain preparation during perfusion with 4-AP. We observed that 90% of seizure-like events recorded in principal cells were preceded by outward currents coupled with extracellular potassium shifts, abolished by pharmacological blockade of GABAA receptors. Potassium elevations associated to GABAA receptor-mediated population events were confirmed in the entorhinal cortex of the in vitro isolated whole guinea pig brain. Fast-rising and sustained extracellular potassium increases associated to interneuronal network activity consistently preceded the initiation of seizure-like events. We conclude that in the 4-AP seizure model, interneuronal network activity occurs before 4-AP-induced seizures and therefore supports a role of interneuron activity in focal seizure generation.SIGNIFICANCE STATEMENT The paper focuses on the mechanisms of ictogenesis, a topic that requires a step beyond the simplistic view that seizures, and epilepsy, are due to an increase of excitatory network activity. Focal temporal lobe seizures in humans and in several experimental epilepsies likely correlate with a prevalent activation of interneurons. The potassium channel blocker 4-aminopyridine reliably induces seizure-like events in temporal lobe structures. Herein, we show that a majority of seizures in the entorhinal cortex starts with interneuronal network activity accompanied by a fast and sustained increase in extracellular potassium. Our new findings reinforce and add a new piece of evidence to the proposal that limbic seizures can be supported by GABAergic hyperactivity.

New Tools to Study Astrocyte Ca2+ Signal Dynamics in Brain Networks In Vivo.

Sensory information processing is a fundamental operation in the brain that is based on dynamic interactions between different neuronal populations. Astrocytes, a type of glial cells, have been proposed to represent active elements of brain microcircuits that, through dynamic interactions with neurons, provide a modulatory control of neuronal network activity. Specifically, astrocytes in different brain regions have been described to respond to neuronal signals with intracellular Ca2+ elevations that represent a key step in the functional recruitment of astrocytes to specific brain circuits. Accumulating evidence shows that Ca2+ elevations regulate the release of gliotransmitters that, in turn, modulate synaptic transmission and neuronal excitability. Recent studies also provided new insights into the spatial and temporal features of astrocytic Ca2+ elevations revealing a surprising complexity of Ca2+ signal dynamics in astrocytes. Here we discuss how recently developed experimental tools such as the genetically encoded Ca2+ indicators (GECI), optogenetics and chemogenetics can be applied to the study of astrocytic Ca2+ signals in the living brain.