A Novel Diselenide-Probucol-Analogue Protects Against Methylmercury-Induced Toxicity in HT22 Cells by Upregulating Peroxide Detoxification Systems: a Comparison with Diphenyl Diselenide.

Methylmercury (MeHg) is a ubiquitous environmental neurotoxicant whose mechanisms of action involve oxidation of endogenous nucleophilic groups (mainly thiols and selenols), depletion of antioxidant defenses, and disruption of neurotransmitter homeostasis. Diphenyl diselenide-(PhSe)2-a model diaryl diselenide, has been reported to display significant protective effects against MeHg-induced neurotoxicity under both in vitro and in vivo experimental conditions. In this study, we compared the protective effects of (PhSe)2 with those of RC513 (4,4'-diselanediylbis(2,6-di-tert-butylphenol), a novel diselenide-probucol-analog) against MeHg-induced toxicity in the neuronal (hippocampal) cell line HT22. Although both (PhSe)2 and RC513 significantly mitigated MeHg- and tert-butylhydroperoxide (t-BuOOH)-cytotoxicity, the probucol analog exhibited superior protective effects, which were observed earlier and at lower concentrations compared to (PhSe)2. RC513 treatment (at either 0.5 µM or 2 µM) significantly increased glutathione peroxidase (GPx) activity, which has been reported to counteract MeHg-toxicity. (PhSe)2 was also able to increase GPx activity, but only at 2 µM. Although both compounds increased the Gpx1 transcripts at 6 h after treatments, only RC513 was able to increase mRNA levels of Prx2, Prx3, Prx5, and Txn2, which are also involved in peroxide detoxification. RC513 (at 2 µM) significantly increased GPx-1 protein expression in HT22 cells, although (PhSe)2 displayed a minor (nonsignificant) effect in this parameter. In agreement, RC513 induced a faster and superior capability to cope with exogenously-added peroxide (t-BuOOH). In summary, when compared to the prototypical organic diaryl diselenide [(PhSe)2], RC513 displayed superior protective properties against MeHg-toxicity in vitro; this was paralleled by a more pronounced upregulation of defenses related to detoxification of peroxides, which are well-known MeHg-derived intermediate oxidant species.

Metabolic activity in cryopreserved and grafted ovarian tissue using high-resolution respirometry.

Cryopreservation of ovarian tissue followed by transplantation represents a strategy to restore ovarian function and fertility. Stress from cryopreservation-thawing processes can lead to alterations and/or damage to mitochondrial structure and functionality. High resolution respirometry and histological analysis were used to evaluate the effect of cryopreservation and transplantation on ovarian tissue. Four different conditions were performed: Fresh non-transplanted tissue, Fresh transplanted tissue, Cryopreserved non-transplanted tissue and Cryopreserved transplanted tissue. All groups were able to respond to the substrates-uncoupler-inhibitor protocol. We found a dramatic decrease in general oxygen consumption in hemi-ovaries submitted to cryopreservation and/or transplantation. The effect of cryopreservation on mitochondrial metabolism was less intense than effect of transplantation, since the transplantation affected all of the mitochondrial states. A total of 2644 follicles were analyzed. Of these, 2198 were classified as morphologically normal. The percentage of morphologically normal follicles was significantly lower in the Cryopreserved transplanted group when compared to the Cryopreserved non-transplanted group and the Fresh transplanted group (p-value < 0.05). Despite decreased follicular viability and mitochondrial activity, the cryopreservation followed by transplantation of ovarian tissue proved feasible for attempts to restore ovarian function.

High Cholesterol Diet Exacerbates Blood-Brain Barrier Disruption in LDLr-/- Mice: Impact on Cognitive Function.

BACKGROUND: Evidence has revealed an association between familial hypercholesterolemia and cognitive impairment. In this regard, a connection between cognitive deficits and hippocampal blood-brain barrier (BBB) breakdown was found in low-density lipoprotein receptor knockout mice (LDLr-/-), a mouse model of familial hypercholesterolemia. OBJECTIVE: Herein we investigated the impact of a hypercholesterolemic diet on cognition and BBB function in C57BL/6 wild-type and LDLr-/-mice. METHODS: Animals were fed with normal or high cholesterol diets for 30 days. Thus, wild-type and LDLr-/-mice were submitted to memory paradigms. Additionally, BBB integrity was evaluated in the mice's prefrontal cortices and hippocampi. RESULTS: A tenfold elevation in plasma cholesterol levels of LDLr-/-mice was observed after a hypercholesterolemic diet, while in wild-type mice, the hypercholesterolemic diet exposure increased plasma cholesterol levels only moderately and did not induce cognitive impairment. LDLr-/-mice presented memory impairment regardless of the diet. We observed BBB disruption as an increased permeability to sodium fluorescein in the prefrontal cortices and hippocampi and a decrease on hippocampal claudin-5 and occludin mRNA levels in both wild-type and LDLr-/-mice treated with a hypercholesterolemic diet. The LDLr-/-mice fed with a regular diet already presented BBB dysfunction. The BBB-increased leakage in the hippocampi of LDLr-/-mice was related to high microvessel content and intense astrogliosis, which did not occur in the control mice. CONCLUSION: Therefore, LDLr-/-mice seem to be more susceptible to cognitive impairments and BBB damage induced by exposure to a high cholesterol diet. Finally, BBB disruption appears to be a relevant event in hypercholesterolemia-induced brain alterations.

LDL Receptor Deficiency Does not Alter Brain Amyloid-ß Levels but Causes an Exacerbation of Apoptosis.

Familial hypercholesterolemia (FH) is a genetic disorder caused by dysfunction of low density lipoprotein receptors (LDLr), resulting in elevated plasma cholesterol levels. FH patients frequently exhibit cognitive impairment, a finding recapitulated in LDLr deficient mice (LDLr-/-), an animal model of FH. In addition, LDLr-/- mice are more vulnerable to the deleterious memory impact of amyloid-ß (Aß), a peptide linked to Alzheimer's disease. Here, we investigated whether the expression of proteins involved in Aß metabolism are altered in the brains of adult or middle-aged LDLr-/- mice. After spatial memory assessment, Aß levels and gene expression of LDLr related-protein 1, proteins involved in Aß synthesis, and apoptosis-related proteins were evaluated in prefrontal cortex and hippocampus. Moreover, the location and cell-specificity of apoptosis signals were evaluated. LDLr-/- mice presented memory impairment, which was more severe in middle-aged animals. Memory deficit in LDLr-/- mice was not associated with altered expression of proteins involved in Aß processing or changes in Aß levels in either hippocampus or prefrontal cortex. We further found that the expression of Bcl-2 was reduced while the expression of Bax was increased in both prefrontal cortex and hippocampus in 3- and 14-month-old LDLr-/-mice Finally, LDLr-/- mice presented increased immunoreactivity for activated caspase-3 in the prefrontal cortex and hippocampus. The activation of caspase 3 was predominantly associated with neurons in LDLr-/- mice. Cognitive impairment in LDLr-/- mice is thus accompanied by an exacerbation of neuronal apoptosis in brain regions related to memory formation, but not by changes in Aß processing or levels.

Hypercholesterolemia impairs contextual fear conditioning memory formation in female mice: evidence for cholinergic dysfunction.

The present study evaluated the effects of hypercholesterolemia in response to conditioned aversive stimuli in mice. Specifically, (a) young (3 months old) and aged (24 months old) female C57Bl/6 mice were fed daily for 4 weeks with a standard rodent diet or an enriched cholesterol diet (ECD) and then subjected to the contextual fear conditioning test. In another experimental set, 3-month-old C576Bl/6 female mice, fed daily during the 4 weeks with the standard rodent diet or ECD, were subjected to the contextual fear conditioning test and received vehicle or scopolamine (0.37 mg/kg; intraperitoneally) immediately after the training session. (b) 12-month-old C576Bl/6 and low-density lipoprotein receptor knockout mice (LDLr) female mice were subjected to the contextual fear conditioning test. In another experimental set, they were subjected to the contextual fear conditioning test and received vehicle or donepezil (3.0 mg/kg; intraperitoneally) immediately after the training session. The present results show that (a) the ECD specifically impaired retrieval of contextual fear memory in aged mice; (b) an ineffective dose of scopolamine impaired fear memory consolidation in young mice fed the ECD; (c) LDLr mice presented impaired contextual fear memory retrieval; and (d) boosting cholinergic neurotransmission with a single donepezil administration at the consolidation window led to improved fear memory consolidation in LDLr mice. These findings suggest that high levels of cholesterol induced by either an ECD or a genetic deletion of LDLr decreased freezing behavior on the contextual fear conditioning test, which seemed to involve dysfunction of the cholinergic system.

Antidepressant effects of creatine on amyloid ß1-40-treated mice: The role of GSK-3ß/Nrf2 pathway.

Alzheimer's disease (AD) is characterized by progressive synaptic dysfunction and neuronal lost in specific brain areas including hippocampus, resulting in memory/learning deficits and cognitive impairments. In addition, non-cognitive symptoms are reported in AD patients, such as anxiety, apathy and depressed mood. The current antidepressant drugs present reduced efficacy to improve depressive symptoms in AD patients. Here, we investigated the ability of creatine, a compound with neuroprotective and antidepressant properties, to counteract amyloid ß1-40 peptide-induced depressive-like behavior in mice. Moreover, we addressed the participation of the intracellular signaling pathway mediated by glycogen synthase kinase-3ß (GSK-3ß)/nuclear factor erythroid-2-related factor 2 (Nrf2) in the creatine effects. Aß1-40 administration (400 pmol/mouse, i.c.v.) increased the immobility time in the tail suspension test and decreased the grooming time and increased latency to grooming in the splash test, indicative of depressive-like behavior. These impairments were attenuated by creatine (0.01 and 10 mg/kg, p.o.) and fluoxetine (10 mg/kg, p.o., positive control). No significant alterations on locomotor performance were observed in the open field. Aß1-40 administration did not alter hippocampal phospho-GSK-3ß (Ser9)/total GSK-3ß, total GSK-3ß and heme oxygenase-1 (HO-1) immunocontents. However, Aß1-40-infused mice treated with creatine (0.01 mg/kg) presented increased phosphorylation of GSK-3ß(Ser9) and HO-1 immunocontent in the hippocampus. Fluoxetine per se increased GSK-3ß(Ser9) phosphorylation, but did not alter HO-1 levels. In addition, Aß1-40 administration increased hippocampal glutathione (GSH) levels as well as glutathione reductase (GR) and thioredoxin reductase (TrxR) activities, and these effects were abolished by creatine and fluoxetine. This study provides the first evidence of the antidepressive-like effects of creatine in Aß1-40-treated mice, which were accompanied by hippocampal inhibition of GSK-3ß and modulation of antioxidant defenses. These findings indicate the potential of creatine for the treatment of depression associated with AD.

Syzygium cumini leaf extract inhibits LDL oxidation, but does not protect the liproprotein from glycation.

ETNOPHARMACOLOGICAL RELEVANCE: Syzygium cumini (L.) Skeels is a plant widely used in folk medicine to treat diabetes mellitus (DM). The tea from its leaves is frequently used by diabetics for lowering hyperglycemia. There is a close relationship between DM and atherosclerosis, a chronic immuno-inflammatory disease, were the early stages encompass oxidative and glycative modifications in the structure of low density lipoprotein (LDL). AIM OF THIS STUDY: To investigate the potential protective effects of aqueous-leaf extract from Syzygium cumini (S.cExt) against CuSO4-induced oxidation and methylglyoxal (MG)-induced glycation of human LDL in vitro. MATERIALS AND METHODS: LDL oxidative changes were evaluated by measuring conjugated dienes (CD) formation, thiobarbituric acid reactive substances (TBARS) levels, quenching of tryptophan (Trp) fluorescence and structural modifications in LDL particle. In LDL glycated by MG (glyLDL), we determined the levels of fluorescent advanced glycation end products (AGEs) and mobility by agarose gel electrophoresis. RESULTS: S.cExt blocked oxidative events induced by CuSO4 in human LDL, plasma and serum. Fourier transform infrared spectroscopy (FT-IR) revealed that specific regions of apoB100 were oxidized by CuSO4 in human LDL and that S.cExt reduced these oxidations. Unlike, the increased AGEs levels and eletrophoretic mobility observed in LDL MG-glycated were not modified by S.cExt. CONCLUSION: The findings herein indicate that S.cExt could be tested in atherogenesis models as potential protective agent against LDL oxidation.

Brain-Defective Insulin Signaling Is Associated to Late Cognitive Impairment in Post-Septic Mice.

Sepsis survivors frequently develop late cognitive impairment. Because little is known on the mechanisms of post-septic memory deficits, there are no current effective approaches to prevent or treat such symptoms. Here, we subjected mice to severe sepsis induced by cecal ligation and puncture (CLP) and evaluated the sepsis-surviving animals in the open field, novel object recognition (NOR), and step-down inhibitory avoidance (IA) task at different times after surgery. Post-septic mice (30 days post-surgery) failed in the NOR and IA tests but exhibited normal performance when re-evaluated 45 days after surgery. Cognitive impairment in post-septic mice was accompanied by reduced hippocampal levels of proteins involved in synaptic plasticity, including synaptophysin, cAMP response element-binding protein (CREB), CREB phosphorylated at serine residue 133 (CREBpSer133), and GluA1 phosphorylated at serine residue 845 (GluA1pSer845). Expression of tumor necrosis factor α (TNF-α) was increased and brain insulin signaling was disrupted, as indicated by increased hippocampal IRS-1 phosphorylation at serine 636 (IRS-1pSer636) and decreased phosphorylation of IRS-1 at tyrosine 465 (IRS-1pTyr465), in the hippocampus 30 days after CLP. Phosphorylation of Akt at serine 473 (AktpSer473) and of GSK3 at serine 9 (GSK3ßpSer9) were also decreased in hippocampi of post-septic animals, further indicating that brain insulin signaling is disrupted by sepsis. We then treated post-septic mice with liraglutide, a GLP-1 receptor agonist with insulinotropic activity, or TDZD-8, a GSK3ß inhibitor, which rescued NOR memory. In conclusion, these results establish that hippocampal inflammation and disrupted insulin signaling are induced by sepsis and are linked to late memory impairment in sepsis survivors.

Organoselenium compounds as mimics of selenoproteins and thiol modifier agents.

Selenium is an essential trace element for animals and its role in the chemistry of life relies on a unique functional group: the selenol (-SeH) group. The selenol group participates in critical redox reactions. The antioxidant enzymes glutathione peroxidase (GPx) and thioredoxin reductase (TrxR) exemplify important selenoproteins. The selenol group shares several chemical properties with the thiol group (-SH), but it is much more reactive than the sulfur analogue. The substitution of S by Se has been exploited in organic synthesis for a long time, but in the last 4 decades the re-discovery of ebselen (2-phenyl-1,2-benzisoselenazol-3(2H)-one) and the demonstration that it has antioxidant and therapeutic properties has renovated interest in the field. The ability of ebselen to mimic the reaction catalyzed by GPx has been viewed as the most important molecular mechanism of action of this class of compound. The term GPx-like or thiol peroxidase-like reaction was previously coined in the field and it is now accepted as the most important chemical attribute of organoselenium compounds. Here, we will critically review the literature on the capacity of organoselenium compounds to mimic selenoproteins (particularly GPx) and discuss some of the bottlenecks in the field. Although the GPx-like activity of organoselenium compounds contributes to their pharmacological effects, the superestimation of the GPx-like activity has to be questioned. The ability of these compounds to oxidize the thiol groups of proteins (the thiol modifier effects of organoselenium compounds) and to spare selenoproteins from inactivation by soft-electrophiles (MeHg+, Hg2+, Cd2+, etc.) might be more relevant for the explanation of their pharmacological effects than their GPx-like activity. In our view, the exploitation of the thiol modifier properties of organoselenium compounds can be harnessed more rationally than the use of low mass molecular structures to mimic the activity of high mass macromolecules that have been shaped by millions to billions of years of evolution.

MPP+-Lesioned Mice: an Experimental Model of Motor, Emotional, Memory/Learning, and Striatal Neurochemical Dysfunctions.

The neurotoxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) induces motor and nonmotor dysfunctions resembling Parkinson's disease (PD); however, studies investigating the effects of 1-methyl-4-phenylpyridinium (MPP+), an active oxidative product of MPTP, are scarce. This study investigated the behavioral and striatal neurochemical changes (related to oxidative damage, glial markers, and neurotrophic factors) 24 h after intracerebroventricular administration of MPP+ (1.8-18 µg/mouse) in C57BL6 mice. MPP+ administration at high dose (18 µg/mouse) altered motor parameters, since it increased the latency to leave the first quadrant and reduced crossing, rearing, and grooming responses in the open-field test and decreased rotarod latency time. MPP+ administration at low dose (1.8 µg/mouse) caused specific nonmotor dysfunctions as it produced a depressive-like effect in the forced swim test and tail suspension test, loss of motivational and self-care behavior in the splash test, anxiety-like effect in the elevated plus maze test, and short-term memory deficit in the step-down inhibitory avoidance task, without altering ambulation. MPP+ at doses of 1.8-18 µg/mouse increased tyrosine hydroxylase (TH) immunocontent and at 18 µg/mouse increased α-synuclein and decreased parkin immunocontent. The astrocytic calcium-binding protein S100B and glial fibrillary acidic protein (GFAP)/S100B ratio was decreased following MPP+ administration (18 µg/mouse). At this highest dose, MPP+ increased the ionized calcium-binding adapter molecule 1 (Iba-1) immunocontent, suggesting microglial activation. Also, MPP+ at a dose of 18 µg/mouse increased thiobarbituric acid reactive substances (TBARS) and glutathione (GSH) levels and increased glutathione peroxidase (GPx) and hemeoxygenase-1 (HO-1) immunocontent, suggesting a significant role for oxidative stress in the MPP+-induced striatal damage. MPP+ (18 µg/mouse) also increased striatal fibroblast growth factor 2 (FGF-2) and brain-derived neurotrophic factor (BDNF) levels. Moreover, MPP+ decreased tropomyosin receptor kinase B (TrkB) immunocontent. Finally, MPP+ (1.8-18 µg/mouse) increased serum corticosterone levels and did not alter acetylcholinesterase (AChE) activity in the striatum but increased it in cerebral cortex and hippocampus. Collectively, these results indicate that MPP+ administration at low doses may be used as a model of emotional and memory/learning behavioral deficit related to PD and that MPP+ administration at high dose could be useful for analysis of striatal dysfunctions associated with motor deficits in PD.

Guanosine prevents nitroxidative stress and recovers mitochondrial membrane potential disruption in hippocampal slices subjected to oxygen/glucose deprivation.

Guanosine, the endogenous guanine nucleoside, prevents cellular death induced by ischemic events and is a promising neuroprotective agent. During an ischemic event, nitric oxide has been reported to either cause or prevent cell death. Our aim was to evaluate the neuroprotective effects of guanosine against oxidative damage in hippocampal slices subjected to an in vitro ischemia model, the oxygen/glucose deprivation (OGD) protocol. We also assessed the participation of nitric oxide synthase (NOS) enzymes activity on the neuroprotection promoted by guanosine. Here, we showed that guanosine prevented the increase in ROS, nitric oxide, and peroxynitrite production induced by OGD. Moreover, guanosine prevented the loss of mitochondrial membrane potential in hippocampal slices subjected to OGD. Guanosine did not present an antioxidant effect per se. The protective effects of guanosine were mimicked by inhibition of neuronal NOS, but not of inducible NOS. The neuroprotective effect of guanosine may involve activation of cellular mechanisms that prevent the increase in nitric oxide production, possibly via neuronal NOS.

Atorvastatin prevents cell damage via modulation of oxidative stress, glutamate uptake and glutamine synthetase activity in hippocampal slices subjected to oxygen/glucose deprivation.

Oxygen-glucose deprivation (OGD) in brain cells increases extracellular glutamate concentration leading to excitotoxicity. Glutamate uptake from the synaptic cleft is carried out by glutamate transporters, which are likely to be modulated by oxidative stress. Therefore, oxidative stress is associated with reduced activity of glutamate transporters and glutamine synthetase, thus increasing extracellular glutamate levels that may aggravate damage to brain cells. Atorvastatin, a cholesterol-lowering agent, has been shown to exert neuroprotective effects. The aim of this study was to investigate if in vivo atorvastatin treatment would have protective effects against hippocampal slices subjected to OGD, ex vivo. Atorvastatin pretreatment promoted increased cell viability after OGD and reoxygenation of hippocampal slices. Atorvastatin-induced neuroprotection may be related to diminished oxidative stress, since it prevented OGD-induced decrement of non-proteic thiols (NPSH) levels and increase in the production of reactive oxygen species (ROS). Atorvastatin pretreatment also prevented the OGD-induced decrease in glutamate uptake and glutamine synthetase activity, although it had no effect on OGD-induced excitatory aminoacids release. Addition of cholesterol before OGD and reoxygenation, abolished the protective effect of atorvastatin on cellular viability as well as on glutamate uptake and glutamine synthetase activity. Therefore, atorvastatin is capable of preventing OGD-induced cell death, an effect achieved due to modulation of glutamate uptake and glutamine synthetase activity, and associated with diminished oxidative stress. Additionally, atorvastatin effects were dependent on its action on cholesterol synthesis inhibition. Thus, atorvastatin might be a useful strategy in the prevention of glutamate exitotoxicity involved in brain injuries such as vascular disorders.

Diphenyl diselenide-modulation of macrophage activation: down-regulation of classical and alternative activation markers.

Diphenyl diselenide (PhSe)(2), a simple organoselenium compound, possesses interesting pharmacological properties that are under extensive research. As macrophages respond to microenvironmental stimuli and can display activities engaged in the initiation and the resolution of inflammation, in the present report we describe the ability of (PhSe)(2) to modulate the macrophage activation. Our data indicate that (PhSe)(2) could inhibit the NO production in a dose-dependent fashion in peritoneal macrophages activated by LPS or treated with vehicle alone. We could demonstrate that this effect correlated with a reduction in the expression of the inducible NO synthase in (PhSe)(2)-treated cells. Furthermore, (PhSe)(2) suppressed the production of reactive oxygen species, diminished the activity of the arginase enzyme, and the accumulation of nitrotyrosine modified proteins in LPS-stimulated macrophages. This compound also diminished the antigen presentation capacity of classically activated macrophages, as it reduced MHCII and CD86 expression. In addition, (PhSe)(2) modulated the alternative activation phenotype of macrophages. Dexamethasone-activated macrophages presented higher production of IL-10 and CD206, which were both down-regulated by the addition of (PhSe)(2). These results suggest that (PhSe)(2) possesses antioxidant and anti-inflammatory activities in classically-activated macrophages. We could demonstrate that (PhSe)(2) can be also utilized to modulate the alternative activation phenotype of macrophages.

Effect of diphenyl diselenide on the development of experimental autoimmune encephalomyelitis.

Experimental autoimmune encephalomyelitis (EAE) is a T cell-mediated inflammatory and demyelinating disease of the central nervous system with clinical and pathological similarities with multiple sclerosis. The oxidative stress is one of the major mediators of demyelination and axonal damage in both, multiple sclerosis and EAE. Therefore, several studies are being performed to assess whether treatment with antioxidants prevents the progression of these diseases. Some organic forms of selenium that exhibit glutathione peroxidase-like activity have become good candidates for disease prevention and therapy since they catalytically remove oxidative stressors. Particularly, diphenyl diselenide ((PhSe)(2)) exerts antioxidant activity and has neuroprotective effects in several systems. The aim of the present study was to prove the therapeutic activity of (PhSe)(2) on the development of EAE. Intraperitoneally administered (PhSe)(2) (1-25 µmoles/kg body weight/day) reduced the incidence of the disease but was also deleterious for the animals. Conversely, (PhSe)(2) given orally (80 µmoles/kg body weight/day) produced a significant inhibition of EAE without any toxic effect. In addition, there was a reduction of the characteristic histological alterations and a diminished in vivo and in vitro T-cell response against the encephalitogenic myelin basic protein. These results show an effective suppression of the autoimmune response that could be the base for future developments of successful antioxidants therapies in EAE as well as in multiple sclerosis.

The intranasal administration of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP): a new rodent model to test palliative and neuroprotective agents for Parkinson's disease.

Parkinson's disease (PD) is the second most common neurodegenerative disorder affecting approximately 1% of the population older than 60 years. Classically, PD is considered to be a motor system disease and its diagnosis is based on the presence of a set of cardinal motor signs that are consequence of a pronounced death of dopaminergic neurons in the substantia nigra pars compacta (SNc). Nowadays there is considerable evidence showing that non-dopaminergic degeneration also occurs in other brain areas which seems to be responsible for the deficits in olfactory, emotional and memory functions that precede the classical motor symptoms in PD. Dopamine-replacement therapy has dominated the treatment of PD and although the currently approved antiparkinsonian agents offer effective relief of the motor deficits, they have not been found to alleviate the non-motor features as well as the underlying dopaminergic neuron degeneration and thus drug efficacy is gradually lost. Another major limitation of chronic dopaminergic therapy is the numerous adverse effects such as dyskinesias, psychosis and behavioral disturbance. The development of new therapies in PD depends on the existence of representative animal models to facilitate the evaluation of new pharmacological agents before they are applied in clinical trials. We have recently proposed a new experimental model of PD consisting of a single intranasal (i.n.) administration of the proneurotoxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP, 1 mg/nostril) in rodents. Our findings demonstrated that rats and mice treated intranasally with MPTP suffer impairments in olfactory, cognitive, emotional and motor functions conceivably analogous to those observed during different stages of PD. Such infusion causes time-dependent loss of tyrosine hydroxylase in the olfactory bulb and SNc, resulting in significant dopamine depletion in different brain areas. We have also identified some pathogenic mechanisms possibly involved in the neurodegeneration induced by i.n. administration of MPTP including mitochondrial dysfunction, oxidative stress, activation of apoptotic cell death mechanisms and glutamatergic excitotoxicity. Therefore, the present review attempts to provide a comprehensive picture of the i.n. MPTP model and to highlight recent findings from our group showing its potential as a valuable rodent model for testing novel drugs that may provide alternative or adjunctive treatment for both motor and non-motor symptoms relief with a reduced side-effect profile as well as the discovery of compounds to modify the course of PD.

Proanthocyanidin-rich fraction from Croton celtidifolius Baill confers neuroprotection in the intranasal 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine rat model of Parkinson's disease.

We have recently demonstrated that rodents treated intranasally with 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) suffered impairments in olfactory, cognitive and motor functions associated with time-dependent disruption of dopaminergic neurotransmission in different brain structures conceivably analogous to those observed during different stages of Parkinson's disease (PD). On the other hand, the proanthocyanidin-rich fraction (PRF) obtained from the bark of Croton celtidifolius Baill (Euphorbiaceae), a tree frequently found in the Atlantic forest in south Brazil, has been described to have several neurobiological activities including antioxidant and anti-inflammatory properties, which may be of interest in the treatment of PD. The present data indicated that the pretreatment with PRF (10 mg/kg, i.p.) during five consecutive days was able to prevent mitochondrial complex-I inhibition in the striatum and olfactory bulb, as well as a decrease of the enzyme tyrosine hydroxylase expression in the olfactory bulb and substantia nigra of rats infused with a single intranasal administration of MPTP (1 mg/nostril). Moreover, pretreatment with PRF was found to attenuate the short-term social memory deficits, depressive-like behavior and reduction of locomotor activity observed at different periods after intranasal MPTP administration in rats. Altogether, the present findings provide strong evidence that PRF from C. celtidifolius may represent a promising therapeutic tool in PD, thus being able to prevent both motor and non-motor early symptoms of PD, together with its neuroprotective potential.

Atorvastatin prevents hippocampal cell death, neuroinflammation and oxidative stress following amyloid-ß(1-40) administration in mice: evidence for dissociation between cognitive deficits and neuronal damage.

The accumulation of amyloid-beta (Aß) peptides in the brain of human and rodents has been associated with the activation of glial cells, neuroinflammatory and oxidative responses, and cognitive deficits. These oxidative changes leave glutamate transporters more vulnerable and may result in reduction of their functions, resulting in excitotoxic damage. Herein, we evaluated the effects of atorvastatin, a HMG-CoA reductase inhibitor, in molecular and behavioral alterations induced by a single intracerebroventricular injection of aggregated Aß(1-40) (400 pmol) in mice. An increased glial fibrillar acidic protein (GFAP) expression and cyclooxygenase-2 (COX-2) levels, as well as increased lipid peroxidation and impairment in the glutathione antioxidant system and cell degeneration was found in the hippocampus of Aß(1-40)-treated mice. Aß(1-40) also induced a marked decrease in glutamatergic transporters (GLAST and GLT-1) expression and in l-[³H] glutamate uptake in mice hippocampus, in addition to spatial learning and memory deficits. Atorvastatin (10 mg/kg/day v.o.) was administered after Aß(1-40) injection and through seven consecutive days. Atorvastatin treatment was neuroprotective against cell degeneration induced by Aß(1-40), reducing inflammatory and oxidative responses and increasing the expression of glutamatergic transporters. On the other hand, atorvastatin did not reverse the cognitive impairments and failed to alter the hippocampal glutamate uptake in Aß(1-40)-treated mice. These results reinforce and extend the notion of the potential neuroprotective action of atorvastatin against the neuronal toxicity induced by Aß(1-40). In addition, the present findings suggest that the spatial learning and memory deficits induced by Aß peptides in rodents may not be entirely related to neuronal damage.

Synergistic neurotoxicity induced by methylmercury and quercetin in mice.

Methylmercury (MeHg) is a highly neurotoxic pollutant, whose mechanisms of toxicity are related to its pro-oxidative properties. A previous report showed under in vivo conditions the neuroprotective effects of plants of the genus Polygala against MeHg-induced neurotoxicity. Moreover, the flavonoid quercetin, isolated from Polygala sabulosa, displayed beneficial effects against MeHg-induced oxidative damage under in vitro conditions. In this study, we sought for potential beneficial effects of quercetin against the neurotoxicity induced by MeHg in Swiss female mice. Animals were divided into six experimental groups: control, quercetin low dose (5 mg/kg), quercetin high dose (50 mg/kg), MeHg (40 mg/L, in tap water), MeHg+quercetin low dose, and MeHg+quercetin high dose. After the treatment (21 days), a significant motor deficit was observed in MeHg+quercetin groups. Biochemical parameters related to oxidative stress showed that the simultaneous treatment with quercetin and MeHg caused a higher cerebellar oxidative damage when compared to the individual exposures. MeHg plus quercetin elicited a higher cerebellar lipid peroxidation than MeHg or quercetin alone. The present results indicate that under in vivo conditions quercetin and MeHg cause additive pro-oxidative effects toward the mice cerebellum and that such phenomenon is associated with the observed motor deficit.