6-OHDA Injection Research Articles

Growth/differentiation factor-15 deficiency compromises dopaminergic neuron survival and microglial response in the 6-hydroxydopamine mouse model of Parkinson's disease

Growth/differentiation factor-15 (Gdf-15) is a member of the TGF-β superfamily and a pleiotropic, widely distributed cytokine, which has been shown to play roles in various pathologies, including inflammation. Analysis of Gdf-15−/− mice has revealed that it serves the postnatal maintenance of spinal cord motor neurons and sensory neurons. In a previous study, exogenous Gdf-15 rescued 6-hydroxydopamine (6-OHDA) lesioned Gdf-15+/+ nigrostriatal dopaminergic (DAergic) neurons in vitro and in vivo. Whether endogenous Gdf-15 serves the physiological maintenance of nigrostriatal DAergic neurons in health and disease is not known and was addressed in the present study. Stereotactic injection of 6-OHDA into the medial forebrain bundle (MFB) led to a significant decline in the numbers of DAergic neurons in both Gdf-15+/+ and Gdf-15−/− mice over a time-period of 14days. However, this decrease was exacerbated in the Gdf-15−/− mice, with only 5.5% surviving neurons as compared to 24% in the Gdf-15+/+ mice. Furthermore, the microglial response to the 6-OHDA lesion was reduced in Gdf-15−/− mice, with significantly lower numbers of total and activated microglia and a differential cytokine expression as compared to the Gdf-15+/+ mice. Using in vitro models, we could demonstrate the importance of endogenous Gdf-15 in promoting DAergic neuron survival thus highlighting its relevance in a direct neurotrophic supportive role. Taken together, these results indicate the importance of Gdf-15 in promoting survival of DAergic neurons and regulating the inflammatory response post 6-OHDA lesion.

Intraventricular administration of endoneuraminidase-N facilitates ectopic migration of subventricular zone-derived neural progenitor cells into 6-OHDA lesioned striatum of mice

Polysialic acid (PSA), a carbohydrate polymer associated with the neural cell adhesion molecule (NCAM), plays an important role in the migration, differentiation and maturation of neuroblasts. Endoneuraminidase-N (Endo-N) can specifically cleave PSA from NCAM. The objective of the present study was to examine: the effect of Endo-N on characteristics of subventricular zone (SVZ)-derived neural progenitor cells (NPCs) in vitro; whether intraventricular administration of Endo-N could increase ectopic migration of SVZ-derived NPCs into 6-hydroxydopamine (6-OHDA)-lesioned striatum, and whether migrated NPCs could differentiate into neuronal and glial cells. In in vitro study, Endo-N was found to inhibit the migration of NPCs, and to enhance the differentiation of NPCs. In in vivo study, mice sequentially received injections of 6-OHDA into the right striatum, Endo-N into the right lateral ventricle, and bromodeoxyuridine (BrdU) intraperitoneally. The data showed that intraventricular injections of Endo-N disorganized the normal structure of the rostral migratory stream (RMS), and drastically increased the number of BrdU-immunoreactive (IR) cells in 6-OHDA-lesioned striatum. In addition, a number of BrdU-IR cells were double labeled for doublecortin (DCX), NeuN or glial fibrillary acidic protein (GFAP). The results suggest that interruption of neuroblast chain pathway with Endo-N facilitates ectopic migration of SVZ-derived NPCs into the lesioned striatum, and migrated NPCs can differentiate into neurons and astrocytes.

Decreased synaptic plasticity in the medial prefrontal cortex underlies short-term memory deficits in 6-OHDA-lesioned rats

Parkinson’s disease (PD) is characterized by motor dysfunction associated with dopaminergic degeneration in the dorsolateral striatum (DLS). However, motor symptoms in PD are often preceded by short-term memory deficits, which have been argued to involve deregulation of medial prefrontal cortex (mPFC). We now used a 6-hydroxydopamine (6-OHDA) rat PD model to explore if alterations of synaptic plasticity in DLS and mPFC underlie short-term memory impairments in PD prodrome. The bilateral injection of 6-OHDA (20μg/hemisphere) in the DLS caused a marked loss of dopaminergic neurons in the substantia nigra (>80%) and decreased monoamine levels in the striatum and PFC, accompanied by motor deficits evaluated after 21 days in the open field and accelerated rotarod. A lower dose of 6-OHDA (10μg/hemisphere) only induced a partial degeneration (about 60%) of dopaminergic neurons in the substantia nigra with no gross motor impairments, thus mimicking an early premotor stage of PD. Notably, 6-OHDA (10μg)—lesioned rats displayed decreased monoamine levels in the PFC as well as short-term memory deficits evaluated in the novel object discrimination and in the modified Y-maze tasks; this was accompanied by a selective decrease in the amplitude of long-term potentiation in the mPFC, but not in DLS, without changes of synaptic transmission in either brain regions. These results indicate that the short-term memory dysfunction predating the motor alterations in the 6-OHDA model of PD is associated with selective changes of information processing in PFC circuits, typified by persistent changes of synaptic plasticity.

Ventral tegmental area dopaminergic lesion-induced depressive phenotype in the rat is reversed by deep brain stimulation of the medial forebrain bundle

DBS of the medial forebrain bundle (MFB) has been investigated clinically in major depressive disorder patients with rapid and long-term reduction of symptoms. In the context of chronic bilateral high frequency deep brain stimulation (DBS) of the MFB, the current study looked at the impact of lesioning the ascending dopaminergic pathway at the level of the ventral tegmental area (VTA). Sprague-Dawley female rats were given bilateral injection of 6-OHDA into the VTA (VTA-lx group) or were left unlesioned (control group). Later, all animals received bilateral microelectrode implantation into the MFB followed by chronic continuous stimulation for 3 weeks. Behavioral tests were performed as baseline and following MFB-DBS, along with histological analysis. Pre-stimulation baseline testing of the VTA-lx animals indicated depressive-like phenotype in comparison with controls. Response to MFB-DBS varied according to (i) the degree of dopaminergic depletion: animals with severe mesocorticolimbic dopamine depletion did not, whilst those with mild dopamine loss responded well to stimulation; (ii) environmental conditions and the nature of the behavioral tests, e.g., stressful vs non-stressful situations. Neuromodulation-induced c-fos expression in the prelimbic frontal cortex and nucleus accumbens was also dependent upon integrity of the dopaminergic ascending projections.Our results confirm a potential role for dopamine in symptom relief observed in clinical MFB-DBS. Although mechanisms are not fully understood, the data suggests that the rescue of depressive phenotype in rodents can work via both dopamine-dependent and independent mechanisms. Further investigations concerning the network of depression using neuromodulation platforms in animal models might give insight into genesis and treatment of major depression disorder.

Effect of Buspirone, Fluoxetine and 8-OH-DPAT on Striatal Expression of Bax, Caspase-3 and Bcl-2 Proteins in 6-Hydroxydopamine-Induced Hemi-Parkinsonian Rats.

The exact pathogenesis of sporadic parkinson's disease (PD) is still unclear. Numerous evidences suggest involvement of apoptosis in the death of dopaminergic neurons. In this study we investigated the effect of sub-chronic administration of buspirone, fluoxetine and 8-hydroxy-2-[di-n-propylamino]tetralin (8-OH-DPAT) in 6-hydroxydopamine (6-OHDA)-lesioned rats and assayed striatal concentrations of apoptotic (Bax, Caspase3) and anti-apoptotic (Bcl-2) proteins. 6-OHDA (8μg/2μl/rat) was injected unilaterally into the central region of the substantia nigra pars copmacta (SNc) of male Wistar rats and then, after 21 days lesioned rats were treated with intraperitonel (i.p) 1 mg/kg injections of buspirone, fluoxetine and 8-OH-DPAT for 10 consecutive days. Striatum of rats was removed at tenth day of drugs administration and were analyzed by western blotting method to measure Bax, caspase3 and Bcl-2 expression. The results showed that the expression of Bax and caspase3 proteins was increased three weeks after 6-OHDA injection while they were decreased significantly in parkinsonian rats which were treated by buspirone, fluoxetine and 8-OH-DPAT. Bcl-2 was decreased and increased in parkinsonian rats and parkinsonian rats treated with buspirone, fluoxetine and 8-OH-DPAT, respectively. Our study indicates that sub-chronic administration of serotonergic drugs such as buspirone, fluoxetine and 8-OH-DPAT restores striatal concentration of apoptotic and anti-apoptotic factors to the basal levels of normal non-lesioned rats. We suggest that these drugs can be used as a potential adjunctive therapy in PD through attenuating neuronal apoptotic process.

Intracranial application of near-infrared light in a hemi-parkinsonian rat model: the impact on behavior and cell survival.

OBJECT The authors of this study used a newly developed intracranial optical fiber device to deliver near-infrared light (NIr) to the midbrain of 6-hydroxydopamine (6-OHDA)-lesioned rats, a model of Parkinson's disease. The authors explored whether NIr had any impact on apomorphine-induced turning behavior and whether it was neuroprotective. METHODS Two NIr powers (333 nW and 0.16 mW), modes of delivery (pulse and continuous), and total doses (634 mJ and 304 J) were tested, together with the feasibility of a midbrain implant site, one considered for later use in primates. Following a striatal 6-OHDA injection, the NIr optical fiber device was implanted surgically into the midline midbrain area of Wistar rats. Animals were tested for apomorphine-induced rotations, and then, 23 days later, their brains were aldehyde fixed for routine immunohistochemical analysis. RESULTS The results showed that there was no evidence of tissue toxicity by NIr in the midbrain. After 6-OHDA lesion, regardless of mode of delivery or total dose, NIr reduced apomorphine-induced rotations at the stronger, but not at the weaker, power. The authors found that neuroprotection, as assessed by tyrosine hydroxylase expression in midbrain dopaminergic cells, could account for some, but not all, of the observed behavioral improvements; the groups that were associated with fewer rotations did not all necessarily have a greater number of surviving cells. There may have been other "symptomatic" elements contributing to behavioral improvements in these rats. CONCLUSIONS In summary, when delivered at the appropriate power, delivery mode, and dosage, NIr treatment provided both improved behavior and neuroprotection in 6-OHDA-lesioned rats.

Adaptive down-regulation of the serotonin transporter in the 6-hydroxydopamine-induced rat model of preclinical stages of Parkinson’s disease and after chronic pramipexole treatment

Our recent study has indicated that a moderate lesion induced by bilateral 6-hydroxydopamine (6-OHDA) injections into the ventrolateral region of the caudate-putamen (CP) in rats, modeling preclinical stages of Parkinson’s disease, induces a “depressive-like” behavior which is reversed by chronic treatment with pramipexole (PRA).The aim of the present study was to examine the influence of the above lesion and chronic PRA treatment on binding to the serotonin transporter (SERT) in different brain regions. As before, 6-OHDA (15μg/2.5μl) was administered bilaterally into the CP. PRA (1mg/kg) was injected subcutaneously twice a day for 2weeks. Serotonergic and dopaminergic neurons of the dorsal raphe (DR) were immunostained for tryptophan hydroxylase and tyrosine hydroxylase, respectively, and were counted stereologically. Binding of [3H]GBR 12,935 to the dopamine transporter (DAT) and [3H]citalopram to SERT was analyzed autoradiographically. Intrastriatal 6-OHDA injections decreased the number of dopaminergic, but not serotonergic neurons in the DR. 6-OHDA reduced the DAT binding in the CP, and SERT binding in the nigrostriatal system (CP, substantia nigra (SN)), limbic system (ventral tegmental area (VTA), nucleus accumbens (NAC), amygdala, prefrontal cortex (PFCX), habenula, hippocampus) and DR. A significant positive correlation was found between DAT and SERT binding in the CP. Chronic PRA did not influence DAT binding but reduced SERT binding in the above structures, and deepened the lesion-induced losses in the core region of the NAC, SN, VTA and PFCX.The present study indicates that both the lesion of dopaminergic neurons and chronic PRA administration induce adaptive down-regulation of SERT binding. Moreover, although involvement of stimulation of dopaminergic transmission by chronic PRA in its “antidepressant” effect seems to be prevalent, additional contribution of SERT inhibition cannot be excluded.

Enteric Dysfunctions in Experimental Parkinson's Disease: Alterations of Excitatory Cholinergic Neurotransmission Regulating Colonic Motility in Rats.

Parkinson's disease is frequently associated with gastrointestinal symptoms, mostly represented by constipation and defecatory dysfunctions. This study examined the impact of central dopaminergic denervation, induced by injection of 6-hydroxydopamine (6-OHDA) into the medial forebrain bundle, on distal colonic excitatory cholinergic neuromotor activity in rats. Animals were euthanized 4 and 8 weeks after 6-OHDA injection. In vivo colonic transit was evaluated by radiologic assay. Electrically induced and carbachol-induced cholinergic contractions were recorded in vitro from longitudinal and circular muscle colonic preparations, whereas acetylcholine levels were assayed in the incubation media. Choline acetyltransferase (ChAT), HuC/D (pan-neuronal marker), muscarinic M2 and M3 receptors were assessed by immunohistochemistry or western blot assay. As compared with control rats, at week 4, 6-OHDA-treated animals displayed the following changes: decreased in vivo colonic transit rate, impaired electrically evoked neurogenic cholinergic contractions, enhanced carbachol-induced contractions, decreased basal and electrically stimulated acetylcholine release from colonic tissues, decreased ChAT immunopositivity in the neuromuscular layer, unchanged density of HuC/D immunoreactive myenteric neurons, and increased expression of colonic muscarinic M2 and M3 receptors. The majority of such alterations were also detected at week 8 post 6-OHDA injection. These findings indicate that central nigrostriatal dopaminergic denervation is associated with an impaired excitatory neurotransmission characterized by a loss of myenteric neuronal ChAT positivity and decrease in acetylcholine release, resulting in a dysregulated smooth muscle motor activity, which likely contributes to the concomitant decrease in colonic transit rate.

Anti-high mobility group box 1 antibody exerts neuroprotection in a rat model of Parkinson's disease

The high mobility group box-1 (HMGB1) exists as an architectural nuclear protein in the normal state, but displays an inflammatory cytokine-like activity in the extracellular space under pathological condition. Inflammation in the pathogenesis of Parkinson's disease (PD) has been documented. In this study, we investigated the involvement of HMGB1 in the pathology and the neuroprotective effects of neutralizing anti-HMGB1 monoclonal antibody (mAb) on an animal model of PD. Adult female Sprague–Dawley rats were initially injected with 6-hydroxydopmaine (6-OHDA, 20μg/4μl) into the right striatum, then anti-HMGB1 mAb (1mg/kg), or control mAb was intravenously administered immediately, at 6 and 24h after 6-OHDA injection. The treatment with anti-HMGB1 mAb significantly preserved dopaminergic neurons in substantia nigra pars compacta and dopaminergic terminals inherent in the striatum, and attenuated PD behavioral symptoms compared to the control mAb-treated group. HMGB1 was retained in the nucleus of neurons and astrocytes by inhibiting the proinflammation-induced oxidative stress in the anti-HMGB1 mAb-treated group, whereas HMGB1 translocation was observed in neurons at 1day and astrocytes at 7days after 6-OHDA injection in the control mAb-treated group. Anti-HMGB1 mAb inhibited the activation of microglia, disruption of blood–brain-barrier (BBB), and the expression of inflammation cytokines such as IL-1β and IL-6. These results suggested that HMGB1 released from neurons and astrocytes was at least partly involved in the mechanism and pathway of degeneration of dopaminergic neurons induced by 6-OHDA exposure. Intravenous administration of anti-HMGB1 mAb stands as a novel therapy for PD possibly acting through the suppression of neuroinflammation and the attenuation of disruption of BBB associated with the disease.

PET imaging of dopamine transporters with [18F]FE-PE2I: Effects of anti-Parkinsonian drugs

PurposeThis study aimed to assess the striatal [18F]FE-PE2I binding and the immunohistochemical stain of tyrosine hydroxylase (TH) in the striatum, and to evaluate the effects of therapeutic drugs on [18F]FE-PE2I binding. MethodsDynamic PET/CT of [18F]FE-PE2I was performed in Parkinson’s disease (PD) rat models, induced by the unilateral injection of 6-OHDA into the striatum. A simplified reference tissue model method was used to calculate the striatal binding potential (striatal BPND). Each of the four normal rats was pretreated with pramipexole, amantadine, and escitalopram 30min before [18F]FE-PE2I injection. The effect of l-DOPA combined with benserazide was assessed in the normal and PD rats. ResultsThe BPND was significantly lower in the lesioned striatum than in the striatum of the normal rats. After the pretreatment with pramipexole, amantadine, and escitalopram, the values of the striatal BPND did not differ from those of the controls. The pretreatment with l-DOPA/benserazide, however, significantly reduced the striatal BPND. The striatal BPND of the PD rats with l-DOPA/benserazide pretreatment was not different from that of the same PD rats with placebo treatment. Conclusion[18F]FE-PE2I may be used as a radioligand for the in-vivo imaging of the DAT. In the normal rats, [18F]FE-PE2I binding is unaffected by pramipexole, amantadine, and escitalopram. l-DOPA/benserazide does not affect the striatal [18F]FE-PE2I binding in PD rats

Amelioration of L-Dopa-Associated Dyskinesias with Triterpenoic Acid in a Parkinsonian Rat Model.

Levo-Dopa (L-Dopa) is widely used for the oral treatment of Parkinson's disease. However, chronic treatment with L-Dopa produces abnormal involuntary movements (AIMs) known as dyskinesias. In this study, commercially available oleanolic acid (OA) that has been previously shown to ameliorate the toxic effects of 6-hydroxydopamine (6-OHDA) in preconditioning studies was used to treat AIMs in a rat model for Parkinson's disease. The forelimb-use asymmetry test was used to measure Parkinson's disease-associated motor impairment. AIMs were measured after 21 days of L-Dopa administration. Glutathione levels were measured in blood, and catalase levels were measured in the substantia nigra and striatum of both the left and right hemispheres. We found that L-Dopa alone as well as L-Dopa and OA combination treatment attenuated the limb-use asymmetry caused by the unilateral injection of 6-OHDA. Chronic L-Dopa administration produced AIMs which were attenuated by treatment with OA. Catalase concentration decreased significantly in the striatum but not in the substantia nigra of the lesioned hemisphere. L-Dopa alone as well as the combined L-Dopa and OA treatment ameliorated the effects of 6-OHDA on catalase concentration. However, intervention with L-Dopa alone as well as with L-Dopa and OA did not affect plasma glutathione concentration. These results suggest that OA administration enhances the effect of catalase on reactive oxygen species following 6-OHDA injection. OA may provide possibilities as an adjunct treatment to prevent or attenuate the development of AIMs following chronic L-Dopa treatment in Parkinson's disease.

The effects of fluvoxamine maleate in a post-natal stress model of neurodegeneration

Parkinson’s disease (PD) is the most common movement disorder characterized by loss of dopamine neurons in the substantia nigra. Treatment strategies focus mainly on retarding the progression of the disease since the serological evaluation of new treatments is difficult. Motor symptoms of PD appear late as opposed to non-motor symptoms (i.e. depression, anxiety) frequently preceding motor symptoms. Fluvoxamine maleate (FM) is an antidepressant widely used to treat depression and anxiety disorders. In patients with both depression and Parkinsonism, treatment with FM may delay motor symptoms and protect dopamine neurons. We aimed to use an animal model of depression to investigate the neuroprotective effects of FM on a postnatal stressed model of neurodegeneration. The ethics clearance was 061/14/Animal.Sprague-Dawley rats (N = 80) were divided into eight groups. Maternal separation was used to create an animal model of depression. Behavioural tests (open field, elevated plus-maze, step and cylinder test) were conducted prior and after the injection of 6-OHDA into the medial forebrain bundle to mimic Parkinsonism. Animals received FM (25 mg/kg, i.p/day) pre and post-lesion and were sacrificed on postnatal day 76. Blood and brain tissues were collected for corticosterone, lipid peroxidation, dopamine and serotonin analyses.Maternal separation caused depressive-like symptoms prior to the 6-OHDA lesion and motor impairment after the lesion. Plasma corticosterone levels and lipid peroxidation were increased in the striatum and the prefrontal cortex while dopamine and serotonin levels decreased. Treatment with FM reversed these effects in maternally separated animals and showed neuroprotection to dopamine neurons in our parkinsonian rat model. Parkinson’s disease (PD) is the most common movement disorder characterized by loss of dopamine neurons in the substantia nigra. Treatment strategies focus mainly on retarding the progression of the disease since the serological evaluation of new treatments is difficult. Motor symptoms of PD appear late as opposed to non-motor symptoms (i.e. depression, anxiety) frequently preceding motor symptoms. Fluvoxamine maleate (FM) is an antidepressant widely used to treat depression and anxiety disorders. In patients with both depression and Parkinsonism, treatment with FM may delay motor symptoms and protect dopamine neurons. We aimed to use an animal model of depression to investigate the neuroprotective effects of FM on a postnatal stressed model of neurodegeneration. The ethics clearance was 061/14/Animal. Sprague-Dawley rats (N = 80) were divided into eight groups. Maternal separation was used to create an animal model of depression. Behavioural tests (open field, elevated plus-maze, step and cylinder test) were conducted prior and after the injection of 6-OHDA into the medial forebrain bundle to mimic Parkinsonism. Animals received FM (25 mg/kg, i.p/day) pre and post-lesion and were sacrificed on postnatal day 76. Blood and brain tissues were collected for corticosterone, lipid peroxidation, dopamine and serotonin analyses. Maternal separation caused depressive-like symptoms prior to the 6-OHDA lesion and motor impairment after the lesion. Plasma corticosterone levels and lipid peroxidation were increased in the striatum and the prefrontal cortex while dopamine and serotonin levels decreased. Treatment with FM reversed these effects in maternally separated animals and showed neuroprotection to dopamine neurons in our parkinsonian rat model.

Magnetic resonance imaging as a tool to image neuroinflammation in a rat model of Parkinson's disease--phagocyte influx to the brain is promoted by bilberry-enriched diet.

Neuroinflammation is a chronic event in neurodegenerative disorders. In the rat model of Parkinson's disease, including a striatal injection of the neurotoxin 6-hydroxydopamine (6-OHDA), antioxidant treatment affects the inflammatory process. Despite a heavy accumulation of microglia early after the injury, dopamine nerve fibre regeneration occurs. It remains unclear why this heavy accumulation of microglia is found early after the lesion in antioxidant-treated animals, or even more, what is the origin of these microglia. In this study magnetic resonance imaging (MRI) was used to elucidate whether the inflammatory response was generated from the blood or from activated brain microglia. Superparamagnetic iron oxide (SPIO) nanoparticles were injected intravenously prior to a striatal 6-OHDA injection to tag phagocytes in the blood. Rats were fed either with bilberry-enriched or control diet. T2*-weighted MRI scans were performed 1 week after the lesion, and hypointense areas were calculated from T2*-weighted images, to monitor the presence of SPIO particles. The results revealed that feeding the animals with bilberries significantly promoted accumulation of blood-derived immune cells. Gadolinium-enhanced MRI demonstrated no difference in leakage of the blood-brain barrier independent of diets. To conclude, bilberry-enriched diet promotes an influx of periphery-derived immune cells to the brain early after injury.

Neuroprotective effect of Spirulina fusiform and amantadine in the 6-OHDA induced Parkinsonism in rats.

BackgroundMulti-factorial etiology exists in pathophysiology of neurodegenerative diseases. The imbalance of anti-oxidant enzymes and dopamine level leads to Parkinsonism. The objective of this study was to assess the protective effect of Spirulina fusiform alone and in combination with amantadine against Parkinsonism effect in 6-hydroxydopamine (6-OHDA) induced rat model.MethodsS. fusiform was administered in different groups (500 mg/kg, once daily and twice daily) and a combination of spirulina (500 mg/kg, once daily) with amantadine (20 mg/kg once daily) for 30 days before and 14 days after a single injection of 6-OHDA into the dorsal striatum. Post lesion produced rotational behavior which was measured at two week intervals (37th and 44th day). Locomotors activity was also done at 44th and muscle coordination at 48th day. Dorsal striatum was isolated from rat brain for evaluating the antioxidant assays and dopamine content at 49th day.ResultsBoth the body rotations (ipsilateral and contralateral) were found to have a statistically significant (p < 0.001) decrease by 34.26 and 52 % after treatment with spirulina (Twice a day) in spirulina treated lesioned group. A higher percentage of improvement was shown in the reduction of ipsilateral (57.34 %) and contralateral (78.3 %) rotations in combination of spirulina with amantadine treated lesioned group rather than spirulina alone treated lesioned groups when compared with positive control lesioned group. Body movements and locomotor activity were improved statistically (p < 0.0001) significant in both treated lesioned groups (Combination of spirulina with amantadine and spirulina twice daily). Similar results were also seen in anti-oxidant levels which later on reached to the normal value. The levels of dopamine content had a statistically significant (p < 0.0001) increase by 78.3 % only in case of spirulina with amantadine treated lesioned group.ConclusionSpirulina is a potent nutraceutical supplement all over the world, so my preclinical study may contribute to give an additional adjuvant drug therapy in aging related disorders (Neurodegenerative as well as diabetes associated neurodegenerative disorders).Electronic supplementary materialThe online version of this article (doi:10.1186/s12906-015-0815-0) contains supplementary material, which is available to authorized users.

Lymphocytes reduce nigrostriatal deficits in the 6-hydroxydopamine mouse model of Parkinson's disease.

Neuroinflammation is a well-known neuropathological feature of Parkinson's disease (PD), but it remains controversial whether it is causal or consequential to neurodegeneration. While the role of microglia in the pathogenesis has been thoroughly investigated in human and different rodent models, data concerning the impact of the adaptive immune system on the pathogenesis of PD are still rare, although lymphocyte populations were found in brain tissue of PD patients and have been implicated in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-mediated neurodegeneration in mice. To test the hypothesis that the adaptive immune system contributes to the progression of PD in the murine 6-hydroxydopamine (6-OHDA) model, we performed unilateral 6-OHDA injection into the medial forebrain bundle and compared wild-type mice with recombination activating gene-1 deficient mice (RAG-1(-/-)), that lack mature lymphocytes. After 6-OHDA injection, immune-deficient mice moved significantly slower and less often than wild-type mice. Rotarod analysis displayed a shorter latency to fall in RAG-1(-/-) mice. Immunohistochemical analysis in wild-type mice demonstrated a higher CD8+ T cell density in the ipsilesional striatum compared to sham-operated animals. Cell counts of tyrosine hydroxylase positive dopaminergic neurons of the substantia nigra in immune compromised mice were significantly reduced compared to wild-type mice. Wild type bone marrow reconstitution into RAG-1(-/-) recipients rescued the clinical deterioration as well as the neurodegeneration in RAG-1(-/-) deficient recipients ameliorated clinical symptoms and neurodegeneration after 6-OHDA treatment. Our data indicate that lymphocytes reduce the clinical and neuropathological impact of 6-OHDA lesioning and thus may play a protective role in this toxic mouse model of PD.

Cardiac Dysregulation and Myocardial Injury in a 6-Hydroxydopamine-Induced Rat Model of Sympathetic Denervation.

BackgroundCardiac sympathetic denervation is found in various cardiac pathologies; however, its relationship with myocardial injury has not been thoroughly investigated.MethodsTwenty-four rats were assigned to the normal control group (NC), sympathectomy control group (SC), and a sympathectomy plus mecobalamin group (SM). Sympathectomy was induced by injection of 6-OHDA, after which, the destruction and distribution of sympathetic and vagal nerve in the left ventricle (LV) myocardial tissue were determined by immunofluorescence and ELISA. Heart rate variability (HRV), ECG and echocardiography, and assays for myocardial enzymes in serum before and after sympathectomy were examined. Morphologic changes in the LV by HE staining and transmission electron microscope were used to estimate levels of myocardial injury and concentrations of inflammatory cytokines were used to reflect the inflammatory reaction.ResultsInjection of 6-OHDA decreased NE (933.1 ± 179 ng/L for SC vs. 3418.1± 443.6 ng/L for NC, P < 0.01) and increased NGF (479.4± 56.5 ng/mL for SC vs. 315.85 ± 28.6 ng/mL for NC, P < 0.01) concentrations. TH expression was reduced, while ChAT expression showed no change. Sympathectomy caused decreased HRV and abnormal ECG and echocardiography results, and histopathologic examinations showed myocardial injury and increased collagen deposition as well as inflammatory cell infiltration in the cardiac tissue of rats in the SC and SM groups. However, all pathologic changes in the SM group were less severe compared to those in the SC group.ConclusionsChemical sympathectomy with administration of 6-OHDA caused dysregulation of the cardiac autonomic nervous system and myocardial injuries. Mecobalamin alleviated inflammatory and myocardial damage by protecting myocardial sympathetic nerves.

Intracarotid Infusion of Mesenchymal Stem Cells in an Animal Model of Parkinson's Disease, Focusing on Cell Distribution and Neuroprotective and Behavioral Effects.

Mesenchymal stem cells (MSCs) have been proposed as a potential therapeutic tool for Parkinson's disease (PD) and systemic administration of these cells has been tested in preclinical and clinical studies. However, no information on survival and actual capacity of MSCs to reach the brain has been provided. In this study, we evaluated homing of intraarterially infused rat MSCs (rMSCs) in the brain of rats bearing a 6-hydroxydopamine (6-OHDA)-induced lesion of the nigrostriatal tract, to establish whether the toxin-induced damage is sufficient to grant MSC passage across the blood-brain barrier (BBB) or if a transient BBB disruption is necessary. The rMSC distribution in peripheral organs and the effects of cell infusion on neurodegenerative process and motor deficits were also investigated. rMSCs were infused 14 days after 6-OHDA injection. A hyperosmolar solution of mannitol was used to transiently permeabilize the BBB. Behavioral impairment was assessed by adjusting step test and response to apomorphine. Animals were sacrificed 7 and 28 days after cell infusion. Our work shows that appreciable delivery of rMSCs to the brain of 6-OHDA-lesioned animals can be obtained only after mannitol pretreatment. A notable percentage of infused cells accumulated in peripheral organs. Infusion of rMSCs did not modify the progression of 6-OHDA-induced damage or the motor impairment at the stepping test, but induced progressive normalization of the pathological response (contralateral turning) to apomorphine administration. These findings suggest that many aspects should be further investigated before considering any translation of MSC systemic administration into the clinical setting for PD treatment. This study demonstrates that mesenchymal stem cells infused through the carotid artery do not efficiently cross the blood-brain barrier in rats with a Parkinson's disease-like degeneration of nigrostriatal neurons, unless a permeabilizing agent (e.g., mannitol) is used. The infusion did not reduce the neuronal damage and associated motor impairment, but abolished the motor abnormalities these animals typically show when challenged with a dopaminergic agonist. Therefore, although arterially infused mesenchymal stem cells did not show neurorestorative effects in this study's Parkinson's disease model, they appeared to normalize the pathological responsiveness of striatal neurons to dopaminergic stimulation. This capability should be further explored in future studies.

Chronic deep brain stimulation of the medial forebrain bundle reverses depressive-like behavior in a hemiparkinsonian rodent model.

Preclinical and clinical evidence suggests that depression might be associated with a dysfunction in the reward/motivation circuitry. Deep brain stimulation (DBS) of the superolateral branch of the medial forebrain bundle (MFB) has been shown in a recent clinical trial to provide a prompt and consistent improvement of depressive symptoms in treatment-resistant patients. In order to better understand the underlying mechanisms of neuromodulation in the context of depression, the effects of chronic bilateral MFB-DBS were assessed in a combined rodent model of depression and Parkinson’s disease. Female Sprague-Dawley rats received unilateral 6-OHDA injection in the right MFB and were divided into three groups: CMS-STIM, CMS-noSTIM and control group. The CMS groups were submitted to chronic unpredictable mild stress (CMS) protocol for 6 weeks. MFB-DBS was applied only to the CMS-STIM group for 1 week. All groups were repeatedly probed on a series of behavioral tasks following each intervention, and to a postmortem histological analysis. CMS led to an increase in immobility in the forced swim test, to a decrease in sucrose solution consumption in the sucrose preference test, as well as to an increased production of ultrasonic vocalizations in the 22 kHz range, indicating increased negative affect. MFB-DBS reversed the anhedonic-like and despair-like behaviors. The results suggest that unilateral dopamine depletion did not preclude MFB-DBS in reversing depressive-like and anhedonic-like behavior in the rodent. Further understanding of the importance of hemispheric dominance in neuropsychiatric disorders is essential in order to optimize stimulation as a therapeutic strategy in these diseases.

Tanshinone IIA protects dopaminergic neurons against 6-hydroxydopamine-induced neurotoxicity through miR-153/NF-E2-related factor 2/antioxidant response element signaling pathway

Parkinson’s disease (PD) is the second most common progressive neurodegenerative disorder with increased oxidative stress, the underlying vital process contributing to cell death. Tanshinone IIA (Tan IIA), a major bioactive diterpene quinone of Salva miltiorrhiza, had been proved effective in the MPTP model through its anti-inflammatory activity. Here in this research, we found that Tan IIA prevented the loss of nigrostriatal dopaminergic neurons by activating the NF-E2-related factor 2 (Nrf2)–antioxidant response element (ARE) pathway. The cytotoxicity of 6-hydroxydopamine (6-OHDA) was attenuated by the treatment of Tan IIA in SH-SY5Y cells, which significantly reduced 6-OHDA-induced lactic dehydrogenase release and reactive oxygen species production. Further study indicated that Tan IIA contributed to the nuclear accumulation of Nrf2, which bound to the ARE sequence, and activated ARE-regulated genes, including heme oxygenase-1, glutamate cysteine ligase catalytic subunit (GCLC) and glutamate cysteine ligase modifier subunit (GCLM). Tan IIA also protected against damage to mitochondrial membrane potential, reduced the translocation of cytochrome c from the mitochondria to the cytoplasm and the activation of Caspase-9 and Caspase-3. Moreover, we demonstrated the above effects were performed in Nrf2-dependent manner. Further studies revealed that Tan IIA reduced the enhancement of miR-153 by 6-OHDA, which targeted the 3′-UTR of Nrf2, and suppressed its expression and activation. Additionally, neurodegeneration caused by in vivo stereotaxic injection of 6-OHDA could also be ameliorated by the administration of Tan IIA. Taken together, our results strongly suggest that Tan IIA may be beneficial for the treatment of PD, and also confirm that targeting the Nrf2/ARE pathway is a promising strategy for therapeutic intervention in PD.

Differential degradation of motor deficits during gradual dopamine depletion with 6-hydroxydopamine in mice

Parkinson’s disease (PD) is a movement disorder whose cardinal motor symptoms arise due to the progressive loss of dopamine. Although this dopamine loss typically progresses slowly over time, currently there are very few animal models that enable incremental dopamine depletion over time within the same animal. This type of gradual dopamine depletion model would be useful in studies aimed at the prodromal phase of PD, when dopamine levels are pathologically low but motor symptoms have not yet presented. Utilizing the highly characterized neurotoxin 6-hydroxydopamine (6-OHDA), we have developed a paradigm to gradually deplete dopamine levels in the striatum over a user-defined time course – spanning weeks to months – in C57BL/6 mice. Dopamine depletions were achieved by administration of five low-dose injections (0.75μg) of 6-OHDA through an implanted intracranial bilateral cannula targeting the medial forebrain bundle. Levels of dopamine within the striatum declined linearly with successive injections, quantified using tyrosine hydroxylase immunostaining and high-performance liquid chromatography. Behavioral testing was carried out at each time point to study the onset and progression of motor impairments as a function of dopamine loss over time. We found that spontaneous locomotion, measured in an open field, was robust until ∼70% of striatal dopamine was lost. Beyond this point, additional dopamine loss caused a sharp decline in motor performance, reaching a final level comparable to that of acutely depleted mice. Similarly, although rearing behavior was more sensitive to dopamine loss and declined linearly as a function of dopamine levels, it eventually declined to levels similar to those seen in acutely depleted mice. In contrast, motor coordination, measured on a vertical pole task, was only moderately impaired in gradually depleted mice, despite severe impairments observed in acutely depleted mice. These results demonstrate the importance of the temporal profile of dopamine loss on the magnitude and progression of behavioral impairments. Our gradual depletion model thus establishes a new paradigm with which to study how circuits respond and adapt to dopamine loss over time, information which could uncover important cellular events during the prodromal phase of PD that ultimately impact the presentation or treatability of behavioral symptoms.