Glycine receptors are involved in hippocampal neuronal damage caused by oxygen-glucose deficiency.

Glycine receptors (GlyRs) belong to the family of ligand-gated cys-loop receptors and effectuate fast inhibitory neurotransmission in central nervous system (CNS). They are involved in numerous physiological processes, such as movement, respiration, and processing of sensory information, as well as in regulation of neuronal excitability in different brain regions. GlyRs play important role in the maintenance of excitatory/inhibitory balance in the hippocampus and participate in the development of various brain pathologies. In the present study, we have examined a surface expression of GlyRs by pyramidal neurons and astrocytes in control and after 30 min of oxygen-glucose deprivation (OGD) in the organotypic culture of hippocampal slices. Our investigation has demonstrated a decrease in GlyR-positive staining associated with pyramidal neurons and relative stability of GlyRs expression at the surface of astrocytes 4 hs after OGD. These data indicate that GlyRs dysfunction may represent a significant additional factor leading to enhanced neuronal damage induced by OGD. Pharmacological modulation of GlyRs is a promising venue of research for the correction of negative consequences of oxygen-glucose deficiency.

Cooperation of HIF- and NCAM-mediated mechanisms in cell viability of hippocampal cultures after oxygen-glucose deprivation.

Neurodegenerative diseases of different genesis are the result of cellular damages including those caused by oxygen and glucose deficit. Neuronal survival or death in brain pathologies depends on a variety of interrelated molecular mechanisms. A key role in modulation of neuron viability belongs to HIF (hypoxia-inducible factor) and NCAM (neural cell adhesion molecules) signaling pathways. In this work, we used organotypic and dissociated hippocampal cultures to analyze cell viability and HIF-1α immunopositive (HIF-1α+ ) signal after 30 min oxygen-glucose deprivation (OGD) followed by 24 h of reoxygenation in the presence of FGL (synthetic NCAM-derived mimetic peptide). According to LDH- and MTS-assay of cell viability, FGL showed a neuroprotective effect, which was attributed to the association with FGFR. We showed that these effects correlated with changes of the HIF-1α+ level suggesting the communications of HIF and NCAM signaling pathways. These data extend our knowledge of neurodegeneration mechanisms and open additional potential for the development of neuroprotection strategies.

mTOR/α-ketoglutarate signaling: impact on brain cell homeostasis under ischemic conditions.

The multifunctional molecules mechanistic target of rapamycin (mTOR) and α-ketoglutarate (αKG) are crucial players in the regulatory mechanisms that maintain cell homeostasis in an ever-changing environment. Cerebral ischemia is associated primarily with oxygen-glucose deficiency (OGD) due to circulatory disorders. Upon exceeding a threshold of resistance to OGD, essential pathways of cellular metabolism can be disrupted, leading to damage of brain cells up to the loss of function and death. This mini-review focuses on the role of mTOR and αKG signaling in the metabolic homeostasis of brain cells under OGD conditions. Integral mechanisms concerning the relative cell resistance to OGD and the molecular basis of αKG-mediated neuroprotection are discussed. The study of molecular events associated with cerebral ischemia and endogenous neuroprotection is relevant for improving the effectiveness of therapeutic strategies.

mTOR/α-ketoglutarate-mediated signaling pathways in the context of brain neurodegeneration and neuroprotection.

Cerebral disorders are largely associated with impaired cellular metabolism, despite the regulatory mechanisms designed to ensure cell viability and adequate brain function. Mechanistic target of rapamycin (mTOR) signaling is one of the most crucial factors in the regulation of energy homeostasis and its imbalance is linked with a variety of neurodegenerative diseases. Recent advances in the metabolic pathways' modulation indicate the role of α-ketoglutarate (AKG) as a major signaling hub, additionally highlighting its anti-aging and neuroprotective properties, but the mechanisms of its action are not entirely clear. In this review, we analyzed the physiological and pathophysiological aspects of mTOR in the brain. We also discussed AKG's multifunctional properties, as well as mTOR/AKG-mediated functional communications in cellular metabolism. Thus, this article provides a broad overview of the mTOR/AKG-mediated signaling pathways, in the context of neurodegeneration and endogenous neuroprotection, with the aim to find novel therapeutic strategies.

Mesenchymal stem cell application for treatment of neuroinflammation-induced cognitive impairment in mice.

Background: The present research has been undertaken to study the therapeutic potential of mesenchymal stem cells (MSCs) for the treatment of neuroinflammation-induced cognitive disorders. Methods: Either umbilical cord or adipose MSCs were injected into mice treated with lipopolysaccharide. The mice were studied in behavioral tests, and their brains were examined by means of immunohistochemistry, electron microscopy and sandwich ELISA. Results: MSCs, introduced either intravenously or intraperitoneally, restored episodic memory of mice disturbed by inflammation, normalized nAChR and Aß1-42 levels and stimulated proliferation of neural progenitor cells in the brain. The effect of MSCs was observed for months, whereas that of MSC-conditioned medium was transient and stimulated an immune reaction. SDF-1α potentiated the effects of MSCs on the brain and memory. Conclusion: MSCs of different origins provide a long-term therapeutic effect in the treatment of neuroinflammation-induced episodic memory impairment.

Cardiac-specific ß-catenin deletion dysregulates energetic metabolism and mitochondrial function in perinatal cardiomyocytes.

ß-Catenin signaling pathway regulates cardiomyocytes proliferation and differentiation, though its involvement in metabolic regulation of cardiomyocytes remains unknown. We used one-day-old mice with cardiac-specific knockout of ß-catenin and neonatal rat ventricular myocytes treated with ß-catenin inhibitor to investigate the role of ß-catenin metabolism regulation in perinatal cardiomyocytes. Transcriptomics of perinatal ß-catenin-ablated hearts revealed a dramatic shift in the expression of genes involved in metabolic processes. Further analysis indicated an inhibition of lipolysis and glycolysis in both in vitro and in vivo models. Finally, we showed that ß-catenin deficiency leads to mitochondria dysfunction via the downregulation of Sirt1/PGC-1α pathway. We conclude that cardiac-specific ß-catenin ablation disrupts the energy substrate shift that is essential for postnatal heart maturation, leading to perinatal lethality of homozygous ß-catenin knockout mice.

Adipose-Derived Stem Cells Reduce Lipopolysaccharide-Induced Myelin Degradation and Neuroinflammatory Responses of Glial Cells in Mice.

Brain inflammation is a key event triggering the pathological process associated with many neurodegenerative diseases. Current personalized medicine and translational research in neurodegenerative diseases focus on adipose-derived stem cells (ASCs), because they are patient-specific, thereby reducing the risk of immune rejection. ASCs have been shown to exert a therapeutic effect following transplantation in animal models of neuroinflammation. However, the mechanisms by which transplanted ASCs promote cell survival and/or functional recovery are not fully understood. We investigated the effects of ASCs in in vivo and in vitro lipopolysaccharide (LPS)-induced neuroinflammatory models. Brain damage was evaluated immunohistochemically using specific antibody markers of microglia, astroglia and oligodendrocytes. ASCs were used for intracerebral transplantation, as well as for non-contact co-culture with brain slices. In both in vivo and in vitro models, we found that LPS caused micro- and astroglial activation and oligodendrocyte degradation, whereas the presence of ASCs significantly reduced the damaging effects. It should be noted that the observed ASCs protection in a non-contact co-culture suggested that this effect was due to humoral factors via ASC-released biomodulatory molecules. However, further clinical studies are required to establish the therapeutic mechanisms of ASCs, and optimize their use as a part of a personalized medicine strategy.

Dynamic presenilin 1 and synaptotagmin 1 interaction modulates exocytosis and amyloid ß production.

BACKGROUND: Alzheimer's disease (AD)-linked protein, presenilin 1 (PS1), is present at the synapse, and the knock-out of presenilin in mice leads to synaptic dysfunction. On the other hand, synaptic activity was shown to influence PS1-dependent generation of distinct amyloid ß (Aß) species. However, the precise nature of these regulations remains unclear. The current study reveals novel role of PS1 at the synapse, and deciphers how PS1 and synaptic vesicle-associated protein, synaptotagmin 1 (Syt1) modulate each other functions in neurons via direct activity-triggered interaction. Additionally, the therapeutic potential of fostering PS1-Syt1 binding is investigated as a synapse-specific strategy for AD prevention. METHODS: PS1-based cell-permeable peptide targeting PS1-Syt1 binding site was designed to inhibit PS1-Syt1 interaction in neurons. PS1 conformation, synaptic vesicle exocytosis and trafficking were assayed by fluorescence lifetime imaging microscopy (FLIM), glutamate release/synaptopHluorin assay, and fluorescence recovery after photobleaching, respectively. Syt1 level and interaction with PS1 in control and sporadic AD brains were determined by immunohistochemistry and FLIM. AAV-mediated delivery of Syt1 into mouse hippocampi was used to investigate the therapeutic potential of strengthening PS1-Syt1 binding in vivo. Statistical significance was determined using two-tailed unpaired Student's t-test, Mann-Whitney's U-test or two-way ANOVA followed by a Bonferroni's post-test. RESULTS: We demonstrate that targeted inhibition of the PS1-Syt1 binding in neurons, without changing the proteins' expression level, triggers "pathogenic" conformational shift of PS1, and consequent increase in the Aß42/40 ratio. Moreover, our data indicate that PS1, by binding directly to Syt1, regulates synaptic vesicle trafficking and facilitates exocytosis and neurotransmitter release. Analysis of human brain tissue revealed that not only Syt1 levels but also interactions between remaining Syt1 and PS1 are diminished in sporadic AD. On the other hand, overexpression of Syt1 in mouse hippocampi was found to potentiate PS1-Syt1 binding and promote "protective" PS1 conformation. CONCLUSIONS: The study reports novel functions of PS1 and Syt1 at the synapse, and demonstrates the importance of PS1-Syt1 binding for exocytosis and safeguarding PS1 conformation. It suggests that reduction in the Syt1 level and PS1-Syt1 interactions in AD brain may present molecular underpinning of the pathogenic PS1 conformation, increased Aß42/40 ratio, and impaired exocytosis.

HIF-1α-mediated upregulation of SERCA2b: The endogenous mechanism for alleviating the ischemia-induced intracellular Ca(2+) store dysfunction in CA1 and CA3 hippocampal neurons.

Pyramidal neurons of the hippocampus possess differential susceptibility to the ischemia-induced damage with the highest vulnerability of CA1 and the lower sensitivity of CA3 neurons. This damage is triggered by Ca(2+)-dependent excitotoxicity and can result in a delayed cell death that might be potentially suspended through activation of endogenous neuroprotection with the hypoxia-inducible transcription factors (HIF). However, the molecular mechanisms of this neuroprotection remain poorly understood. Here we show that prolonged (30min) oxygen and glucose deprivation (OGD) in situ impairs intracellular Ca(2+) regulation in CA1 rather than in CA3 neurons with the differently altered expression of genes coding Ca(2+)-ATPases: the mRNA level of plasmalemmal Ca(2+)-ATPases (PMCA1 and PMCA2 subtypes) was downregulated in CA1 neurons, whereas the mRNA level of the endoplasmic reticulum Ca(2+)-ATPases (SERCA2b subtype) was increased in CA3 neurons at 4h of re-oxygenation after prolonged OGD. These demonstrate distinct susceptibility of CA1 and CA3 neurons to the ischemic impairments in intracellular Ca(2+) regulation and Ca(2+)-ATPase expression. Stabilization of HIF-1α by inhibiting HIF-1α hydroxylation prevented the ischemic decrease in both PMCA1 and PMCA2 mRNAs in CA1 neurons, upregulated the SERCA2b mRNA level and eliminated the OGD-induced Ca(2+) store dysfunction in these neurons. Cumulatively, these findings reveal the previously unknown HIF-1α-driven upregulation of Ca(2+)-ATPases as a mechanism opposing the ischemic impairments in intracellular Ca(2+) regulation in hippocampal neurons. The ability of HIF-1α to modulate expression of genes coding Ca(2+)-ATPases suggests SERCA2b as a novel target for HIF-1 and may provide potential implications for HIF-1α-stabilizing strategy in activating endogenous neuroprotection.

Brief anoxia preconditioning and HIF prolyl-hydroxylase inhibition enhances neuronal resistance in organotypic hippocampal slices on model of ischemic damage.

It is well known that a brief anoxia or hypoxia episodes can render brain resistant to a subsequent ischemia. Recent investigations indicate that mechanisms of such stimulated endogenous neuroprotection are related to the family of hypoxia-inducible factors (HIF), however there are still little data available on the role of HIF family members in hippocampus-a brain structure, highly sensitive to oxygen deficiency. We have used the model of cultured hippocampal slices and single-cell quantitative RT-PCR to study HIF-1α and HIF-3α mRNA expression following triple 5-min mild anoxia, 30-min oxygen-glucose deprivation and their combination. We also tested the effects of HIF prolyl-hydroxylase inhibition with 2,4-pyridinedicarboxylic acid diethyl ester pre-treatment followed by a 30-min oxygen-glucose deprivation. It was found that neuronal damage induced by oxygen-glucose deprivation was accompanied by a significant decrease in both HIF-1α and HIF-3α mRNA levels in CA1 but not CA3 neurons. Anoxia preconditioning did not affect cell viability and HIF mRNA levels but applied before oxygen-glucose deprivation prevented neuronal damage and suppression of HIF-1α and HIF-3α mRNA expression. It was also found that effects of the prolyl-hydroxylase inhibitor were similar to anoxia preconditioning. These results suggest that anoxia preconditioning increases anti-ischemic neuronal resistance which to a certain extent correlates with the changes of HIF-1α and HIF-3α expression.

A synthetic NCAM-derived peptide, FGL, protects hippocampal neurons from ischemic insult both in vitro and in vivo.

There is a major unmet need for development of innovative strategies for neuroprotection against ischemic brain injury. Here we show that FGL, a neural cell adhesion molecule (NCAM)-derived peptide binding to and inducing phosphorylation of the fibroblast growth factor receptor (FGFR), acts neuroprotectively after an ischemic insult both in vitro and in vivo. The neuroprotective activity of FGL was tested in vitro on dissociated rat hippocampal neurons and hippocampal slice cultures, using a protocol of oxygen-glucose deprivation (OGD). FGL protected hippocampal neurons from damage and maintained or restored their metabolic and presynaptic activity, both if employed as a pretreatment alone to OGD, and if only applied after the insult. In vivo 24 h pretreatment with a single suboccipital injection of FGL significantly protected hippocampal CA1 neurons from death in a transient global ischemia model in the gerbil. We conclude that FGL promotes neuronal survival after ischemic brain injury.