Myelin regeneration can occur spontaneously in demyelinating diseases such as multiple sclerosis (MS). However, the underlying mechanisms and causes of its frequent failure remain incompletely understood. Here we show, using an in-vivo remyelination model, that demyelinated axons are electrically active and generate de novo synapses with recruited oligodendrocyte progenitor cells (OPCs), which, early after lesion induction, sense neuronal activity by expressing AMPA (α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid)/kainate receptors. Blocking neuronal activity, axonal vesicular release or AMPA receptors in demyelinated lesions results in reduced remyelination. In the absence of neuronal activity there is a ∼6-fold increase in OPC number within the lesions and a reduced proportion of differentiated oligodendrocytes. These findings reveal that neuronal activity and release of glutamate instruct OPCs to differentiate into new myelinating oligodendrocytes that recover lost function. Co-localization of OPCs with the presynaptic protein VGluT2 in MS lesions implies that this mechanism may provide novel targets to therapeutically enhance remyelination.

Although ubiquitous in biological studies, the enhanced green and yellow fluorescent proteins (EGFP and EYFP) were not specifically optimized for neuroscience, and their underwhelming brightness and slow expression in brain tissue limits the fidelity of dendritic spine analysis and other indispensable techniques for studying neurodevelopment and plasticity. We hypothesized that EGFP’s low solubility in mammalian systems must limit the total fluorescence output of whole cells, and that improving folding efficiency could therefore translate into greater brightness of expressing neurons. By introducing rationally selected combinations of folding-enhancing mutations into GFP templates and screening for brightness and expression rate in human cells, we developed mGreenLantern, a fluorescent protein having up to sixfold greater brightness in cells than EGFP. mGreenLantern illuminates neurons in the mouse brain within 72 h, dramatically reducing lag time between viral transduction and imaging, while its high brightness improves detection of neuronal morphology using widefield, confocal, and two-photon microscopy. When virally expressed to projection neurons in vivo, mGreenLantern fluorescence developed four times faster than EYFP and highlighted long-range processes that were poorly detectable in EYFP-labeled cells. Additionally, mGreenLantern retains strong fluorescence after tissue clearing and expansion microscopy, thereby facilitating superresolution and whole-brain imaging without immunohistochemistry. mGreenLantern can directly replace EGFP/EYFP in diverse systems due to its compatibility with GFP filter sets, recognition by EGFP antibodies, and excellent performance in mouse, human, and bacterial cells. Our screening and rational engineering approach is broadly applicable and suggests that greater potential of fluorescent proteins, including biosensors, could be unlocked using a similar strategy.

BackgroundBrain-Derived Neurotrophic Factor (BDNF) is the main candidate for neuroprotective therapy for Huntington's disease (HD), but its conditional administration is one of its most challenging problems.ResultsHere we used transgenic mice that over-express BDNF under the control of the Glial Fibrillary Acidic Protein (GFAP) promoter (pGFAP-BDNF mice) to test whether up-regulation and release of BDNF, dependent on astrogliosis, could be protective in HD. Thus, we cross-mated pGFAP-BDNF mice with R6/2 mice to generate a double-mutant mouse with mutant huntingtin protein and with a conditional over-expression of BDNF, only under pathological conditions. In these R6/2:pGFAP-BDNF animals, the decrease in striatal BDNF levels induced by mutant huntingtin was prevented in comparison to R6/2 animals at 12 weeks of age. The recovery of the neurotrophin levels in R6/2:pGFAP-BDNF mice correlated with an improvement in several motor coordination tasks and with a significant delay in anxiety and clasping alterations. Therefore, we next examined a possible improvement in cortico-striatal connectivity in R62:pGFAP-BDNF mice. Interestingly, we found that the over-expression of BDNF prevented the decrease of cortico-striatal presynaptic (VGLUT1) and postsynaptic (PSD-95) markers in the R6/2:pGFAP-BDNF striatum. Electrophysiological studies also showed that basal synaptic transmission and synaptic fatigue both improved in R6/2:pGAP-BDNF mice.ConclusionsThese results indicate that the conditional administration of BDNF under the GFAP promoter could become a therapeutic strategy for HD due to its positive effects on synaptic plasticity.

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Caffeine has cognitive‐enhancing properties with effects on learning and memory, concentration, arousal and mood. These effects imply changes at circuital and synaptic level, but the mechanism by which caffeine modifies synaptic plasticity remains elusive. Here we report that caffeine, at concentrations representing moderate to high levels of consumption in humans, induces an NMDA receptor‐independent form of LTP (CAFLTP) in the CA1 region of the hippocampus by promoting calcium‐dependent secretion of BDNF, which subsequently activates TrkB‐mediated signaling required for the expression of CAFLTP. Our data include the novel observation that insulin receptor substrate 2 (IRS2) is phosphorylated during induction of CAFLTP, a process that requires cytosolic free Ca2+. Consistent with the involvement of IRS2 signals in caffeine‐mediated synaptic plasticity, phosphorylation of Akt (Ser473) in response to LTP induction is defective in Irs2−/− mice, demonstrating that these plasticity changes are associated with downstream targets of the phosphoinositide 3‐kinase (PI3K) pathway. These findings indicate that TrkB‐IRS2 signals are essential for activation of PI3K during the induction of LTP by caffeine.

SUMMARYPurpose: In this study, we explore the antiepileptic effects of flufenamic acid (FFA) in order to identify the cellular mechanisms that underlie the potential anticonvulsant properties of this nonsteroidal antiinflammatory compound. Methods: The mechanisms of FFA action were analyzed using an in vitro model in which epileptiform activity was induced in hippocampal slices by perfusion with 100 lM 4-aminopyridine (4-AP) added to a modified Mg 2+ -free solution. The activity of CA1 pyramidal neurons as well as the synaptic connection between CA3 and CA1 was monitored using extracellular and patch-clamp recordings. Results: Epileptiform activity was suppressed in hippocampal neurons by FFA at concentrations between 50 and 200 lM. Glutamatergic excitatory synaptic transmission was diminished by FFA without modifying recurrent c-aminobutyric acid (GABA)ergic synaptic inhibition. Several lines of evidence indicated that FFA did not decrease neurotransmitter release probability, implicating a postsynaptic mechanism of action. FFA also potently reduced neuronal excitability, but did not alter the amplitude, duration, or undershoot of action potentials. Conclusions: Our results suggest that FFA exerts an anticonvulsive effect on hippocampal pyramidal neurons by simultaneously decreasing glutamatergic excitatory synaptic activity and reducing neuronal excitability. Therefore, our study provides experimental evidence that FFA may represent an effective pharmacologic agent in the treatment of epilepsy in the mammalian central nervous system.

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