Astrocytes Are the Source of TNF Mediating Homeostatic Synaptic Plasticity.

Tumor necrosis factor α (TNF) mediates homeostatic synaptic plasticity (HSP) in response to chronic activity blockade, and prior work has established that it is released from glia. Here we demonstrate that astrocytes are the necessary source of TNF during HSP. Hippocampal cultures from rats of both sexes depleted of microglia still will increase TNF levels following activity deprivation and still express TTX-driven HSP. Slice cultures from mice of either sex with a conditional deletion of TNF from microglia also express HSP, but critically, slice cultures with a conditional deletion of TNF from astrocytes do not. In astrocytes, glutamate signaling is sufficient to reduce NFκB signaling and TNF mRNA levels. Further, chronic TTX treatment increases TNF in an NFκB-dependent manner, although NFκB signaling is dispensable for the neuronal response to TTX-driven HSP. Thus, astrocytes can sense neuronal activity through glutamate spillover and increase TNF production when activity falls, to drive HSP through the production of TNF.

Early TNF-Dependent Regulation of Excitatory and Inhibitory Synapses on Striatal Direct Pathway Medium Spiny Neurons in the YAC128 Mouse Model of Huntington's Disease.

Huntington's disease (HD) is a neurodegenerative disease caused by a polyglutamine expansion in the huntingtin gene. Neurodegeneration first occurs in the striatum, accompanied by an elevation in inflammatory cytokines. Using the presymptomatic male YAC128 HD model mouse, we examined the synaptic input onto the striatal medium spiny neurons to look for early changes that precede degeneration. We observed an increase in excitatory synaptic strength, as measured by AMPA/NMDA ratios, specifically on direct pathway D1 receptor expressing medium spiny neurons, with no changes on indirect pathway neurons. The changes in excitation were accompanied by a decrease in inhibitory synaptic strength, as measured by the amplitude of miniature inhibitory synaptic currents. The pro-inflammatory cytokine tumor necrosis factor alpha (TNF) was elevated in the striatum of YAC128 at the ages examined. Critically, the changes in excitatory and inhibitory inputs are both dependent on TNF signaling, as blocking TNF signaling genetically or pharmacological normalized synaptic strength. The observed changes in synaptic function are similar to the changes seen in D1 medium spiny neurons treated with high levels of TNF, suggesting that saturating levels of TNF exist in the striatum even at early stages of HD. The increase in glutamatergic synaptic strength and decrease in inhibitory synaptic strength would increase direct pathway neuronal excitability, which may potentiate excitotoxicity during the progress of HD.SIGNIFICANCE STATEMENT The striatum is the first structure to degenerate in Huntington's disease, but the early changes that presage the degeneration are not well defined. Here we identify early synaptic changes in the YAC128 mouse model of Huntington's disease specifically on a subpopulation of striatal neurons. These neurons have stronger excitatory synapses and weaker inhibitory inputs, and thus would increase the susceptibility to excitotoxicity. These changes are dependent on signaling by the pro-inflammatory cytokine TNFα. TNF is elevated even at early presymptomatic stages, and blocking TNF signaling even acutely will reverse the synaptic changes. This suggests early intervention could be important therapeutically.

Elevated TNF-α Leads to Neural Circuit Instability in the Absence of Interferon Regulatory Factor 8.

Interferon regulatory factor 8 (IRF8) is a transcription factor necessary for the maturation of microglia, as well as other peripheral immune cells. It also regulates the transition of microglia and other immune cells to a pro-inflammatory phenotype. Irf8 is also a known risk gene for multiple sclerosis and lupus, and it has recently been shown to be downregulated in schizophrenia. While most studies have focused on IRF8-dependent regulation of immune cell function, little is known about how it impacts neural circuits. Here, we show by RNAseq from Irf8 -/- male and female mouse brains that several genes involved in regulation of neural activity are dysregulated. We then show that these molecular changes are reflected in heightened neural excitability and a profound increase in susceptibility to lethal seizures in male and female Irf8 -/- mice. Finally, we identify that TNF-α is elevated specifically in microglia in the CNS, and genetic or acute pharmacological blockade of TNF-α in the Irf8 -/- CNS rescued the seizure phenotype. These results provide important insights into the consequences of IRF8 signaling and TNF-α on neural circuits. Our data further suggest that neuronal function is impacted by loss of IRF8, a factor involved in neuropsychiatric and neurodegenerative diseases.SIGNIFICANCE STATEMENT Here, we identify a previously unknown and key role for interferon regulator factor 8 (IRF8) in regulating neural excitability and seizures. We further determine that these effects on neural circuits are through elevated TNF-α in the CNS. As IRF8 has most widely been studied in the context of regulating the development and inflammatory signaling in microglia and other immune cells, we have uncovered a novel function. Further, IRF8 is a risk gene for multiple sclerosis and lupus, IRF8 is dysregulated in schizophrenia, and elevated TNF-α has been identified in a multitude of neurologic conditions. Thus, elucidating these IRF8 and TNF-α-dependent effects on brain circuit function has profound implications for understanding underlying, therapeutically relevant mechanisms of disease.

Sustained TNF signaling is required for the synaptic and anxiety-like behavioral response to acute stress.

Acute stress triggers plasticity of forebrain synapses as well as behavioral changes. Here we reveal that Tumor Necrosis Factor α (TNF) is a required downstream mediator of the stress response in mice, necessary for stress-induced synaptic potentiation in the ventral hippocampus and for an increase in anxiety-like behaviour. Acute stress is sufficient to activate microglia, triggering the long-term release of TNF. Critically, on-going TNF signaling specifically in the ventral hippocampus is necessary to sustain both the stress-induced synaptic and behavioral changes, as these could be reversed hours after induction by antagonizing TNF signaling. This demonstrates that TNF maintains the synaptic and behavioral stress response in vivo, making TNF a potential novel therapeutic target for stress disorders.

N-Acetylcysteine Suppresses Microglial Inflammation and Induces Mortality Dose-Dependently via Tumor Necrosis Factor-α Signaling.

N-acetylcysteine (NAC) is an antioxidant that prevents tumor necrosis factor (TNF)-α-induced cell death, but it also acts as a pro-oxidant, promoting reactive oxygen species independent apoptosis. Although there is plausible preclinical evidence for the use of NAC in the treatment of psychiatric disorders, deleterious side effects are still of concern. Microglia, key innate immune cells in the brain, play an important role in inflammation in psychiatric disorders. This study aimed to investigate the beneficial and deleterious effects of NAC on microglia and stress-induced behavior abnormalities in mice, and its association with microglial TNF-α and nitric oxide (NO) production. The microglial cell line MG6 was stimulated by Escherichia coli lipopolysaccharide (LPS) using NAC at varying concentrations for 24 h. NAC inhibited LPS-induced TNF-α and NO synthesis, whereas high concentrations (≥30 mM) caused MG6 mortality. Intraperitoneal injections of NAC did not ameliorate stress-induced behavioral abnormalities in mice, but high-doses induced microglial mortality. Furthermore, NAC-induced mortality was alleviated in microglial TNF-α-deficient mice and human primary M2 microglia. Our findings provide ample evidence for the use of NAC as a modulating agent of inflammation in the brain. The risk of side effects from NAC on TNF-α remains unclear and merits further mechanistic investigations.

Deficient Autophagy in Microglia Aggravates Repeated Social Defeat Stress-Induced Social Avoidance.

Major depressive disorder (MDD) is associated with repeated exposure to environmental stress. Autophagy is activated under various stress conditions that are associated with several diseases in the brain. This study was aimed at elucidating the autophagy signaling changes in the prefrontal cortex (PFC) under repeated social defeat (RSD) to investigate the involvement of microglial autophagy in RSD-induced behavioral changes. We found that RSD stress, an animal model of MDD, significantly induced initial autophagic signals followed by increased transcription of autophagy-related genes (Atg6, Atg7, and Atg12) in the PFC. Similarly, significantly increased transcripts of ATGs (Atg6, Atg7, Atg12, and Atg5) were confirmed in the postmortem PFC of patients with MDD. The protein levels of the prefrontal cortical LC3B were significantly increased, whereas p62 was significantly decreased in the resilient but not in susceptible mice and patients with MDD. This indicates that enhanced autophagic flux may alleviate stress-induced depression. Furthermore, we identified that FKBP5, an early-stage autophagy regulator, was significantly increased in the PFC of resilient mice at the transcript and protein levels. In addition, the resilient mice exhibited enhanced autophagic flux in the prefrontal cortical microglia, and the autophagic deficiency in microglia aggravated RSD-induced social avoidance, indicating that microglial autophagy involves stress-induced behavioral changes.

Divergent Synaptic Scaling of Miniature EPSCs following Activity Blockade in Dissociated Neuronal Cultures.

Neurons can respond to decreased network activity with a homeostatic increase in the amplitudes of miniature EPSCs (mEPSCs). The prevailing view is that mEPSC amplitudes are uniformly multiplied by a single factor, termed "synaptic scaling." Deviations from purely multiplicative scaling have been attributed to biological differences, or to a distortion imposed by a detection threshold limit. Here, we demonstrate in neurons dissociated from cortices of male and female mice that the shift in mEPSC amplitudes observed in the experimental data cannot be reproduced by simulation of uniform multiplicative scaling, with or without the distortion caused by applying a detection threshold. Furthermore, we demonstrate explicitly that the scaling factor is not uniform but is close to 1 for small mEPSCs, and increases with increasing mEPSC amplitude across a substantial portion of the data. This pattern was also observed for previously published data from dissociated mouse hippocampal neurons and dissociated rat cortical neurons. The finding of "divergent scaling" shifts the current view of homeostatic plasticity as a process that alters all synapses on a neuron equally to one that must accommodate the differential effect observed for small versus large mEPSCs. Divergent scaling still accomplishes the essential homeostatic task of modifying synaptic strengths in the opposite direction of the activity change, but the consequences are greatest for those synapses which individually are more likely to bring a neuron to threshold.SIGNIFICANCE STATEMENT In homeostatic plasticity, the responses to chronic increases or decreases in network activity act in the opposite direction to restore normal activity levels. Homeostatic plasticity is likely to play a role in diseases associated with long-term changes in brain function, such as epilepsy and neuropsychiatric illnesses. One homeostatic response is the increase in synaptic strength following a chronic block of activity. Research is focused on finding a globally expressed signaling pathway, because it has been proposed that the plasticity is uniformly expressed across all synapses. Here, we show that the plasticity is not uniform. Our work suggests that homeostatic signaling molecules are likely to be differentially expressed across synapses.

Dystroglycan mediates homeostatic synaptic plasticity at GABAergic synapses.

Dystroglycan (DG), a cell adhesion molecule well known to be essential for skeletal muscle integrity and formation of neuromuscular synapses, is also present at inhibitory synapses in the central nervous system. Mutations that affect DG function not only result in muscular dystrophies, but also in severe cognitive deficits and epilepsy. Here we demonstrate a role of DG during activity-dependent homeostatic regulation of hippocampal inhibitory synapses. Prolonged elevation of neuronal activity up-regulates DG expression and glycosylation, and its localization to inhibitory synapses. Inhibition of protein synthesis prevents the activity-dependent increase in synaptic DG and GABAA receptors (GABAARs), as well as the homeostatic scaling up of GABAergic synaptic transmission. RNAi-mediated knockdown of DG blocks homeostatic scaling up of inhibitory synaptic strength, as does knockdown of like-acetylglucosaminyltransferase (LARGE)--a glycosyltransferase critical for DG function. In contrast, DG is not required for the bicuculline-induced scaling down of excitatory synaptic strength or the tetrodotoxin-induced scaling down of inhibitory synaptic strength. The DG ligand agrin increases GABAergic synaptic strength in a DG-dependent manner that mimics homeostatic scaling up induced by increased activity, indicating that activation of this pathway alone is sufficient to regulate GABAAR trafficking. These data demonstrate that DG is regulated in a physiologically relevant manner in neurons and that DG and its glycosylation are essential for homeostatic plasticity at inhibitory synapses.

An adaptive role of TNFα in the regulation of striatal synapses.

Elevation of inflammatory cytokines in the striatum precedes symptoms in a number of motor dysfunctions, but it is unclear whether this is part of the disease process or an adaptive response to the pathology. In pyramidal cells, TNFα drives the insertion of AMPA-type glutamate receptors into synapses, and contributes to the homeostatic regulation of circuit activity in the developing neocortex. Here we demonstrate that in the mouse dorsolateral striatum, TNFα drives the internalization of AMPARs and reduces corticostriatal synaptic strength, dephosphorylates DARPP-32 and GluA1, and results in a preferential removal of Ca(2+)-permeable AMPARs. Striatal TNFα signaling appears to be adaptive in nature, as TNFα is upregulated in response to the prolonged blockade of D2 dopamine receptors and is necessary to reduce the expression of extrapyramidal symptoms induced by chronic haloperidol treatment. These data indicate that TNFα is a regulator of glutamatergic synaptic strength in the adult striatum in a manner distinct from its regulation of synapses on pyramidal cells and mediates an adaptive response during pathological conditions.

Astrocytic TNFα regulates the behavioral response to antidepressants.

Recent studies have suggested that cytokines, and in particular tumor necrosis factor alpha (TNFα), have a role in modulating antidepressant efficacy. To directly test this idea, we compared the response of TNFα(-/-) mice and astrocyte-specific TNFα(-/-) mice to the antidepressants fluoxetine and desipramine. Using standard behavior models for measuring antidepressant efficacy, the forced swim test (FST) and tail suspension test (TST), we determined that TNFα(-/-) mice were essentially normal in basal behavior in the FST and TST. However, TNFα(-/-) mice showed no behavioral response to a standard dose of chronic antidepressant treatment, in sharp contrast to wildtype mice. Similar results were seen with acute antidepressant treatment, but TNFα(-/-) mice did respond to a very high-dose acute antidepressant treatment. We also assessed in vitro and in vivo effects of fluoxetine on TNFα expression. Glia responded to serotonin in vitro and fluoxetine in vivo by upregulating TNFα mRNA. Consistent with this source of TNFα, mice with an astrocyte-specific deletion of TNFα also did not respond to standard chronic antidepressant treatment. These data suggest that astrocytic TNFα is important to the sensitivity of the behavioral response to administration of antidepressants.

TNF-α downregulates inhibitory neurotransmission through protein phosphatase 1-dependent trafficking of GABA(A) receptors.

Inflammation has been implicated in the progression of neurological disease, yet precisely how inflammation affects neuronal function remains unclear. Tumor necrosis factor-α (TNFα) is a proinflammatory cytokine that regulates synapse function by controlling neurotransmitter receptor trafficking and homeostatic synaptic plasticity. Here we characterize the mechanisms through which TNFα regulates inhibitory synapse function in mature rat and mouse hippocampal neurons. Acute application of TNFα induces a rapid and persistent decrease of inhibitory synaptic strength and downregulation of cell-surface levels of GABA(A)Rs containing α1, α2, ß2/3, and γ2 subunits. We show that trafficking of GABA(A)Rs in response to TNFα is mediated by neuronally expressed TNF receptor 1 and requires activation of p38 MAPK, phosphatidylinositol 3-kinase, protein phosphatase 1 (PP1), and dynamin GTPase. Furthermore, TNFα enhances the association of PP1 with GABA(A)R ß3 subunits and dephosphorylates a site on ß3 known to regulate phospho-dependent interactions with the endocytic machinery. Conversely, we find that calcineurin and PP2A are not essential components of the signaling pathway and that clustering of the scaffolding protein gephyrin is only reduced after the initial receptor endocytosis. Together, these findings demonstrate a distinct mechanism of regulated GABA(A)R endocytosis that may contribute to the disruption of circuit homeostasis under neuroinflammatory conditions.

Persistent synaptic scaling independent of AMPA receptor subunit composition.

Despite long-standing evidence that the specific intracellular domains of AMPA-type glutamate receptor (AMPAR) subunits are critical for trafficking, it has recently been demonstrated that there is no absolute requirement for any AMPAR subunit for the receptor insertion underlying LTP. It is unclear whether this holds true to other forms of plasticity. Homeostatic synaptic plasticity (HSP) is an important form of negative feedback that provides stability to neuronal networks, and results at least in part from the insertion of AMPARs into glutamatergic synapses following chronic reductions in neuronal activity. Similar to LTP, the GluA1 subunit has been suggested to be the requisite subunit for HSP-induced AMPAR insertion and acute treatment with signaling molecules that underlie some forms of HSP results in the preferential incorporation of GluA2-lacking receptors. However, knockdown experiments have instead implicated a requirement for the GluA2 subunit. Here we re-examined the requirement for specific AMPAR subunit during chronic tetrodotoxin-induced HSP using hippocampal cultures derived from AMPAR subunit knock-out mice. We observed HSP in cultures from GluA1⁻/⁻, GluA2⁻/⁻, and GluA2⁻/⁻ GluA3⁻/⁻ mice, and conclude that, as with LTP, there is no subunit requirement for HSP.

An essential role for inhibitor-2 regulation of protein phosphatase-1 in synaptic scaling.

Protein phosphatase-1 (PP1) activity is important for many calcium-dependent neuronal functions including Hebbian synaptic plasticity and learning and memory. PP1 activity is necessary for the induction of long-term depression, whereas downregulation of PP1 activity is required for the normal induction of long-term potentiation. However, how PP1 is activated is not clear. Moreover, it is not known whether PP1 plays a role in homeostatic synaptic scaling, another form of synaptic plasticity which functions to reset the neuronal firing rate in response to chronic neuronal activity perturbations. In this study, we found that PP1 inhibitor-2 (I-2) is phosphorylated at serine 43 (S43) in rat and mouse cortical neurons in response to bicuculine application. Expression of I-2 phosphorylation-blocking mutant I-2 (S43A) blocked the dephosphorylation of GluA2 at serine 880, AMPA receptor trafficking, and synaptic downscaling induced by bicuculline application. Our data suggest that the phosphorylation of I-2 at S43 appears to be mediated by L-type calcium channels and calcium/calmodulin-dependent myosin light-chain kinase. Our work thus reveals a novel calcium-induced PP1 activation pathway critical for homeostatic synaptic plasticity.

Netrin-1 promotes excitatory synaptogenesis between cortical neurons by initiating synapse assembly.

Netrin-1 is a secreted protein that directs long-range axon guidance during early stages of neural circuit formation and continues to be expressed in the mammalian forebrain during the postnatal period of peak synapse formation. Here we demonstrate a synaptogenic function of netrin-1 in rat and mouse cortical neurons and investigate the underlying mechanism. We report that netrin-1 and its receptor DCC are widely expressed by neurons in the developing mammalian cortex during synapse formation and are enriched at synapses in vivo. We detect DCC protein distributed along the axons and dendrites of cultured cortical neurons and provide evidence that newly translated netrin-1 is selectively transported to dendrites. Using gain and loss of function manipulations, we demonstrate that netrin-1 increases the number and strength of excitatory synapses made between developing cortical neurons. We show that netrin-1 increases the complexity of axon and dendrite arbors, thereby increasing the probability of contact. At sites of contact, netrin-1 promotes adhesion, while locally enriching and reorganizing the underlying actin cytoskeleton through Src family kinase signaling and m-Tor-dependent protein translation to locally cluster presynaptic and postsynaptic proteins. Finally, we demonstrate using whole-cell patch-clamp electrophysiology that netrin-1 increases the frequency and amplitude of mEPSCs recorded from cortical pyramidal neurons. These findings identify netrin-1 as a synapse-enriched protein that promotes synaptogenesis between mammalian cortical neurons.

ProNGF induces TNFalpha-dependent death of retinal ganglion cells through a p75NTR non-cell-autonomous signaling pathway.

Neurotrophin binding to the p75 neurotrophin receptor (p75(NTR)) activates neuronal apoptosis following adult central nervous system injury, but the underlying cellular mechanisms remain poorly defined. In this study, we show that the proform of nerve growth factor (proNGF) induces death of retinal ganglion cells in adult rodents via a p75(NTR)-dependent signaling mechanism. Expression of p75(NTR) in the adult retina is confined to Müller glial cells; therefore we tested the hypothesis that proNGF activates a non-cell-autonomous signaling pathway to induce retinal ganglion cell (RGC) death. Consistent with this, we show that proNGF induced robust expression of tumor necrosis factor alpha (TNFalpha) in Müller cells and that genetic or biochemical ablation of TNFalpha blocked proNGF-induced death of retinal neurons. Mice rendered null for p75(NTR), its coreceptor sortilin, or the adaptor protein NRAGE were defective in proNGF-induced glial TNFalpha production and did not undergo proNGF-induced retinal ganglion cell death. We conclude that proNGF activates a non-cell-autonomous signaling pathway that causes TNFalpha-dependent death of retinal neurons in vivo.

Astrocytes display complex and localized calcium responses to single-neuron stimulation in the hippocampus.

Astrocytes show a complex structural and physiological interplay with neurons and respond to neuronal activation in vitro and in vivo with intracellular calcium elevations. These calcium changes enable astrocytes to modulate synaptic transmission and plasticity through various mechanisms. However, the response pattern of astrocytes to single neuronal depolarization events still remains unresolved. This information is critical for fully understanding the coordinated network of neuron-glial signaling in the brain. To address this, we developed a system to map astrocyte calcium responses along apical dendrites of CA1 pyramidal neurons in hippocampal slices using single-neuron stimulation with channelrhodopsin-2. This technique allowed selective neuronal depolarization without invasive manipulations known to alter calcium levels in astrocytes. Light-evoked neuronal depolarization was elicited and calcium events in surrounding astrocytes were monitored using the calcium-sensitive dye Calcium Orange. Stimulation of single neurons caused calcium responses in populations of astrocytes along the apical axis of CA1 cell dendrites. Calcium responses included single events that were synchronized with neuronal stimulation and poststimulus changes in calcium event frequency, both of which were modulated by glutamatergic and purinergic signaling. Individual astrocytes near CA1 cells showed low ability to respond to repeated neuronal depolarization events. However, the response of the surrounding astrocyte population was remarkably accurate. Interestingly, the reliability of responses was graded with respect to astrocyte location along the CA1 cell dendrite, with astrocytes residing in the primary dendrite subregion being most responsive. This study provides a new perspective on the dynamic response property of astrocyte ensembles to neuronal activity.

Synaptic scaling mediated by glial TNF-alpha.

Two general forms of synaptic plasticity that operate on different timescales are thought to contribute to the activity-dependent refinement of neural circuitry during development: (1) long-term potentiation (LTP) and long-term depression (LTD), which involve rapid adjustments in the strengths of individual synapses in response to specific patterns of correlated synaptic activity, and (2) homeostatic synaptic scaling, which entails uniform adjustments in the strength of all synapses on a cell in response to prolonged changes in the cell's electrical activity. Without homeostatic synaptic scaling, neural networks can become unstable and perform suboptimally. Although much is known about the mechanisms underlying LTP and LTD, little is known about the mechanisms responsible for synaptic scaling except that such scaling is due, at least in part, to alterations in receptor content at synapses. Here we show that synaptic scaling in response to prolonged blockade of activity is mediated by the pro-inflammatory cytokine tumour-necrosis factor-alpha (TNF-alpha). Using mixtures of wild-type and TNF-alpha-deficient neurons and glia, we also show that glia are the source of the TNF-alpha that is required for this form of synaptic scaling. We suggest that by modulating TNF-alpha levels, glia actively participate in the homeostatic activity-dependent regulation of synaptic connectivity.

Restorative Action of Vitamin D3 on Motor Dysfunction Through Enhancement of Neurotrophins and Antioxidant Expression in the Striatum.

A number of studies has explored a positive correlation between low levels of serum Vitamin D3 (VD; cholecalciferol) and development of neurodegenerative diseases including Huntington's disease (HD). In the present study, the prophylactic effect of VD on motor dysfunction was studied in an experimental model of HD. An HD-like syndrome was induced in male C57BL/6 mice through an intraperitoneal injection (i.p) of 3-NP for 3 consecutive doses at 12 h interval of time as described previously (Amende et al. 2005). This study investigated thein-vivotherapeutic potential of VD (500 IU/kg/day) supplementation on movement, motor coordination, motor activity and biochemical changes in this HD model. Mice were divided into four groups: Group I: Control (saline); Group II: 3-NP induced HD (HD); Group III: Vitamin D3 (VD) and Group IV: 3-NP induced + post Vitamin D3 injection (HD + VD). All groups of mice were tested for locomotion, gait analysis and rotarod performances over a span of 30-days. VD administration rescued locomotor dysfunction and neuromuscular impairment in HD mice with no change in gait dynamics. In addition, administration of VD to 3-NP treated mice led to a significant enhancement in the expression of key neurotrophic factors including brain-derived neurotrophic factor (BDNF) and nerve-growth factor (NGF), the Vitamin D receptor (VDR), and antioxidant markers (catalases [Cat] and glutathione peroxidase [GpX4]) in the striatum, suggesting a detoxification effect of VD. Altogether, our results show that VD supplementation induces survival signals, diminishes oxidative stress, and reduces movement and motor dysfunction in HD.

Protein phosphatase-1 inhibitor-2 promotes PP1γ positive regulation of synaptic transmission.

Inhibitor-2 (I-2) is a prototypic inhibitor of protein phosphatase-1 (PP1), a major serine-threonine phosphatase that regulates synaptic plasticity and learning and memory. Although I-2 is a potent inhibitor of PP1 in vitro, our previous work has elucidated that, in vivo, I-2 may act as a positive regulator of PP1. Here we show that I-2 and PP1γ, but not PP1α, positively regulate synaptic transmission in hippocampal neurons. Moreover, we demonstrated that I-2 enhanced PP1γ interaction with its major synaptic scaffold, neurabin, by Förster resonance energy transfer (FRET)/Fluorescence lifetime imaging microscopy (FLIM) studies, while having a limited effect on PP1 auto-inhibitory phosphorylation. Furthermore, our study indicates that the effect of I-2 on PP1 activity in vivo is dictated by I-2 threonine-72 phosphorylation. Our work thus demonstrates a molecular mechanism by which I-2 positively regulates PP1 function in synaptic transmission.

TNF-Mediated Homeostatic Synaptic Plasticity: From in vitro to in vivo Models.

Since it was first described almost 30 years ago, homeostatic synaptic plasticity (HSP) has been hypothesized to play a key role in maintaining neuronal circuit function in both developing and adult animals. While well characterized in vitro, determining the in vivo roles of this form of plasticity remains challenging. Since the discovery that the pro-inflammatory cytokine tumor necrosis factor-α (TNF-α) mediates some forms of HSP, it has been possible to probe some of the in vivo contribution of TNF-mediated HSP. Work from our lab and others has found roles for TNF-HSP in a variety of functions, including the developmental plasticity of sensory systems, models of drug addiction, and the response to psychiatric drugs.