Ciliary Neurotrophic Factor Protects Striatal Neurons against Excitotoxicity by Enhancing Glial Glutamate Uptake

Ciliary neurotrophic factor (CNTF) is a potent neuroprotective cytokine in different animal models of glutamate-induced excitotoxicity, although its action mechanisms are still poorly characterized. We tested the hypothesis that an increased function of glial glutamate transporters (GTs) could underlie CNTF-mediated neuroprotection. We show that neuronal loss induced by in vivo striatal injection of the excitotoxin quinolinic acid (QA) was significantly reduced (by ∼75%) in CNTF-treated animals. In striatal slices, acute QA application dramatically inhibited corticostriatal field potentials (FPs), whose recovery was significantly higher in CNTF rats compared to controls (∼40% vs. ∼7%), confirming an enhanced resistance to excitotoxicity. The GT inhibitor dl-threo-β-benzyloxyaspartate greatly reduced FP recovery in CNTF rats, supporting the role of GT in CNTF-mediated neuroprotection. Whole-cell patch-clamp recordings from striatal medium spiny neurons showed no alteration of basic properties of striatal glutamatergic transmission in CNTF animals, but the increased effect of a low-affinity competitive glutamate receptor antagonist (γ-d-glutamylglycine) also suggested an enhanced GT function. These data strongly support our hypothesis that CNTF is neuroprotective via an increased function of glial GTs, and further confirms the therapeutic potential of CNTF for the clinical treatment of progressive neurodegenerative diseases involving glutamate overflow

Dysfonction des astrocytes du striatum induite par expression sélective de huntingtine mutée in vivo

La maladie de Huntington (MH) est une pathologie neurodégénérative héréditaire due à la mutation du gène codant la huntingtine (Htt). La mutation de la Htt (Httm) entraîne la perte sélective des neurones épineux de taille moyenne du striatum et ceci malgré son expression ubiquitaire. Plusieurs études suggèrent que les astrocytes pourraient participer à la pathologie en particulier dans les mécanismes d excitotoxicité impliqués dans la mort neuronale. Grâce à un nouveau vecteur lentiviral développé au laboratoire, nous avons exprimé un fragment court de la Httm dans les astrocytes du striatum de souris adulte. Nous avons observé une activation progressive des astrocytes, réaction jusqu à présent considérée comme secondaire à la dysfonction neuronale. L expression sélective de la Httm dans les astrocytes du striatum entraine une diminution d expression et d activité des transporteurs astrocytaires au glutamate GLT-1 et GLAST, 12 semaines après l injection des vecteurs. Ces résultats sont corrélés avec l observation de cerveaux de patients de la MH présentant une activation astrocytaire et une perte du transporteur de glutamate GLT-1 dès le stade pré-symptomatique. Ces résultats montrent que les astrocytes sont affectés par leur propre expression de Httm et qu ils pourraient participer précocement à la pathologie. Nous avons étudié le phénotype des neurones voisins des astrocytes dysfonctionnels et nous avons observé la diminution de deux marqueurs neuronaux, DARPP-32 et NR2B. La Httm astrocytaire pourrait donc altérer la fonction des neurones ce qui implique les interactions neurones-astrocytes dans la MH.PARIS-BIUSJ-Physique recherche (751052113) / SudocSudocFranceF

The Inside-Out Amyloid Hypothesis and Synapse Pathology in Alzheimer's Disease.

Cumulative evidence in brains and cultured neurons of Alzheimer's disease (AD) transgenic mouse models, as well as in human postmortem AD brains, highlights that age-related increases in β-amyloid peptide (Aβ), particularly in endosomes near synapses, are involved in early synapse dysfunction. Our immunoelectron microscopy and high-resolution immunofluorescence microscopy studies show that this early subcellular Aβ accumulation leads to progressive Aβ aggregation and pathology, particularly within dystrophic neurites and synapses. These studies confirm that neuritic/synaptic Aβ accumulation is the nidus of plaque formation. Aβ-dependent synapse pathology in AD models is modulated by synaptic activity and is plaque independent. The amyloid precursor protein (APP) is normally transported down neurites and appears to be preferentially processed to Aβ at synapses. Synapses are sites of early Aβ accumulation and aberrant tau phosphorylation in AD, which alter the synaptic composition at early stages of the disease. Elucidating the normal role of APP, and potentially of Aβ, at synapses should provide important insights into the mechanism(s) of Aβ-induced synapse dysfunction in AD and how to therapeutically mitigate these dysfunctions. © 2013 S. Karger AG, Basel

Detection of pre-plaque amyloid aggregation using FTIR

Background: Alzheimer's disease (AD) is characterized by misfolding and aggregation of naturally occurring beta-amyloid peptides (Aβ). These aggregates are thought to be pathogenic to neurons, although the conformation of the pathogenic Aβ species remains unclear. Biochemical extraction methods and different microscopy techniques (TEM, confocal) can be used to identify pathogenic Aβ species in the brain, although such methods can alter protein conformation or are n ot designed to determine structural details of protein assemblies

Ciliary neurotrophic factor (CNTF) activation of astrocytes decreases spreading depolarization susceptibility and increases potassium clearance

International audienceWaves of spreading depolarization (SD) have been implicated in the progressive expansion of acute brain injuries. SD can persist over several days, coincident with the time course of astrocyte activation, but little is known about how astrocyte activation may influence SD susceptibility. We examined whether activation of astrocytes modified SD threshold in hippocampal slices. Injection of a lentiviral vector encoding Ciliary neurotrophic factor (CNTF) into the hippocampus in vivo, led to sustained astrocyte activation, verified by up-regulation of glial fibrillary acidic protein (GFAP) at the mRNA and protein levels, as compared to controls injected with vector encoding LacZ. In acute brain slices from LacZ controls, localized 1M KCl microinjections invariably generated SD in CA1 hippocampus, but SD was never induced with this stimulus in CNTF tissues. No significant change in intrinsic excitability was observed in CA1 neurons, but excitatory synaptic transmission was significantly reduced in CNTF samples. mRNA levels of the predominantly astrocytic Na(+) /K(+) -ATPase pump α2 subunit were higher in CNTF samples, and the kinetics of extracellular K(+) transients during matched synaptic activation were consistent with increased K(+) uptake in CNTF tissues. Supporting a role for the Na(+) /K(+) -ATPase pump in increased SD threshold, ouabain, an inhibitor of the pump, was able to generate SD in CNTF tissues. These data support the hypothesis that activated astrocytes can limit SD onset via increased K(+) clearance and suggest that therapeutic strategies targeting these glial cells could improve the outcome following acute brain injuries associated with SD

APP depletion alters selective pre- and post-synaptic proteins

The normal role of Alzheimer's disease (AD)-linked amyloid precursor protein (APP) in the brain remains incompletely understood. Previous studies have reported that lack of APP has detrimental effects on spines and electrophysiological parameters. APP has been described to be important in synaptic pruning during development. The effect of APP knockout on mature synapses is complicated by this role in development. We previously reported on differential changes in synaptic proteins and receptors in APP mutant AD transgenic compared to wild-type neurons, which revealed selective decreases in levels of pre- and post-synaptic proteins, including of surface glutamate receptors. In the present study, we undertook a similar analysis of synaptic composition but now in APP knockout compared to wild-type mouse neurons. Here we demonstrate alterations in levels of selective pre- and post-synaptic proteins and receptors in APP knockout compared to wild-type mouse primary neurons in culture and brains of mice in youth and adulthood. Remarkably, we demonstrate selective increases in levels of synaptic proteins, such as GluA1, in neurons with APP knockout and with RNAi knockdown, which tended to be opposite to the reductions seen in AD transgenic APP mutant compared to wild-type neurons. These data reinforce that APP is important for the normal composition of synapses

CNTF activates astrocytes and protects striatal neurons against QA excitotoxicity <i>in vivo</i>.

<p><b>A:</b> CNTF activates astrocytes that re-express vimentin (red) and overexpress GFAP around GFP-positive MSNs neurons of the striatum. <b>B:</b> rats from Vehicle (Veh.), LacZ and CNTF groups were injected with 80 nmol QA and the lesion (*) volume was assessed 15 days later on NeuN-immunostained sections. CNTF significantly decreased lesion size (^§p<0.001 vs. Veh. and LacZ groups, ANOVA and Scheffé's test).</p