SummarySynaptic loss is the best pathological correlate of the cognitive decline in Alzheimer's Disease; yet, the molecular mechanisms underlying synaptic failure are unknown. Here we report a non-apoptotic baseline caspase-3 activity in hippocampal dendritic spines, and an enhancement of this activity at the onset of memory decline in the Tg2576-APPswe mouse model of Alzheimer's Disease. We show that, in spines, caspase-3 activates calcineurin which, in turn, triggers dephosphorylation and removal of the GluR1 subunit of AMPA-type receptor from post-synaptic sites. These molecular modifications lead to alterations of glutamatergic synaptic transmission and plasticity, and correlate with spine degeneration and a deficit in hippocampal-dependent memory. Importantly, pharmacological inhibition of caspase-3 activity in Tg2576 mice rescues the observed Alzheimerlike phenotypes. Therefore, we identify a novel caspase-3-dependent mechanism driving synaptic failure and contributing to cognitive dysfunction in Alzheimer's Disease. These findings point to caspase-3 as possible avenues for pharmacological therapy during early disease stages.Episodic hippocampal-dependent memory loss, the earliest clinical sign of Alzheimer's disease, is thought to be due to changes in synaptic function rather than neuronal loss 1,2 . Specifically, functional brain imaging studies revealed hippocampal mild abnormalities prior to clinical diagnosis and in the absence of structural brain atrophy, suggesting an altered functional connectivity of hippocampus at early stages of disease [3][4][5] .Dendritic spines are likely to be the first affected synaptic elements during early cognitive decline 6 . This is supported by several lines of evidence, such as: i) hippocampal spine-mediated plasticity underlies learning and memory 7 ; ii) post-mortem hippocampus from Alzheimer patients shows a significant decrease in dendritic spine density compared to age-matched controls 8 and iii) transgenic 3 mice expressing mutated forms of the amyloid precursor protein (APP), associated with familial Alzheimer's Disease, show age-dependent reductions in spine density, prior to plaque deposition 9 .Nevertheless, the molecular link between APP mutations triggering Alzheimer's Disease, and the occurrence of early spine loss remains elusive. Interestingly, a localized caspase-3 activity, causing synaptic failure, has been observed in vitro 10 , but the molecular mechanism linking caspase-3 activity to synaptic loss is far from being elucidated.Here, we analyzed caspase-3 activity in hippocampal synapses of the Tg2576 transgenic mouse model, harboring the human APPswe mutant allele linked to familial Alzheimer's Disease 11 . These mice develop early synaptic deficits 12 and several neuropathological features at older age, including amyloid plaques and dystrophic neurites 13 . Although Tg2576 mice lack neurofibrillary tangles and significant neuronal loss 14 , there is strong evidence that accumulation of the amyloid-β (Aβ) peptide, derived via APP proteolysis, is r...

Adult neurogenesis in the dentate gyrus plays a critical role in hippocampus-dependent spatial learning. It remains unknown, however, how new neurons become functionally integrated into spatial circuits and contribute to hippocampus-mediated forms of learning and memory. To investigate these issues, we used a mouse model in which the differentiation of adult-generated dentate gyrus neurons can be anticipated by conditionally expressing the pro-differentiative gene PC3 (Tis21/BTG2) in nestin-positive progenitor cells. In contrast to previous studies that affected the number of newly generated neurons, this strategy selectively changes their timing of differentiation. New, adult-generated dentate gyrus progenitors, in which the PC3 transgene was expressed, showed accelerated differentiation and significantly reduced dendritic arborization and spine density. Functionally, this genetic manipulation specifically affected different hippocampus-dependent learning and memory tasks, including contextual fear conditioning, and selectively reduced synaptic plasticity in the dentate gyrus. Morphological and functional analyses of hippocampal neurons at different stages of differentiation, following transgene activation within defined time-windows, revealed that the new, adult-generated neurons up to 3–4 weeks of age are required not only to acquire new spatial information but also to use previously consolidated memories. Thus, the correct unwinding of these key memory functions, which can be an expression of the ability of adult-generated neurons to link subsequent events in memory circuits, is critically dependent on the correct timing of the initial stages of neuron maturation and connection to existing circuits.

The role of autophagy in cancer onset and progression appears still controversial. On one hand, autophagy allows cancer cell to survive in unfavorable environmental conditions, on the other hand, once internal energy resources are exhausted, it leads to cell death. In addition, autophagy interpheres with cell cycle progression, de facto exerting a cytostatic activity. Hence, it represents an important target for anticancer therapy. For example, temozolomide (TMZ), of use for glioblastoma (GBM) treatment, appears as capable of inducing autophagy partially inhibiting cancer cell proliferation. However, GBM, a very aggressive brain tumor with poor prognosis even after surgery and radio-chemotherapy, invariably recurs and leads to patient death. Since cancer stem cells have been hypothesized to play a role in refractory/relapsing cancers, in the present work we investigated if autophagy could represent a constitutive cytoprotection mechanism for glioblastoma stem-like cells (GSCs) and if the modulation of autophagic process could affect GBM growth and survival. Thus, in the present study we first evaluated the relevance of autophagy in GBM tumor specimens, then its occurrence in GSCs and, finally, if modulation of autophagy could influence GSC response to TMZ. Our results suggested that, in vitro, the impairing autophagic process with quinacrine, a compound able to cross the blood-brain barrier, increased GSC susceptibility to TMZ. Death of GSCs was apparently due to the iron dependent form of programmed cell death characterized by the accumulation of lipid peroxides called ferroptosis. These results underscore the relevance of the modulation of autophagy in the GSC survival and death and suggest that triggering of ferroptosis in GSCs could represent a novel and important target for the management of glioblastoma.

Control of pyramidal neuron excitability is vital for the functioning of the neocortex. Somatodendritic slow self-inhibition (SSI) allows inhibitory neurons to regulate their own activity, but the existence of similar mechanisms in excitatory cells has not been shown. We found that in rodents endocannabinoids mediated SSI and long-term modulation of inhibitory connections in layer 2/3 pyramidal neurons with a distinct dendritic morphology, suggesting that a glutamatergic network in cortical circuits is self-regulated.

Long-term potentiation of inhibitory GABAergic transmission controls synaptic integration and action potential generation of specific neocortical neurons.

In the CNS, endocannabinoids are identified mainly as two endogenous lipids: anandamide, the ethanolamide of arachidonic acid, and 2-arachidonoylglycerol (2-AG). Endocannabinoids are known to inhibit transmitter release from presynaptic terminals; however we have recently demonstrated that they are also involved in slow self-inhibition (SSI) of layer V low-threshold spiking (LTS) interneurons in rat somatosensory cortex. SSI is induced by repetitive firing in LTS cells, which can express either cholecystokinin or somatostatin. SSI is triggered by an endocannabinoid-dependent activation of a prolonged somatodendritic K ϩ conductance and associated hyperpolarization in the same cell. The synthesis of both endocannabinoids is dependent on elevated [Ca 2ϩ ] i such as occurs during sustained neuronal activity. To establish whether 2-AG mediates autocrine LTS-SSI, we blocked its biosynthesis from phospholipase C (PLC) and diacylglycerol lipases (DAGLs). Current-clamp recordings from LTS interneurons in acute neocortical slices showed that inclusion of DAGL inhibitors in the whole-cell pipette prevented the long-lasting hyperpolarization triggered by LTS cell repetitive firing. Similarly, extracellular applications of a PLC inhibitor prevented SSI in LTS interneurons. Moreover, metabotropic glutamate receptor-dependent activation of PLC produced a long-lasting hyperpolarization which was prevented by the CB1 antagonist AM251, as well as by PLC and DAGL inhibitors. The loss of SSI in the presence of intracellular DAGL blockers confirms that endocannabinoid production occurs in the same interneuron undergoing the persistent hyperpolarization. Since DAGLs produce no endocannabinoid other than 2-AG, these results identify this compound as the autocrine mediator responsible for the postsynaptic slow self-inhibition of neocortical LTS interneurons.

BackgroundMesenchymal stem/stromal cells (MSCs) represent an attractive tool for cell-based cancer therapy mainly because of their ability to migrate to tumors and to release bioactive molecules. However, the impact of MSCs on tumor growth has not been fully established. We previously demonstrated that murine MSCs show a strong tropism towards glioblastoma (GBM) brain xenografts and that these cells are able to uptake and release the chemotherapeutic drug paclitaxel (PTX), maintaining their tropism towards the tumor. Here, we address the therapy-relevant issue of using MSCs from human donors (hMSCs) for local or systemic administration in orthotopic GBM models, including xenografts of patient-derived glioma stem cells (GSCs).MethodsU87MG or GSC1 cells expressing the green fluorescent protein (GFP) were grafted onto the striatum of immunosuppressed rats. Adipose hMSCs (Ad-hMSCs), fluorescently labeled with the mCherry protein, were inoculated adjacent to or into the tumor. In rats bearing U87MG xenografts, systemic injections of Ad-hMSCs or bone marrow (BM)-hMSCs were done via the femoral vein or carotid artery. In each experiment, either PTX-loaded or unloaded hMSCs were used. To characterize the effects of hMSCs on tumor growth, we analyzed survival, tumor volume, tumor cell proliferation, and microvascular density.ResultsOverall, the AD-hMSCs showed remarkable tropism towards the tumor. Intracerebral injection of Ad-hMSCs significantly improved the survival of rats with U87MG xenografts. This effect was associated with a reduction in tumor growth, tumor cell proliferation, and microvascular density. In GSC1 xenografts, intratumoral injection of Ad-hMSCs depleted the tumor cell population and induced migration of resident microglial cells. Overall, PTX loading did not significantly enhance the antitumor potential of hMSCs. Systemically injected Ad- and BM-hMSCs homed to tumor xenografts. The efficiency of hMSC homing ranged between 0.02 and 0.5% of the injected cells, depending both on the route of cell injection and on the source from which the hMSCs were derived. Importantly, systemically injected PTX-loaded hMSCs that homed to the xenograft induced cytotoxic damage to the surrounding tumor cells.ConclusionshMSCs have a therapeutic potential in GBM brain xenografts which is also expressed against the GSC population. In this context, PTX loading of hMSCs seems to play a minor role.Electronic supplementary materialThe online version of this article (doi:10.1186/s13287-017-0516-3) contains supplementary material, which is available to authorized users.

Tumors exhibit complex organization and contain a variety of cell populations. The realization that the regenerative properties of a tumor may be largely confined to a cell subpopulation (cancer stem cell) is driving a new era of anti-cancer research. Cancer stem cells from Glioblastoma Multiforme tumors express markers that are also expressed in non-cancerous neural stem cells, including nestin and Sox2. We previously showed that the transcription factor Hes3 is a marker of neural stem cells, and that its expression is inhibited by JAK activity. Here we show that Hes3 is also expressed in cultures from glioblastoma multiforme which express neural stem cell markers, can differentiate into neurons and glia, and can recapitulate the tumor of origin when transplanted into immunocompromised mice. Similar to observations in neural stem cells, JAK inhibits Hes3 expression. Hes3 RNA interference reduces the number of cultured glioblastoma cells suggesting a novel therapeutic strategy.