Knowing that expression of metabotropic glutamate 2 (mGlu2) receptors in the dorsal root ganglia is regulated by acetylation mechanisms, we examined the effect of two selective and chemically ...unrelated histone deacetylase (HDAC) inhibitors, N-(2-aminophenyl)-4-N-(pyridine-3-ylmethoxy-carbonyl)aminomethylbenzamide (MS-275) and suberoylanilide hydroamic acid (SAHA), in a mouse model of persistent inflammatory pain. Although a single subcutaneous injection of MS-275 (3 mg/kg) or SAHA (5-50 mg/kg) was ineffective, a 5-day treatment with either of the two HDAC inhibitors substantially reduced the nociceptive response in the second phase of the formalin test, which reflects the development of central sensitization in the dorsal horn of the spinal cord. Analgesia was abrogated by a single injection of the mGlu2/3 receptor antagonist (alphaS)-alpha-amino-alpha-(1S,2S)-2-carboxycyclopropyl-9H-xantine-9-propanoic acid (LY341495; 1 mg/kg, i.p.), which was inactive per se. Both MS-275 and SAHA up-regulated the expression of mGlu2 receptors in the dorsal root ganglion (DRG) and spinal cord under conditions in which they caused analgesia, without changing the expression of mGlu1a, mGlu4, or mGlu5 receptors. Induction of DRG mGlu2 receptors in response to SAHA was associated with increased acetylation of p65/RelA on lysine 310, a process that enhances the transcriptional activity of p65/RelA at nuclear factor-kappaB-regulated genes. Transcription of the mGlu2 receptor gene is known to be activated by p65/RelA in DRG neurons. We conclude that HDAC inhibition produces analgesia by up-regulating mGlu2 receptor expression in the DRG, an effect that results from the amplification of NF-kappaB transcriptional activity. These data provide the first evidence that HDAC inhibitors cause analgesia and suggest that HDACs are potential targets for the epigenetic treatment of pain.
In animals, sporadic injections of the mitochondrial toxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) selectively damage dopaminergic neurons but do not fully reproduce the features of human ...Parkinson's disease. We have now developed a mouse Parkinson's disease model that is based on continuous MPTP administration with an osmotic minipump and mimics many features of the human disease. Although both sporadic and continuous MPTP administration led to severe striatal dopamine depletion and nigral cell loss, we find that only continuous administration of MPTP produced progressive behavioral changes and triggered formation of nigral inclusions immunoreactive for ubiquitin and α-synuclein. Moreover, only continuous MPTP infusions caused long-lasting activation of glucose uptake and inhibition of the ubiquitin-proteasome system. In mice lacking α-synuclein, continuous MPTP delivery still induced metabolic activation, but induction of behavioral symptoms and neuronal cell death were almost completely alleviated. Furthermore, the inhibition of the ubiquitin-proteasome system and the production of inclusion bodies were reduced. These data suggest that continuous low-level exposure of mice to MPTP causes a Parkinson-like syndrome in an α-synucleindependent manner.
Locus Coeruleus (LC) is the main noradrenergic nucleus of the brain, which is involved in many physiological functions including cognition; its impairment may be crucial in the neurobiology of a ...variety of brain diseases. Locus Coeruleus-Magnetic Resonance Imaging (LC-MRI) allows to identify
in vivo
LC in humans. Thus, a variety of research teams have been using LC-MRI to estimate LC integrity in normal aging and in patients affected by neurodegenerative disorders, where LC integrity my work as a biomarker. A number of variations between LC-MRI studies exist, concerning post-acquisition analysis and whether this had been performed within MRI native space or in
ad hoc
-built MRI template space. Moreover, the reproducibility and reliability of this tool is still to be explored. Therefore, in the present study, we analyzed a group of neurologically healthy, cognitively intact elderly subjects, using both a native space- and a template space-based LC-MRI analysis. We found a good inter-method agreement, particularly considering the LC Contrast Ratio. The template space-based approach provided a higher spatial resolution, lower operator-dependency, and allowed the analysis of LC topography. Our
ad hoc
-developed LC template showed LC morphological data that were in line with templates published very recently. Remarkably, present data significantly overlapped with a recently published LC “metaMask”, that had been obtained by averaging the results of a variety of previous LC-MRI studies. Thus, such a template space-based approach may pave the way to a standardized LC-MRI analysis and to be used in future clinic–anatomical correlations.
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.
Increasing findings indicate that a dysfunction in the autophagy machinery is common during retinal degeneration. The present article provides evidence showing that an autophagy defect in the outer ...retinal layers is commonly described at the onset of retinal degeneration. These findings involve a number of structures placed at the border between the inner choroid and the outer retina encompassing the choriocapillaris, the Bruch's membrane, photoreceptors and Mueller cells. At the center of these anatomical substrates are placed cells forming the retinal pigment epithelium (RPE), where autophagy seems to play most of its effects. In fact, a failure of the autophagy flux is mostly severe at the level of RPE. Among various retinal degenerative disorders, age-related macular degeneration (AMD) is mostly affected by a damage to RPE, which can be reproduced by inhibiting the autophagy machinery and it can be counteracted by the activation of the autophagy pathway. In the present manuscript evidence is provided that such a severe impairment of retinal autophagy may be counteracted by administration of a number of phytochemicals, which possess a strong stimulatory activity on autophagy. Likewise, natural light stimulation administered in the form of pulsatile specific wavelengths is capable of inducing autophagy within the retina. This dual approach to stimulate autophagy is further strengthened by the interaction of light with phytochemicals which is shown to activate the chemical properties of these natural molecules in sustaining retinal integrity. The beneficial effects of photo-biomodulation combined with phytochemicals is based on the removal of toxic lipid, sugar and protein species along with the stimulation of mitochondrial turn-over. Additional effects of autophagy stimulation under the combined effects of nutraceuticals and light pulses are discussed concerning stimulation of retinal stem cells which partly correspond to a subpopulation of RPE cells.
This work represents a detailed methodological description of automated stereology dedicated to all brainstem catecholamine nuclei. Each tyrosine-hydroxylase-containing nucleus was analyzed to count ...the following features: i) nuclear volume; ii) neuron number per nucleus; iii) neuron area per each nucleus.A number of reports described catecholamine-containing neurons within brainstem of a variety of animal species. In a recently published work, we reported a simultaneous quantitative analysis of tyrosine hydroxylase-positive neurons in the whole brainstem. Here we report the detailed step by step stereological procedure which allowed to perform a morphometric assessment of each catecholamine nucleus. This protocol provides the method chance to analyze simultaneously various morphological features in the same experimental setting to avoid variability when single nuclei are analyzed in different experiments. This improves the reliability of comparisons between brainstem catecholamine nuclei within the reticular formation to increase our insight about the key functional roles played by these cells in the mammalian brain. In fact, despite being a discrete number of neurons scattered in a small brain area, these cells provide remarkable axonal collateralization which allows the modulation of neuronal activity in the entire CNS. The step by step description of brainstem stereology provided here is reported in order to share these methods and enhance quantitative studies about these fascinating nuclei. At the same time we aim to provide a tool to be used routinely when analyzing the morphology and physiology of brainstem catecholamine cells.
The cellular prion protein (PrPc) is physiologically expressed within selective brain areas of mammals. Alterations in the secondary structure of this protein lead to scrapie-like prion protein ...(PrPsc), which precipitates in the cell. PrPsc has been detected in infectious, inherited or sporadic neurodegenerative disorders. Prion protein metabolism is dependent on autophagy and ubiquitin proteasome. Despite not being fully elucidated, the physiological role of prion protein relates to chaperones which rescue cells under stressful conditions.Methamphetamine (METH) is a widely abused drug which produces oxidative stress in various brain areas causing mitochondrial alterations and protein misfolding. These effects produce a compensatory increase of chaperones while clogging cell clearing pathways. In the present study, we explored whether METH administration modifies the amount of PrPc. Since high levels of PrPc when the clearing systems are clogged may lead to its misfolding into PrPsc, we further tested whether METH exposure triggers the appearance of PrPsc. We analysed the effects of METH and dopamine administration in PC12 and striatal cells by using SDS-PAGE Coomassie blue, immune- histochemistry and immune-gold electron microscopy. To analyze whether METH administration produces PrPsc aggregates we used antibodies directed against PrP following exposure to proteinase K or sarkosyl which digest folded PrPc but misfolded PrPsc. We fond that METH triggers PrPsc aggregates in DA-containing cells while METH is not effective in primary striatal neurons which do not produce DA. In the latter cells exogenous DA is needed to trigger PrPsc accumulation similarly to what happens in DA containing cells under the effects of METH. The present findings, while fostering novel molecular mechanisms involving prion proteins, indicate that, cell pathology similar to prion disorders can be mimicked via a DA-dependent mechanism by a drug of abuse.
Mutation of genes encoding for various components of a metabolic pathway named the ubiquitin-proteasome system (UP) leads to inherited forms of Parkinson's disease (PD), whereas various components of ...the UP are constantly present within neuronal inclusions, Lewy bodies, that characterize most genetic and sporadic forms of PD. It has been hypothesized that impairment of this metabolic pathway might be a common mechanism for the onset of PD, and a recent study demonstrated a dysfunction of the UP system within the substantia nigra of patients affected by sporadic PD. In search for the mechanisms underlying the selective toxicity for nigral neurons after inhibition of the UP system, we explored the selective effects after striatal microinfusions of lactacystin or epoxomycin and potential retrograde changes within the ipsilateral substantia nigra. We found that neurotoxicity was selective for striatal dopamine (DA) components and led to retrograde apoptosis within nigral DA cells, which developed neuronal inclusions staining for antigens of the UP system. We found the same ultrastructural features characterizing inclusions obtained in vivo and in vitro after UP inhibition. In vivo, lactacystin-epoxomycin-induced toxicity was suppressed by inhibiting DA synthesis. Similarly, in vitro inclusions and apoptosis were prevented by reducing endogenous DA. On the other hand, toxicity of proteasome inhibition was enhanced by drugs augmenting DA availability: l-3,4-dihydroxyphenylalanine, monoamine oxidase blockers, and DA beta-hydroxylase blockers. These findings demonstrate that impairment of the UP system produces cell death and neuronal inclusions selectively for DA-containing neurons that depend on the occurrence of endogenous DA.
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.
Abstract Methamphetamine produces locomotor activation and typical stereotyped motor patterns, which are commonly related with increased catecholamine activity within the basal ganglia, including the ...dorsal and ventral striatum. Since the cerebellum is critical for movement control, and for learning of motor patterns, we hypothesized that cerebellar catecholamines might be a target of methamphetamine. To test this experimental hypothesis we injected methamphetamine into C57 Black mice at the doses of 5 mg/kg two or three times, 2 h apart. This dosing regimen is known to be toxic for striatal dopamine terminals. However, we found that in the cerebellum, methamphetamine increased the expression of the primary transcript of the tyrosine hydroxylase (TH) gene, followed by an increased expression of the TH protein. Increased TH was localized within Purkinje cells, where methamphetamine increased the number of TH-immunogold particles, and produced a change in the distribution of the enzyme by increasing the cytoplasmic percentage. Increased TH expression was accompanied by a slight increase in noradrenaline content. This effect was highly site-specific for the cortex of posterior vermal lobules, while only slight effects were detectable in the hemispheres. The present data indicate that the cerebellum does represent a target of methamphetamine, which produces specific and fine alterations of the catecholamine system involving synthesis, amount, and compartmentalization of TH as well as increased noradrenaline levels. This may be relevant for motor alterations induced by methamphetamine. In line with this, inherited cerebellar movement disorders in various animal species including humans are associated with increased TH immunoreactivity within intrinsic neurons of the same lobules of the cerebellar cortex.
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