The use of neural progenitor cells (NPCs) is limited by the incomplete knowledge of the extracellular signals regulating their proliferation and survival. We report that cultured mouse NPCs express functional mGlu3 and mGlu5 metabotropic glutamate receptors. Pharmacological blockade of both receptors reduced NPC proliferation and survival, whereas activation of mGlu5 receptors substantially enhanced cell proliferation. Adult mice lacking mGlu5 receptors or treated with mGlu5 or mGlu3 receptor antagonists showed a dramatic reduction in the number of dividing neuroprogenitors present in the subventricular zone and in the dentate gyrus of the hippocampus. These data disclose a novel function of mGlu receptors and offer new potential strategies for the optimization of cell replacement therapy in neurodegenerative disorders.

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Expression of Dickkopf-1 (Dkk-1), a secreted protein that negatively modulates the Wnt pathway, was induced in the hippocampus of gerbils and rats subjected to transient global cerebral ischemia as well as in cultured cortical neurons challenged with an excitotoxic pulse. In ischemic animals, the temporal and regional pattern of Dkk-1 expression correlated with the profile of neuronal death, as assessed by Nissl staining and Dkk-1 immunostaining in adjacent hippocampal sections. Treatment of ischemic animals with either Dkk-1 antisense oligonucleotides or lithium ions (which rescue the Wnt pathway acting downstream of the Dkk-1 blockade) protected vulnerable hippocampal neurons against ischemic damage. The same treatments protected cultured cortical neurons against NMDA toxicity. We conclude that induction of Dkk-1 with the ensuing inhibition of the canonical Wnt signaling pathway is required for the development of ischemic and excitotoxic neuronal death.

Methamphetamine (METH) is abused worldwide, and it represents a threat for public health. METH exposure induces a variety of detrimental effects. In fact, METH produces a number of oxidative species, which lead to lipid peroxidation, protein misfolding, and nuclear damage. Cell clearing pathways such as ubiquitin-proteasome (UP) and autophagy (ATG) are involved in METH-induced oxidative damage. Although these pathways were traditionally considered to operate as separate metabolic systems, recent studies demonstrate their interconnection at the functional and biochemical level. Very recently, the convergence between UP and ATG was evidenced within a single organelle named autophagoproteasome (APP), which is suppressed by mTOR activation. In the present research study, the occurrence of APP during METH toxicity was analyzed. In fact, coimmunoprecipitation indicates a binding between LC3 and P20S particles, which also recruit p62 and alpha-synuclein. The amount of METH-induced toxicity correlates with APP levels. Specific markers for ATG and UP, such as LC3 and P20S in the cytosol, and within METH-induced vacuoles, were measured at different doses and time intervals following METH administration either alone or combined with mTOR modulators. Western blotting, coimmunoprecipitation, light microscopy, confocal microscopy, plain transmission electron microscopy, and immunogold staining were used to document the effects of mTOR modulation on METH toxicity and the merging of UP with ATG markers within APPs. METH-induced cell death is prevented by mTOR inhibition, while it is worsened by mTOR activation, which correlates with the amount of autophagoproteasomes. The present data, which apply to METH toxicity, are also relevant to provide a novel insight into cell clearing pathways to counteract several kinds of oxidative damage.

Summary:  Inhibition of the Wnt pathway by the secreted glycoprotein, Dickkopf‐1 (Dkk‐1) has been related to processes of excitotoxic and ischemic neuronal death. We now report that Dkk‐1 is induced in neurons of the rat olfactory cortex and hippocampus degenerating in response to seizures produced by systemic injection of kainate (12 mg/kg, i.p.). There was a tight correlation between Dkk‐1 expression and neuronal death in both regions, as shown by the different expression profiles in animals classified as “high” and “low” responders to kainate. For example, no induction of Dkk‐1 was detected in the hippocampus of low responder rats, in which seizures did not cause neuronal loss. Induction of Dkk‐1 always anticipated neuronal death and was associated with a reduction in nuclear levels of β‐catenin, which reflects an ongoing inhibition of the canonical Wnt pathway. Intracerebroventricular injections of Dkk‐1 antisense oligonucleotides (12 nmol/2 μL) substantially reduced kainate‐induced neuronal damage, as did a pretreatment with lithium ions (1 mEq/kg, i.p.), which rescue the Wnt pathway by acting downstream of the Dkk‐1 blockade. Taken collectively, these data suggest that an early inhibition of the Wnt pathway by Dkk‐1 contributes to neuronal damage associated with temporal lobe epilepsy. We also examined Dkk‐1 expression in the hippocampus of epileptic patients and their controls. A strong Dkk‐1 immunolabeling was found in six bioptic samples and in one autoptic sample from patients with mesial temporal lobe epilepsy associated with hippocampal sclerosis. Dkk‐1 expression was undetectable or very low in autoptic samples from nonepileptic patients or in bioptic samples from patients with complex partial seizures without neuronal loss and/or reactive gliosis in the hippocampus. Our data raise the attractive possibility that drugs able to rescue the canonical Wnt pathway, such as Dkk‐1 antagonists or inhibitors of glycogen synthase kinase‐3β, reduce the development of hippocampal sclerosis in patients with temporal lobe epilepsy.

Inhibition of the canonical Wnt pathway has been implicated in the pathophysiology of neuronal death. Here, we report that the secreted Wnt antagonist, Dickkopf-1 (Dkk-1) is rapidly induced in neurons after induction of focal brain ischemia. In rats undergoing transient focal ischemia in response to brain infusion of endothelin-1, Dkk-1 was induced in neurons of the ischemic core and the penumbra region. Induction of Dkk-1 was associated with a reduced expression of b-catenin (a downstream signaling molecule of the canonical Wnt pathway), and was not observed in neurons expressing the protective protein, heat shock protein-70. Treatment with lithium ions, which, inter alia, rescue the canonical Wnt pathway, was highly protective against ischemic damage. Dkk-1 was also induced in cortical neurons of mice undergoing permanent middle cerebral artery (MCA) occlusion. This model allowed us to compare wild-type mice with doubleridge mice, which are characterized by a reduced expression of Dkk-1. Doubleridge mice showed an attenuated reduction of b-catenin and a reduced infarct volume in response to MCA occlusion, providing a direct demonstration that Dkk-1 contributes to the pathophysiology of ischemic neuronal damage. These data rise the interesting possibility that Dkk-1 antagonists or drugs that rescue the Wnt pathway might be neuroprotective in stroke.

We examined whether selective activation of mGlu4 metabotropic glutamate receptors attenuates 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced nigrostriatal damage in mice. C57BL mice were treated with a single dose of MPTP (30 mg/kg, i.p.) preceded, 30 min earlier, by a systemic injection of the mGlu4 receptor enhancer N-phenyl-7-(hydroxyimino)cyclopropa[b]chromen-1a-carboxamide (PHCCC). PHCCC was injected either subcutaneously in cremophor EL or intraperitoneally in saline containing 50% DMSO. PHCCC treatment (3 or 10 mg/kg) significantly reduced MPTP toxicity, as assessed by measurements of the striatal levels of dopamine and its metabolites and by tyrosine hydroxylase, dopamine transporter, and glial fibrillary acidic protein immunostaining in the corpus striatum and substantia nigra. In another set of experiments, a higher cumulative dose of MPTP (80 mg/kg divided into four injections with 2 h of interval) was injected to mGlu4 Ϫ/Ϫ mice and their Sv129/CD1 wild-type controls. A higher dose was used in these experiments because Sv129/CD1 mice are less sensitive to MPTP toxicity. Systemic administration of PHCCC was protective in wild-type mice but failed to affect nigrostriatal damage in mGlu4 Ϫ/Ϫ mice. Finally, unilateral infusion of PHCCC in the external globus pallidus protected the ipsilateral nigrostriatal pathway against MPTP toxicity. These data support the view that mGlu4 receptors are potential targets for the experimental treatment of parkinsonism.

T he pentraxin (PTX) family includes 2 subgroups of proteins that are structurally divided into short and long forms, are encoded by different genes, and are produced by different cells. [1][2][3] PTX3 is the prototype of the long PTX group, which differs from short PTXs by the presence of an unrelated long N-terminal domain giving it a different ligand recognition capacity. 4 PTX3 is highly conserved from mouse to humans (82% identical and 92% conserved amino acids) and is induced by primary inflammatory stimuli in a variety of cell types, 5,6 including mononuclear phagocytes, fibroblasts, adipocytes, dendritic, and endothelial and smooth muscle cells. 5,7 Marked as an innate immunity protein, PTX3 not only regulates inflammatory responses but also is involved in a range of important biological mechanisms, including vascular pathology. In fact, blood vessels produce large amounts of PTX3 during inflammation, and the level of circulating PTX3 increases in several pathological conditions affecting the cardiovascular system. 8,9 Moreover, in advanced atherosclerotic lesions and in patients with vasculitis, the protein is abundantly present in endothelial cells. 10,11 Thus, PTX3 seems to be a rapid marker of primary local innate immunity and inflammation activation and a novel diagnostic tool for vascular disorders. Clinical Perspective on p 1505In addition, high plasma PTX3 has been linked to vascular endothelial dysfunction in several human diseases, including Background-Pentraxin 3 (PTX3), the prototype of long pentraxins, has been described to be associated with endothelial dysfunction in different cardiovascular disorders. No study has yet evaluated the possible direct effect of PTX3 on vascular function. Methods and Results-Through in vitro experiments of vascular reactivity and ultrastructural analyses, we demonstrate that PTX3 induces dysfunction and morphological changes in the endothelial layer through a P-selectin/matrix metalloproteinase-1 pathway. The latter hampered the detachment of endothelial nitric oxide synthase from caveolin-1, leading to an impairment of nitric oxide signaling. In vivo studies showed that administering PTX3 to wild-type mice induced endothelial dysfunction and increased blood pressure, an effect absent in P-selectin-deficient mice. In isolated human umbilical vein endothelial cells, PTX3 significantly blunted nitric oxide production through the matrix metalloproteinase-1 pathway. Finally, using ELISA, we found that hypertensive patients (n=31) have higher plasma levels of PTX3 and its mediators P-selectin and matrix metalloproteinase-1 than normotensive subjects (n=21). Conclusions-Our data show for the first time a direct role of PTX3 on vascular function and blood pressure homeostasis, identifying the molecular mechanisms involved. The findings in humans suggest that PTX3, P-selectin, and matrix metalloproteinase-1 may be novel biomarkers that predict the onset of vascular dysfunction in hypertensive patients. Here, we demonstrate that (1) PTX3 induces vascular dysfuncti...