δ subunit-containing GABAA IPSCs are driven by both synaptic and diffusional GABA in mouse dentate granule neurons.

KEY POINTS: Current views suggest γ2 subunit-containing GABAA receptors mediate phasic IPSCs while extrasynaptic δ subunits mediate diffusional IPSCs and tonic current. We have re-examined the roles of the two receptor populations using mice with picrotoxin resistance engineered into receptors containing the δ subunit. Using pharmacological separation, we find that in general δ and γ IPSCs are modulated in parallel by manipulations of transmitter output and diffusion, with evidence favouring modestly more diffusional contribution to δ IPSCs. Our findings also reveal that spontaneous δ IPSCs are mainly driven by channel deactivation, rather than by diffusion of GABA. Understanding the functional contributions of the two receptor classes may help us understand the actions of drug therapies with selective effects on one population over the other. ABSTRACT: GABAA receptors mediate transmission throughout the central nervous system and typically contain a δ subunit (δ receptors) or a γ2 subunit (γ2 receptors). δ IPSCs decay slower than γ2 IPSCs, but the reasons are unclear. Transmitter diffusion, rebinding, or slow deactivation kinetics of channels are candidates. We used gene editing to confer picrotoxin resistance on δ receptors in mice, then pharmacologically isolated δ receptors in mouse dentate granule cells to explore IPSCs. γ2 and δ components of IPSCs were modulated similarly by presynaptic manipulations and manipulations of transmitter lifetime, suggesting that GABA release recruits δ receptors proportionally to γ2 receptors. δ IPSCs showed more sensitivity to altered transmitter release and to a rapidly dissociating antagonist, suggesting an additional spillover contribution. Reducing GABA diffusion with 5% dextran increased the peak amplitude and decreased the decay of evoked δ IPSCs but had no effect on δ or dual-component (mainly γ2-driven) spontaneous IPSCs, suggesting that GABA actions can be local for both receptor types. Rapid application of varied [GABA] onto nucleated patches from dentate granule cells demonstrated a deactivation rate of δ receptors similar to that of δ spontaneous IPSCs, consistent with the idea that deactivation and local GABA actions drive δ spontaneous IPSCs. Overall, our results indicate that δ IPSCs are activated by both synaptic and diffusional GABA. Our results are consistent with a functional relationship between δ and γ2 GABAA receptors akin to that of slow NMDA and fast AMPA EPSCs at glutamate synapses.

Chemogenetic Isolation Reveals Synaptic Contribution of δ GABAA Receptors in Mouse Dentate Granule Neurons.

Two major GABAA receptor classes mediate ionotropic GABA signaling, those containing a δ subunit and those with a γ2 subunit. The classical viewpoint equates γ2-containing receptors with IPSCs and δ-containing receptors with tonic inhibition because of differences in receptor localization, but significant questions remain because the populations cannot be pharmacologically separated. We removed this barrier using gene editing to confer a point mutation on the δ subunit in mice, rendering receptors containing the subunit picrotoxin resistant. By pharmacologically isolating δ-containing receptors, our results demonstrate their contribution to IPSCs in dentate granule neurons and weaker contributions to thalamocortical IPSCs. Despite documented extrasynaptic localization, we found that receptor localization does not preclude participation in isolated IPSCs, including mIPSCs. Further, phasic inhibition from δ subunit-containing receptors strongly inhibited summation of EPSPs, whereas tonic activity had little impact. In addition to any role that δ-containing receptors may play in canonical tonic inhibition, our results highlight a previously underestimated contribution of δ-containing receptors to phasic inhibition.SIGNIFICANCE STATEMENT GABAA receptors play key roles in transient and tonic inhibition. The prevailing view suggests that synaptic γ2-containing GABAA receptors drive phasic inhibition, whereas extrasynaptic δ-containing receptors mediate tonic inhibition. To re-evaluate the impact of δ receptors, we took a chemogenetic approach that offers a sensitive method to probe the synaptic contribution of δ-containing receptors. Our results reveal that localization does not strongly limit the contribution of δ receptors to IPSCs and that δ receptors make an unanticipated robust contribution to phasic inhibition.

Contributions of space-clamp errors to apparent time-dependent loss of Mg2+ block induced by NMDA.

N-methyl-d-aspartate receptors (NMDARs) govern synaptic plasticity, development, and neuronal response to insult. Prolonged activation of NMDARs such as during an insult may activate secondary currents or modulate Mg2+ sensitivity, but the conditions under which these occur are not fully defined. We reexamined the effect of prolonged NMDAR activation in juvenile mouse hippocampal slices. NMDA (10 µM) elicited current with the expected negative-slope conductance in the presence of 1.2 mM Mg2+ However, several minutes of continued NMDA exposure elicited additional inward current at -70 mV. A higher concentration of NMDA (100 µM) elicited the current more rapidly. The additional current was not dependent on Ca2+, network activity, or metabotropic NMDAR function and did not persist on agonist removal. Voltage ramps revealed no alteration of either reversal potential or NMDA-elicited conductance between -30 mV and +50 mV. The result was a more linear NMDA current-voltage relationship. The current linearization was also induced in interneurons and in mature dentate granule neurons but not immature dentate granule cells, dissociated cultured hippocampal neurons, or nucleated patches excised from CA1 pyramidal neurons. Comparative simulations of NMDA application to a CA1 pyramidal neuron and to a cultured neuron revealed that linearization can be explained by space-clamp errors arising from gradual recruitment of distal dendritic NMDARs. We conclude that persistent secondary currents do not strongly contribute to NMDAR responses in juvenile mouse hippocampus and careful discernment is needed to exclude contributions of clamp artifacts to apparent secondary currents.NEW & NOTEWORTHY We report that upon sustained activation of NMDARs in juvenile mouse hippocampal neurons there is apparent loss of Mg2+ block at negative membrane potentials. However, the phenomenon is explained by loss of dendritic voltage clamp, leading to a linear current-voltage relationship. Our results give a specific example of how spatial voltage errors in voltage-clamp recordings can readily be misinterpreted as biological modulation.

Endogenous 24S-hydroxycholesterol modulates NMDAR-mediated function in hippocampal slices.

N-methyl-D-aspartate receptors (NMDARs), a major subtype of glutamate receptors mediating excitatory transmission throughout the central nervous system (CNS), play critical roles in governing brain function and cognition. Because NMDAR dysfunction contributes to the etiology of neurological and psychiatric disorders including stroke and schizophrenia, NMDAR modulators are potential drug candidates. Our group recently demonstrated that the major brain cholesterol metabolite, 24S-hydroxycholesterol (24S-HC), positively modulates NMDARs when exogenously administered. Here, we studied whether endogenous 24S-HC regulates NMDAR activity in hippocampal slices. In CYP46A1(-/-) (knockout; KO) slices where endogenous 24S-HC is greatly reduced, NMDAR tone, measured as NMDAR-to-α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor (AMPAR) excitatory postsynaptic current (EPSC) ratio, was reduced. This difference translated into more NMDAR-driven spiking in wild-type (WT) slices compared with KO slices. Application of SGE-301, a 24S-HC analog, had comparable potentiating effects on NMDAR EPSCs in both WT and KO slices, suggesting that endogenous 24S-HC does not saturate its NMDAR modulatory site in ex vivo slices. KO slices did not differ from WT slices in either spontaneous neurotransmission or in neuronal intrinsic excitability, and exhibited LTP indistinguishable from WT slices. However, KO slices exhibited higher resistance to persistent NMDAR-dependent depression of synaptic transmission induced by oxygen-glucose deprivation (OGD), an effect restored by SGE-301. Together, our results suggest that loss of positive NMDAR tone does not elicit compensatory changes in excitability or transmission, but it protects transmission against NMDAR-mediated dysfunction. We expect that manipulating this endogenous NMDAR modulator may offer new treatment strategies for neuropsychiatric dysfunction.

Cofilin under control of ß-arrestin-2 in NMDA-dependent dendritic spine plasticity, long-term depression (LTD), and learning.

Dendritic spines are dynamic, actin-rich structures that form the postsynaptic sites of most excitatory synapses in the brain. The F-actin severing protein cofilin has been implicated in the remodeling of dendritic spines and synapses under normal and pathological conditions, by yet unknown mechanisms. Here we report that ß-arrestin-2 plays an important role in NMDA-induced remodeling of dendritic spines and synapses via translocation of active cofilin to dendritic spines. NMDAR activation triggers cofilin activation through calcineurin and phosphatidylinositol 3-kinase (PI3K)-mediated dephosphorylation and promotes cofilin translocation to dendritic spines that is mediated by ß-arrestin-2. Hippocampal neurons lacking ß-arrestin-2 develop mature spines that fail to remodel in response to NMDA. ß-Arrestin-2-deficient mice exhibit normal hippocampal long-term potentiation, but significantly impaired NMDA-dependent long-term depression and spatial learning deficits. Moreover, ß-arrestin-2-deficient hippocampal neurons are resistant to Aß-induced dendritic spine loss. Our studies demonstrate unique functions of ß-arrestin-2 in NMDAR-mediated dendritic spine and synapse plasticity through spatial control over cofilin activation.

Hyposmolality differentially and spatiotemporally modulates levels of glutamine synthetase and serine racemase in rat supraoptic nucleus.

Prolonged hyposmotic challenge (HOC) has a dual effect on vasopressin (VP) secretion [Yagil and Sladek (1990) Am J Physiol 258(2 Pt 2):R492-R500]. We describe an electrophysiological correlate of this phenomenon, whereby in vitro HOC transiently reduced the firing activity of VP neurons within the supraoptic nucleus of brain slices, which was followed by a rebound increase of their activity; this was paralleled by changes in the level of proteins relevant to astroglia-neuronal interactions. Hence, in vitro HOC transiently (at 5 min) increased the level of astrocyte-specific glial fibrillary acidic protein (GFAP), which then declined to control or base level (at 20 min); this was blocked by the gliotoxin L-aminoadipic acid, but not by tetanus toxin, which was used to inhibit neurotransmission. Similarly, in vivo HOC led to changes in GFAP level, which after an early increase (10 min) returned to normal (30 min). Immunoassays revealed that neuronal, but not astrocytic, expression of serine racemase (SR) was increased at the late stage of HOC in vivo, whereas at an early stage there was a transient increase in level of the astrocyte-specific glutamine synthetase (GS). Furthermore, there was an increased molecular association between GFAP and GS at 10 min, whereas SR increased its association with the neuronal nuclear antigen NeuN at 30 min. These results suggest that the dual effect of HOC on VP neuronal secretion/activity could be related to metabolic/signaling changes in astrocytes (glutamate-glutamine conversion) and neurons (D-serine synthesis/ammonia production), which may account for the rebound in VP neuronal activity, presumably by promoting the activation of neuronal glutamate receptors.

Astrocytic group I mGluR-dependent potentiation of astrocytic glutamate and potassium uptake.

One of the most important functions of astrocytes is removal of glutamate released during synaptic transmission. Surprisingly, the mechanisms by which astrocyte glutamate uptake is acutely modulated remain to be clarified. Astrocytes express metabotropic glutamate receptors (mGluRs) and other G protein-coupled receptors (GPCRs), which are activated during neuronal activity. Here, we test the hypothesis that astrocytic group I mGluRs acutely regulate glutamate uptake by astrocytes in situ. This hypothesis was tested in acute mouse hippocampal slices. Activation of astrocytic mGluRs, using a tetanic high-frequency stimulus (HFS) applied to Schaffer collaterals, led to potentiation of the amplitude of the synaptically evoked glutamate transporter currents (STCs) and associated charge transfer without changes in kinetics. Similar potentiation of STCs was not observed in the presence of group I mGluR antagonists or the PKC inhibitor, PKC 19-36, suggesting that HFS-induced potentiation of astrocyte glutamate uptake is astrocytic group I mGluR and PKC dependent. Pharmacological stimulation of a transgenic GPCR (MrgA1R), expressed exclusively in astrocytes, also potentiated STC amplitude and charge transfer, albeit quicker and shorter lasting compared with HFS-induced potentiation. The amplitude of the slow, inward astrocytic current due to potassium (K(+)) influx was also enhanced following activation of the endogenous mGluRs or the astrocyte-specific MrgA1 Gq GPCRs. Taken together, these findings suggest that astrocytic group I mGluR activation has a synergistic, modulatory effect on the uptake of glutamate and K(+).

GABAergic inhibition through synergistic astrocytic neuronal interaction transiently decreases vasopressin neuronal activity during hypoosmotic challenge.

The neuropeptide vasopressin is crucial to mammalian osmotic regulation. Local hypoosmotic challenge transiently decreases and then increases vasopressin secretion. To investigate mechanisms underlying this transient response, we examined the effects of hypoosmotic challenge on the electrical activity of rat hypothalamic supraoptic nucleus (SON) vasopressin neurons using patch-clamp recordings. We found that 5 min exposure of hypothalamic slices to hypoosmotic solution transiently increased inhibitory postsynaptic current (IPSC) frequency and reduced the firing rate of vasopressin neurons. Recovery occurred by 10 min of exposure, even though the osmolality remained low. The γ-aminobutyric acid (GABA)A receptor blocker, gabazine, blocked the IPSCs and the hypoosmotic suppression of firing. The gliotoxin l-aminoadipic acid blocked the increase in IPSC frequency at 5 min and the recovery of firing at 10 min, indicating astrocytic involvement in hypoosmotic modulation of vasopressin neuronal activity. Moreover, ß-alanine, an osmolyte of astrocytes and GABA transporter (GAT) inhibitor, blocked the increase in IPSC frequency at 5 min of hypoosmotic challenge. Confocal microscopy of immunostained SON sections revealed that astrocytes and magnocellular neurons both showed positive staining of vesicular GATs (VGAT). Hypoosmotic stimulation in vivo reduced the number of VGAT-expressing neurons, and increased co-localisation and molecular association of VGAT with glial fibrillary acidic protein that increased significantly by 10 min. By 30 min, neuronal VGAT labelling was partially restored, and astrocytic VGAT was relocated to the ventral portion while it decreased in the somatic zone of the SON. Thus, synergistic astrocytic and neuronal GABAergic inhibition could ensure that vasopressin neuron firing is only transiently suppressed under hypoosmotic conditions.

Multiscale photoacoustic tomography of neural activities with GCaMP calcium indicators.

SIGNIFICANCE: Optical imaging of responses in fluorescently labeled neurons has progressed significantly in recent years. However, there is still a need to monitor neural activities at divergent spatial scales and at depths beyond the optical diffusion limit. AIM: To meet these needs, we aim to develop multiscale photoacoustic tomography (PAT) to image neural activities across spatial scales with a genetically encoded calcium indicator GCaMP. APPROACH: First, using photoacoustic microscopy, we show that depth-resolved GCaMP signals can be monitored in vivo from a fly brain in response to odor stimulation without depth scanning and even with the cuticle intact. In vivo monitoring of GCaMP signals was also demonstrated in mouse brains. Next, using photoacoustic computed tomography, we imaged neural responses of a mouse brain slice at depths beyond the optical diffusion limit. RESULTS: We provide the first unambiguous demonstration that multiscale PAT can be used to record neural activities in transgenic flies and mice with select neurons expressing GCaMP. CONCLUSIONS: Our results indicate that the combination of multiscale PAT and fluorescent neural activity indicators provides a methodology for imaging targeted neurons at various scales.

Sex Differences in the Role of CNIH3 on Spatial Memory and Synaptic Plasticity.

BACKGROUND: CNIH3 is an AMPA receptor (AMPAR) auxiliary protein prominently expressed in the dorsal hippocampus (dHPC), a region that plays a critical role in spatial memory and synaptic plasticity. However, the effects of CNIH3 on AMPAR-dependent synaptic function and behavior have not been investigated. METHODS: We assessed a gain-of-function model of Cnih3 overexpression in the dHPC and generated and characterized a line of Cnih3-/- C57BL/6 mice. We assessed spatial memory through behavioral assays, protein levels of AMPAR subunits and synaptic proteins by immunoblotting, and long-term potentiation in electrophysiological recordings. We also utilized a super-resolution imaging workflow, SEQUIN (Synaptic Evaluation and Quantification by Imaging of Nanostructure), for analysis of nanoscale synaptic connectivity in the dHPC. RESULTS: Overexpression of Cnih3 in the dHPC improved short-term spatial memory in female mice but not in male mice. Cnih3-/- female mice exhibited weakened short-term spatial memory, reduced dHPC synapse density, enhanced expression of calcium-impermeable AMPAR (GluA2-containing) subunits in synaptosomes, and attenuated long-term potentiation maintenance compared with Cnih3+/+ control mice; Cnih3-/- males were unaffected. Further investigation revealed that deficiencies in spatial memory and changes in AMPAR composition and synaptic plasticity were most pronounced during the metestrus phase of the estrous cycle in female Cnih3-/- mice. CONCLUSIONS: This study identified a novel effect of sex and estrous on CNIH3's role in spatial memory and synaptic plasticity. Manipulation of CNIH3 unmasked sexually dimorphic effects on spatial memory, synaptic function, AMPAR composition, and hippocampal plasticity. These findings reinforce the importance of considering sex as a biological variable in studies of memory and hippocampal synaptic function.

Mild chronic perturbation of inhibition severely alters hippocampal function.

Pentameric GABAA receptors mediate a large share of CNS inhibition. The γ2 subunit is a typical constituent. At least 11 mutations in the γ2 subunit cause human epilepsies, making the role of γ2-containing receptors in brain function of keen basic and translational interest. How small changes to inhibition may cause brain abnormalities, including seizure disorders, is unclear. In mice, we perturbed fast inhibition with a point mutation T272Y (T6'Y in the second membrane-spanning domain) to the γ2 subunit. The mutation imparts resistance to the GABAA receptor antagonist picrotoxin, allowing verification of mutant subunit incorporation. We confirmed picrotoxin resistance and biophysical properties in recombinant receptors. T6'Y γ2-containing receptors also exhibited faster deactivation but unaltered steady-state properties. Adult T6'Y knockin mice exhibited myoclonic seizures and abnormal cortical EEG, including abnormal hippocampal-associated theta oscillations. In hippocampal slices, picrotoxin-insensitive inhibitory synaptic currents exhibited fast decay. Excitatory/inhibitory balance was elevated by an amount expected from the IPSC alteration. Partial pharmacological correction of γ2-mediated IPSCs with diazepam restored total EEG power toward baseline, but had little effect on the abnormal low-frequency peak in the EEG. The results suggest that at least part of the abnormality in brain function arises from the acute effects of truncated inhibition.

24S-hydroxycholesterol and 25-hydroxycholesterol differentially impact hippocampal neuronal survival following oxygen-glucose deprivation.

N-methyl-D-aspartate receptors (NMDARs), a major subtype of glutamate receptor mediating excitatory transmission throughout the CNS, participate in ischemia-induced neuronal death. Unfortunately, undesired side effects have limited the strategy of inhibiting/blocking NMDARs as therapy. Targeting endogenous positive allosteric modulators of NMDAR function may offer a strategy with fewer downsides. Here, we explored whether 24S-hydroxycholesterol (24S-HC), an endogenous positive NMDAR modulator characterized recently by our group, participates in NMDAR-mediated excitotoxicity following oxygen-glucose deprivation (OGD) in primary neuron cultures. 24S-HC is the major brain cholesterol metabolite produced exclusively in neurons near sites of glutamate transmission. By selectively potentiating NMDAR current, 24S-HC may participate in NMDAR-mediated excitotoxicity following energy failure, thus impacting recovery after stroke. In support of this hypothesis, our findings indicate that exogenous application of 24S-HC exacerbates NMDAR-dependent excitotoxicity in primary neuron culture following OGD, an ischemic-like challenge. Similarly, enhancement of endogenous 24S-HC synthesis reduced survival rate. On the other hand, reducing endogenous 24S-HC synthesis alleviated OGD-induced cell death. We found that 25-HC, another oxysterol that antagonizes 24S-HC potentiation, partially rescued OGD-mediated cell death in the presence or absence of exogenous 24S-HC application, and 25-HC exhibited NMDAR-dependent/24S-HC-dependent neuroprotection, as well as NMDAR-independent neuroprotection in rat tissue but not mouse tissue. Our findings suggest that both endogenous and exogenous 24S-HC exacerbate OGD-induced damage via NMDAR activation, while 25-HC exhibits species dependent neuroprotection through both NMDAR-dependent and independent mechanisms.

24(S)-Hydroxycholesterol as a Modulator of Neuronal Signaling and Survival.

The major cholesterol metabolite in brain, 24(S)-hydroxycholesterol (24S-HC), serves as a vehicle for cholesterol removal. Its effects on neuronal function, however, have only recently begun to be investigated. Here, we review that nascent work. Our own studies have demonstrated that 24S-HC has potent positive modulatory effects on N-methyl-d-aspartate (NMDA) receptor (NMDAR) function. This could have implications not only for brain plasticity but also for pathological NMDAR overuse. Other work has demonstrated effects of 24S-HC on neuronal survival and as a possible biomarker of neurodegenerative disease. Depending on circumstances, both upregulation/mimicry of 24S-HC signaling and down-regulation/antagonism may have therapeutic potential. We are interested in the possibility that synthetic analogues of 24S-HC with positive effects at NMDARs may hold neurotherapeutic promise, given the role of NMDA receptor hypofunction in certain neuropsychiatric disorders.

Specificity and efficiency of reporter expression in adult neural progenitors vary substantially among nestin-CreER(T2) lines.

Transgenic lines expressing a controllable form of Cre recombinase have become valuable tools for manipulating gene expression in adult neural progenitors and their progeny. Neural progenitors express several proteins that distinguish them from mature neurons, and the promoters for these genes have been co-opted to produce selective transgene expression within this population. To date, nine CreER(T2) transgenic lines have been designed using the nestin promoter; however, only a subset are capable of eliciting expression within both neurogenic zones of the adult brain. Here we compare three such nestin-CreER(T2) lines to evaluate specificity of expression and efficiency of recombination. Each line was examined by using three different Cre reporter strains that varied in sensitivity. We found that all three nestin-CreER(T2) strains induced reporter expression within the main neurogenic areas, albeit to varying degrees depending on the reporter. Unexpectedly, we found that two of the three lines induced substantial reporter expression outside of neurogenic areas. These lines produced strong labeling in cerebellar granule neurons, with additional expression in the cortex, hippocampus, striatum, and thalamus. Reporter expression in the third nestin-CreER(T2) line was considerably more specific, but was also less efficient, labeling a smaller percentage of the target population than the other two drivers. Our findings suggest that each nestin-CreER(T2) line may best serve different experimental needs, depending on whether specificity or efficiency is of greatest concern. Our study further demonstrates that each new pair of driver and responder lines should be evaluated independently, as both components can significantly influence the resulting expression pattern.

Astrocyte calcium microdomains are inhibited by bafilomycin A1 and cannot be replicated by low-level Schaffer collateral stimulation in situ.

Astrocyte Gq GPCR and IP3 receptor-dependent Ca(2+) elevations occur spontaneously in situ and in vivo. These events vary considerably in size, often remaining confined to small territories of astrocyte processes called "microdomains" and sometimes propagating over longer distances that can include the soma. It has remained unclear whether these events are driven by constitutive (basal) GPCR signaling activity, neuronal action potential-dependent or quantal vesicular release, or some combination of these mechanisms. Here, we applied manipulations to increase or inhibit neuronal vesicular neurotransmitter release together with low-level stimulation of Schaffer collaterals in acute mouse hippocampal slices in an effort to determine the mechanisms underlying spontaneous astrocyte Ca(2+) events. We found no significant change in spontaneous microdomain astrocyte Ca(2+) elevations when neuronal action potentials were significantly enhanced or blocked. The astrocyte Ca(2+) activity was also not affected by inhibitors of group I mGluRs. However, blockade of miniature neurotransmitter release using Bafilomycin A1 significantly reduced the frequency of microdomain astrocyte Ca(2+) elevations. We then tested whether astrocyte Ca(2+) microdomains can be evoked by low intensity SC stimulation. Importantly, microdomains could not be reproduced even using single, low intensity pulses to the SCs at a minimum distance from the astrocyte. Evoked astrocyte Ca(2+) responses most often included the cell soma, were reduced by group I mGluR antagonists, and were larger in size compared to spontaneous Ca(2+) microdomains. Overall, our findings suggest that spontaneous microdomain astrocyte Ca(2+) elevations are not driven by neuronal action potentials but require quantal release of neurotransmitter which cannot be replicated by stimulation of Schaffer collaterals.

Calcium Signaling and Gliotransmission in Normal vs. Reactive Astrocytes.

A prominent area of neuroscience research over the past 20 years has been the acute modulation of neuronal synaptic activity by Ca(2+)-dependent release of the transmitters ATP, D-serine, and glutamate (called gliotransmitters) by astrocytes. Although the physiological relevance of this mechanism is under debate, emerging evidence suggests that there are critical factors in addition to Ca(2+) that are required for gliotransmitters to be released from astrocytes. Interestingly, these factors include activated microglia and the proinflammatory cytokine Tumor Necrosis Factor α (TNFα), chemotactic cytokine Stromal cell-Derived Factor-1α (SDF-1α), and inflammatory mediator prostaglandin E2 (PGE(2)). Of note, microglial activation and release of inflammatory molecules from activated microglia and reactive astrocytes can occur within minutes of a triggering stimulus. Therefore, activation of astrocytes by inflammatory molecules combined with Ca(2+) elevations may lead to gliotransmitter release, and be an important step in the early sequence of events contributing to hyperexcitability, excitotoxicity, and neurodegeneration in the damaged or diseased brain. In this review, we will first examine evidence questioning Ca(2+)-dependent gliotransmitter release from astrocytes in healthy brain tissue, followed by a close examination of recent work suggesting that Ca(2+)-dependent gliotransmitter release occurs as an early event in the development of neurological disorders and neuroinflammatory and neurodegenerative diseases.

Bidirectional scaling of astrocytic metabotropic glutamate receptor signaling following long-term changes in neuronal firing rates.

Very little is known about the ability of astrocytic receptors to exhibit plasticity as a result of changes in neuronal activity. Here we provide evidence for bidirectional scaling of astrocytic group I metabotropic glutamate receptor signaling in acute mouse hippocampal slices following long-term changes in neuronal firing rates. Plasticity of astrocytic mGluRs was measured by recording spontaneous and evoked Ca²âº elevations in both astrocytic somata and processes. An exogenous astrocytic Gq G protein-coupled receptor was resistant to scaling, suggesting that the alterations in astrocyte Ca²âº signaling result from changes in activity of the surface mGluRs rather than a change in intracellular G protein signaling molecules. These findings suggest that astrocytes actively detect shifts in neuronal firing rates and adjust their receptor signaling accordingly. This type of long-term plasticity in astrocytes resembles neuronal homeostatic plasticity and might be important to ensure an optimal or expected level of input from neurons.