Rotenone Administration Research Articles

Characterisation of a novel model of Parkinson's disease by intra-striatal infusion of the pesticide rotenone

One of the most promising models of Parkinson's disease to have emerged in recent years is one in which the pesticide, rotenone, is administered systemically to laboratory rats. However, this model is associated with peripheral toxicity and high mortality rates which impede its widespread application in preclinical drug discovery research. This study sought to determine if administration of rotenone directly into the rat striatum could also mimic the motor dysfunction and neuropathological features of the human condition while overcoming the toxicity associated with systemic administration. Male Sprague–Dawley rats were infused with control or rotenone solutions into the striatum. The effect of the pesticide on body weight and spontaneous motor function (Corridor, Stepping and Whisker Tests) was assessed ante mortem, and its effect on nigrostriatal integrity (quantitative tyrosine hydroxylase immunohistochemistry), α-synuclein expression (quantitative α-synuclein immunohistochemistry), and striatal neurotransmitter content (HLPC for dopamine, GABA and noradrenaline) was assessed post mortem. Intra-striatal infusion of rotenone had no detrimental effect on the rats' body weight but caused significant impairments in contralateral motor function. Neuropathologically, rotenone caused significant nigrostriatal degeneration and selective loss of dopamine from the striatum but there was no evidence of any change in α-synuclein expression in the rotenone-infused rats. This study shows intra-striatal rotenone to be capable of modelling some of the main behavioural and neuropathological features of human Parkinsonism, while being less toxic than its systemic counterpart. Thus, this model may prove to be useful in future Parkinson's disease drug discovery programmes.

Parkinsonian Rotenone Mouse Model: Reevaluation of Long-Term Administration of Rotenone in C57BL/6 Mice

Chronic systemic exposure of Lewis rats to rotenone produced many features of Parkinson's disease (PD), including nigrostriatal dopamine (DA) neurodegeneration and the formation of cytoplasmic inclusions in nigral DA neurons. We also reported that chronic oral administration of rotenone at 30 mg/kg for 28 d caused specific nigrostriatal DA neurodegeneration in C57BL/6 mice. To establish a PD model more suitable for evaluating nigrostriatal DA neurodegeneration, the present study has been designed to assess the neurotoxicity of rotenone after daily oral administration at 30 or 100 mg/kg for 56 d in C57BL/6 mice. The survival rate of rotenone-treated mice at 30 mg/kg did not change from 28 to 56 d, although the survival rate of rotenone-treated mice at 30 mg/kg was decreased to about 70% within one week. The survival rate of the rotenone-treated mice at 100 mg/kg was suddenly decreased after 28 d, and finally to about 15% at 56 d. Rotenone at 30 mg/kg, but not 100 mg/kg, for 28 d caused a significant loss of tyrosine hydroxylase (TH)-positive neurons in the substantia nigra. Rotenone at 100 mg/kg caused a highly variable loss of TH-positive neurons among individual mice. Rotenone at 30 mg/kg for 56 d caused a significant loss of TH-positive neurons and behavioral impairment. In addition, α-synuclein immunoreactivity was increased in surviving TH-positive neurons in a time-dependent manner. Thus, this protocol for chronic administration of rotenone at 30 mg/kg for 56 d is more useful for understanding the mechanism of DA neurodegeneration.

Both Stereoselective (R)- and (S)-1-Methyl-1,2,3,4-tetrahydroisoquinoline Enantiomers Protect Striatal Terminals Against Rotenone-Induced Suppression of Dopamine Release

1-Methyl-1,2,3,4-tetrahydroisoquinoline (1MeTIQ) is present in the human and rodent brain as a mixture of stereospecific (R)- and (S)-1MeTIQ enantiomers. The racemate, (R,S)-1MeTIQ, exhibits neuroprotective activity as shown in the earlier study by the authors, and In addition, it was suggested to play a crucial physiological role in the mammalian brain as an endogenous regulator of dopaminergic activity. In this article, we investigated the influence of stereospecific enantiomers of 1MeTIQ, (R)- and (S)-1MeTIQ (50 mg/kg i.p.) on rotenone-induced (3 mg/kg s.c.) behavioral and neurochemical changes in the rat. In behavioral study, in order to record dynamic motor function of rats, we measured locomotor activity using automated locomotor activity boxes. In biochemical studies, we analyzed in rat striatum the concentration of dopamine (DA) and its metabolites: intraneuronal DOPAC, extraneuronal 3-MT, and final HVA using HPLC with electrochemical detection. Otherwise, DA release was estimated by in vivo microdialysis study. The behavioral study has demonstrated that both acute and repeated (3 times) rotenone administration unimportantly depressed a basic locomotor activity in rat. (R)- and (S)-1MeTIQ stereoisomers (50 mg/kg i.p.) produced a modest behavioral activation both in naïve and rotenone-treated rats. The data from ex vivo neurochemical experiments have shown stereospecificity of 1MeTIQ enantiomers in respect of their effects on DA catabolism. (R)-1MeTIQ significantly increased both the level of the final DA metabolite, HVA (by about 70%), and the rate of DA metabolism (by 50%). In contrast to that, (S)-1MeTIQ significantly depressed DOPAC, HVA levels (by 60 and 40%, respectively), and attenuated the rate of DA metabolism (by about 60%). On the other hand, both the enantiomers increased the concentrations of DA and its extraneuronal metabolite, 3-MT in rat striatum. In vivo microdialysis study has shown that repeated but not acute administration of rotenone produced a deep and significant functional impairment of striatal DA release. Both (R)- and (S)- stereospecific enantiomers of 1MeTIQ antagonized rotenone-induced suppression of DA release; however, the effect of (R)-1MeTIQ was more strongly expressed in microdialysis study. In conclusion, we suggest that both chiral isomers of 1MeTIQ offer neuroprotection against rotenone-induced disturbances in the function of dopaminergic neurons and (R,S)-1MeTIQ will be useful as a drug with marked neuroprotective activity in the brain.

Oral Administration of Rotenone using a Gavage and Image Analysis of Alpha-synuclein Inclusions in the Enteric Nervous System

Oral Administration of Rotenone using a Gavage and Image Analysis of Alpha-synuclein Inclusions in the Enteric Nervous System

Oral Administration of Rotenone using a Gavage and Image Analysis of Alpha-synuclein Inclusions in the Enteric Nervous System

In Parkinson's disease (PD) patients, the associated pathology follows a characteristic pattern involving inter alia the enteric nervous system (ENS) (1,2), the olfactory bulb (OB), the dorsal motor nucleus of the vagus (DMV)(3), the intermediolateral nucleus of the spinal cord (4) and the substantia nigra, providing the basis for the neuropathological staging of the disease(4,5). The ENS and the OB are the most exposed nervous structures and the first ones to be affected. Interestingly, PD has been related to pesticide exposure(6-8). Here we show in detail two methods used in our previous study (9). In order to analyze the effects of rotenone acting locally on the ENS, we administered rotenone using a gavage to one-year old C57/BL6 mice. Rotenone is a widely used pesticide that strongly inhibits mitochondrial Complex I (10). It is highly lipophylic and poorly absorbed in the gastrointestinal tract (11). Our results showed that the administration of 5 mg/kg of rotenone did not inhibit mitochondrial Complex I activity in the muscle or the brain. Thus, suggesting that using our administration method rotenone did not cross the hepatoportal system and was acting solely on the ENS. Here we show a method to administer pesticides using a gavage and the image analysis protocol used to analyze the effects of the pesticide in alpha-synuclein accumulation in the ENS. The first part shows a method that allows intragastric administration of pesticides (rotenone) at a desired precise concentration. The second method shows a semi-automatic image analysis protocol to analyze alpha-synuclein accumulation in the ENS using an image analysis software.

Development of a high-throughput screening assay for cytoprotective agents in rotenone-induced cell death

Parkinson’s disease (PD) is a neurodegenerative disease featured by selective loss of substantia nigra neurons. Rotenone administration in animals induces neurodegeneration accompanied by α-synuclein-positive Lewy body-like inclusions, recapturing typical histopathological features of PD. In an effort to screen for small-molecule agents to reverse rotenone-induced cytotoxicity, we developed and validated a sensitive and robust assay with neuroblastoma SK-N-SH cells. This assay was amenable to a high-throughput screening format with Z′ factor of 0.56. Robotic screening of a bioactive compound library led to the identification of carnosic acid that can effectively protect cells from rotenone treatment. Using a high-content image-based assay and Western blot analysis, we demonstrated that carnosic acid protects cells from rotenone stress by significant induction of HSP70 expression. Therefore, the assay reported here can be used to identify novel cytoprotective agents for clinical therapeutics of PD.

Resurrecting Hope for Antioxidant Treatment of Cardiovascular Disease

Oxidative stress has been associated with cardiovascular disease and the development of hypertension in both humans and in animal models. Antioxidant therapy has been successful in ameliorating disease in animals, but the results from human studies have been largely disappointing. Despite these negative results, a wealth of evidence suggests that oxidative stress is involved in the development of cardiovascular disease both in humans and animal models. Because global attenuation of cellular reactive oxygen species (ROS) may not be advantageous, more recent therapeutic strategies target specific subcellular sources of oxidative stress. Most studies implicate plasmalemmal membrane-bound NADPH oxidase as a key mediator of ROS generation in cardiovascular disease. However, in the current issue of Circulation Research , Dikalova et al1 demonstrate that specific attenuation of mitochondrial derived oxidants prevents the development of angiotensin II–induced hypertension in the mouse. Perhaps more interesting is the mechanism: inhibition of a mitochondrial ROS-triggered activation of NADPH oxidase, interrupting a vicious cycle of ROS production. The novel approach of targeting the inciting source of ROS (mitochondria) to prevent accelerated oxidant production may hold more promise for humans by eliminating pathological but not homeostatic sources of ROS. Seminal observations by Harrison et al2 showed evidence of increased oxidative stress within the systemic vasculature in an Angiotensin II–induced rat model of hypertension, piquing interest in a possible etiologic role for ROS in this condition. In animal models, stimulating ROS production can induce hypertension,2,3 whereas treatment with a variety of antioxidants including vitamin C and E, or targeted inhibition of enzymatic oxidant production, can reduce blood pressure.3–6 Hypertensive patients demonstrate increased NAD(P)H oxidase activity7 with elevated levels of hydrogen peroxide in plasma.8 Antioxidant treatment leads to enhanced vasodilation in both forearm and coronary arteries of human hypertensive subjects,9,10 indicating a functional vasomotor …

Successful Amelioration of Mitochondrial Optic Neuropathy Using the Yeast NDI1 Gene in a Rat Animal Model

BackgroundLeber's hereditary optic neuropathy (LHON) is a maternally inherited disorder with point mutations in mitochondrial DNA which result in loss of vision in young adults. The majority of mutations reported to date are within the genes encoding the subunits of the mitochondrial NADH-quinone oxidoreductase, complex I. Establishment of animal models of LHON should help elucidate mechanism of the disease and could be utilized for possible development of therapeutic strategies.Methodology/Principal FindingsWe established a rat model which involves injection of rotenone-loaded microspheres into the optic layer of the rat superior colliculus. The animals exhibited the most common features of LHON. Visual loss was observed within 2 weeks of rotenone administration with no apparent effect on retinal ganglion cells. Death of retinal ganglion cells occurred at a later stage. Using our rat model, we investigated the effect of the yeast alternative NADH dehydrogenase, Ndi1. We were able to achieve efficient expression of the Ndi1 protein in the mitochondria of all regions of retinal ganglion cells and axons by delivering the NDI1 gene into the optical layer of the superior colliculus. Remarkably, even after the vision of the rats was severely impaired, treatment of the animals with the NDI1 gene led to a complete restoration of the vision to the normal level. Control groups that received either empty vector or the GFP gene had no effects.Conclusions/SignificanceThe present study reports successful manifestation of LHON-like symptoms in rats and demonstrates the potential of the NDI1 gene therapy on mitochondrial optic neuropathies. Our results indicate a window of opportunity for the gene therapy to be applied successfully after the onset of the disease symptoms.

The exceptional properties of 9‐methyl‐β‐carboline: stimulation, protection and regeneration of dopaminergic neurons coupled with anti‐inflammatory effects

Beta-carbolines (BCs) are potential endogenous and exogenous neurotoxins that may contribute to the pathogenesis of Parkinson's disease. However, we recently demonstrated protective and stimulatory effects of 9-methyl-BC (9-me-BC) in primary dopaminergic culture. In the present study, treatment with 9-me-BC unmasked a unique tetrad of effects. First, tyrosine hydroxylase (TH) expression was stimulated in pre-existing dopa decarboxylase immunoreactive neurons and several TH-relevant transcription factors (Gata2, Gata3, Creb1, Crebbp) were up-regulated. Neurite outgrowth of TH immunoreactive (THir) neurons was likewise stimulated. The interaction with tyrosine kinases (protein kinase A and C, epidermal growth factor-receptor, fibroblast growth factor-receptor and neural cell adhesion molecule) turned out to be decisive for these observed effects. Second, 9-me-BC protected in acute toxicity models THir neurons against lipopolysaccharide and 2,9-dime-BC(+) toxicity. Third, in a chronic toxicity model when cells were treated with 9-me-BC after chronic rotenone administration, a pronounced regeneration of THir neurons was observed. Fourth, 9-me-BC inhibited the proliferation of microglia induced by toxin treatment and installed an anti-inflammatory environment by decreasing the expression of inflammatory cytokines and receptors. Finally, 9-me-BC lowered the content of alpha-synuclein protein in the cultures. The presented results warrant the exploration of 9-me-BC as a novel potential anti-parkinsonian medication, as 9-me-BC interferes with several known pathogenic factors in Parkinson's disease as outlined above. Further investigations are currently under way.

α-Synuclein Transgenic Mice Reveal Compensatory Increases in Parkinson's Disease-Associated Proteins DJ-1 and Parkin and Have Enhanced α-Synuclein and PINK1 Levels After Rotenone Treatment

Parkinson's disease (PD) is a severe neurodegenerative disorder characterised by loss of dopaminergic neurons of the substantia nigra. The pathological hallmarks are cytoplasmic inclusions termed Lewy bodies consisting primarily of aggregated alpha-synuclein (alphaSN). Different lines of transgenic mice have been developed to model PD but have failed to recapitulate the hallmarks of this disease. Since treatment of rodents with the pesticide rotenone can reproduce nigrostriatal cell loss and other features of PD, we aimed to test chronic oral administration of rotenone to transgenic mice over-expressing human alphaSN with the A53T mutation. Initial assessment of this transgenic line for compensatory molecular changes indicated decreased brain beta-synuclein expression and significantly increased levels of the PD-associated oxidative stress response protein, DJ-1, and the E3 ubiquitin ligase enzyme, Parkin. Rotenone treatment of 30 mg/kg for 25 doses over a 35-day period was tolerated in the transgenic mice and resulted in decreased spontaneous locomotor movement and increased cytoplasmic alphaSN expression. The mitochondrial Parkinson's-associated PTEN-induced kinase 1 protein levels were also increased in transgenic mouse brain after rotenone treatment; there was no change in brain dopamine levels or nigrostriatal cell loss. These hA53T alphaSN transgenic mice provide a useful model for presymptomatic Parkinson's features and are valuable for study of associated compensatory changes in early Parkinson's disease stages.

Progression of Parkinson’s disease pathology is reproduced by intragastric administration of rotenone in mice

AbstractIn Parkinson’s disease patients pathology follows a characteristic pattern involving inter alia the enteric nervous system, the dorsal motor nucleus of the vagus, the intermediolateral nucleus of the spinal cord and the substantia nigra, providing the basis for the neuropathological staging of the disease. Here we report that intragastrically administered rotenone, a commonly used pesticide that inhibits mitochondrial Complex I, is able to reproduce PD pathological staging as found in patients. Our results show that low doses of chronically and intragastrically administered rotenone induce PD pathology in all the above-mentioned nervous system structures in wild-type mice. Interestingly, HPLC analysis showed no rotenone levels in the systemic blood or the central nervous system (CNS) (detection limit [rotenone]<20nM), indicating that rotenone is detoxified by the liver. These alterations are sequential, appearing only in synaptically connected nervous structures, treatment time-dependent and accompanied by inflammatory signs and motor dysfunctions. These results strongly suggest that the local effect of pesticides on the ENS might be sufficient to induce PD progression and to reproduce the neuroanatomical and neurochemical features of PD staging. Thus, providing new insight into how environmental factors could trigger PD and pointing out a transsynaptic mechanism by which PD might propagate to and through the CNS.

Progression of Parkinson's Disease Pathology Is Reproduced by Intragastric Administration of Rotenone in Mice

In patients with Parkinson's disease (PD), the associated pathology follows a characteristic pattern involving inter alia the enteric nervous system (ENS), the dorsal motor nucleus of the vagus (DMV), the intermediolateral nucleus of the spinal cord and the substantia nigra, providing the basis for the neuropathological staging of the disease. Here we report that intragastrically administered rotenone, a commonly used pesticide that inhibits Complex I of the mitochondrial respiratory chain, is able to reproduce PD pathological staging as found in patients. Our results show that low doses of chronically and intragastrically administered rotenone induce alpha-synuclein accumulation in all the above-mentioned nervous system structures of wild-type mice. Moreover, we also observed inflammation and alpha-synuclein phosphorylation in the ENS and DMV. HPLC analysis showed no rotenone levels in the systemic blood or the central nervous system (detection limit [rotenone]<20 nM) and mitochondrial Complex I measurements showed no systemic Complex I inhibition after 1.5 months of treatment. These alterations are sequential, appearing only in synaptically connected nervous structures, treatment time-dependent and accompanied by inflammatory signs and motor dysfunctions. These results strongly suggest that the local effect of pesticides on the ENS might be sufficient to induce PD-like progression and to reproduce the neuroanatomical and neurochemical features of PD staging. It provides new insight into how environmental factors could trigger PD and suggests a transsynaptic mechanism by which PD might spread throughout the central nervous system.

Stereotaxical Infusion of Rotenone: A Reliable Rodent Model for Parkinson's Disease

A clinically-related animal model of Parkinson's disease (PD) may enable the elucidation of the etiology of the disease and assist the development of medications. However, none of the current neurotoxin-based models recapitulates the main clinical features of the disease or the pathological hallmarks, such as dopamine (DA) neuron specificity of degeneration and Lewy body formation, which limits the use of these models in PD research. To overcome these limitations, we developed a rat model by stereotaxically (ST) infusing small doses of the mitochondrial complex-I inhibitor, rotenone, into two brain sites: the right ventral tegmental area and the substantia nigra. Four weeks after ST rotenone administration, tyrosine hydroxylase (TH) immunoreactivity in the infusion side decreased by 43.7%, in contrast to a 75.8% decrease observed in rats treated systemically with rotenone (SYS). The rotenone infusion also reduced the DA content, the glutathione and superoxide dismutase activities, and induced alpha-synuclein expression, when compared to the contralateral side. This ST model displays neither peripheral toxicity or mortality and has a high success rate. This rotenone-based ST model thus recapitulates the slow and specific loss of DA neurons and better mimics the clinical features of idiopathic PD, representing a reliable and more clinically-related model for PD research.

Neuroprotective effect of the antiparkinsonian drug pramipexole against nigrostriatal dopaminergic degeneration in rotenone-treated mice

Pramipexole, an agonist for dopamine (DA) D2/D3-receptors, has been used to treat both early and advanced Parkinson's disease (PD). In this study, we examined the effect of pramipexole on DA neurons in a PD model of C57BL/6 mice, which were treated with rotenone (30 mg/kg, p.o.) daily for 28 days. Pramipexole (1 mg/kg, i.p.) was injected daily 30 min before each oral administration of rotenone. Chronic oral administration of rotenone caused a loss of DA neurons in the substantia nigra pars compacta (SNpc), motor deficits and the up-regulation of α-synuclein immunoreactivity in some surviving DA neurons. Pramipexole inhibited rotenone-induced DA neuronal death and motor deficits, and reduced immunoreactivity for α-synuclein. In addition, pramipexole inhibited the in vitro oligomerization of human wild-type α-synuclein by H 2O 2 plus cytochrome c. To examine the neuroprotective effect of pramipexole against oxidative stress, we used a DJ-1-knockdown SH-SY5Y cell line and electron spin resonance (ESR) spectrometry. Simultaneous treatment with H 2O 2 and pramipexole resulted in the significant protection of DJ-1-knockdown cells against cell death in a concentration-dependent manner. A high concentration of pramipexole directly scavenged hydroxyl radical ( OH) generated from H 2O 2 and Fe 2+. Furthermore, pramipexole increased Bcl-2 immunoreactivity in DA neurons in the SNpc. These results suggest that pramipexole may protect DA neurons against exposure to rotenone by chronic oral administration, and this effect is mediated by multiple functions including scavenging of OH and induction of Bcl-2 protein.

Chronic Administration with Rotenone does not Enhance MPTP Neurotoxicity in C57BL/6 Mice

Systematic administration of rotenone as one of pesticides is known to produce degeneration of nigral dopaminergic neurons and motor deficits in experimental animals. Here, we investigated to determine whether systematic administration of rotenone causes the increased susceptibility in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-treated mice. Rotenone was injected into MPTP-treated mice over a period of 4 weeks. Thereafter, we evaluated the effect of rotenone 1, 3, and 6 weeks after the cessation of treatment with rotenone. In the present study with HPLC analysis, rotenone did not enhance MPTP-induced dopaminergic neurotoxicity in mice. Furthermore, MPTP + rotenone (9 mg/kg)-treated mice exhibit a significant loss of motor activity 1 day after the cessation of treatment with rotenone, However, no significant change of motor activity was found in MPTP-treated and MPTP + rotenone (9 mg/kg)-treated animals 6 weeks after the cessation of treatment with 0.5% carboxymethyl cellulose or rotenone. Our Western blot analysis study demonstrated that the change of tyrosine hydroxylase and glial fibrillary acidic protein protein levels in MPTP-treated mice was similar than that in MPTP + rotenone-treated animals. These results suggest that rotenone did not enhance MPTP neurotoxicity in mice. Our findings suggest that rotenone is not a reliable model for PD. Thus, our findings provide further valuable information for the pathogenesis of PD for exposure to agricultural pesticides.

Valproic Acid is Neuroprotective in the Rotenone Rat Model of Parkinson’s Disease: Involvement of α-Synuclein

Valproic acid (VPA), an established antiepileptic and antimanic drug, has recently emerged as a promising neuroprotective agent. Among its many cellular targets, VPA has been recently demonstrated to be an effective inhibitor of histone deacetylases. Accordingly, we have adopted a schedule of dietary administration (2% VPA added to the chow) that results in a significant inhibition of histone deacetylase activity and in an increase of histone H3 acetylation in brain tissues of 4 weeks-treated rats. We have tested this schedule of VPA treatment in an animal model of Parkinson's disease (PD), in which degeneration of nigro-striatal dopaminergic neurons is obtained through sub-chronic administration of the mitochondrial toxin, rotenone, via osmotic mini pumps implanted to rats. The decrease of the dopaminergic marker tyrosine hydroxylase in substantia nigra and striatum caused by 7 days toxin administration was prevented in VPA-fed rats. VPA treatment also significantly counteracted the death of nigral neurons and the 50% drop of striatal dopamine levels caused by rotenone administration. The PD-marker protein alpha-synuclein decreased, in its native form, in substantia nigra and striatum of rotenone-treated rats, while monoubiquitinated alpha-synuclein increased in the same regions. VPA treatment counteracted both these alpha-synuclein alterations. Furthermore, monoubiquitinated alpha-synuclein increased its localization in nuclei isolated from substantia nigra of rotenone-treated rats, an effect also prevented by VPA treatment. Nuclear localization of alpha-synuclein has been recently described in some models of PD and its neurodegenerative effect has been ascribed to histone acetylation inhibition. Thus, the ability of VPA to increase histone acetylation is a novel candidate mechanism for its neuroprotective action.

Progression of Parkinson’s disease pathology is reproduced by intragastric administration of rotenone in mice

AbstractIn Parkinson’s disease patients pathology follows a characteristic pattern involving inter alia the enteric nervous system, the dorsal motor nucleus of the vagus, the intermediolateral nucleus of the spinal cord and the substantia nigra, providing the basis for the neuropathological staging of the disease. Here we report that intragastrically administered rotenone, a commonly used pesticide that inhibits mitochondrial Complex I, is able to reproduce PD pathological staging as found in patients. Our results show that low doses of chronically and intragastrically administered rotenone induce PD pathology in all the above-mentioned nervous system structures in wild-type mice. Interestingly, HPLC analysis showed no rotenone levels in the systemic blood or the central nervous system (CNS) (detection limit [rotenone]&lt;20nM), indicating that rotenone is detoxified by the liver. These alterations are sequential, appearing only in synaptically connected nervous structures, treatment time-dependent and accompanied by inflammatory signs and motor dysfunctions. These results strongly suggest that the local effect of pesticides on the ENS might be sufficient to induce PD progression and to reproduce the neuroanatomical and neurochemical features of PD staging. Thus, providing new insight into how environmental factors could trigger PD and pointing out a transsynaptic mechanism by which PD might propagate to and through the CNS.

Progression of Parkinson’s disease pathology is reproduced by intragastric administration of rotenone in mice

AbstractIn Parkinson’s disease patients pathology follows a characteristic pattern involving inter alia the enteric nervous system, the dorsal motor nucleus of the vagus, the intermediolateral nucleus of the spinal cord and the substantia nigra, providing the basis for the neuropathological staging of the disease. Here we report that intragastrically administered rotenone, a commonly used pesticide that inhibits mitochondrial Complex I, is able to reproduce PD pathological staging as found in patients. Our results show that low doses of chronically and intragastrically administered rotenone induce PD pathology in all the above-mentioned nervous system structures in wild-type mice. Interestingly, HPLC analysis showed no rotenone levels in the systemic blood or the central nervous system (CNS) (detection limit [rotenone]&lt;20nM), suggesting that rotenone is detoxified by the liver. These alterations are sequential, appearing only in synaptically connected nervous structures, treatment time-dependent and accompanied by inflammatory signs and motor dysfunctions. These results strongly suggest that the local effect of pesticides on the ENS might be sufficient to induce PD progression and to reproduce the neuroanatomical and neurochemical features of PD staging. Thus, providing new insight into how environmental factors could trigger PD and suggesting a transsynaptic mechanism by which PD might propagate to and through the CNS.

Neurons express hemoglobin α‐ and β‐chains in rat and human brains

Hemoglobin is the oxygen carrier in vertebrate blood erythrocytes. Here we report that hemoglobin chains are expressed in mammalian brain neurons and are regulated by a mitochondrial toxin. Transcriptome analyses of laser-capture microdissected nigral dopaminergic neurons in rats and striatal neurons in mice revealed the presence of hemoglobin alpha, adult chain 2 (Hba-a2) and hemoglobin beta (Hbb) transcripts, whereas other erythroid markers were not detected. Quantitative reverse transcriptase-polymerase chain reaction (RT-PCR) analysis confirmed the expression of Hba-a2 and Hbb in nigral dopaminergic neurons, striatal gamma-aminobutyric acid (GABA)ergic neurons, and cortical pyramidal neurons in rats. Combined in situ hybridization histochemistry and immunohistochemistry with the neuronal marker neuronal nuclear antigen (NeuN) in rat brain further confirmed the presence of hemoglobin mRNAs in neurons. Immunohistochemistry identified hemoglobin alpha- and beta-chains in both rat and human brains, and hemoglobin proteins were detected by Western blotting in whole rat brain tissue as well as in cultures of mesencephalic neurons, further excluding the possibility of blood contamination. Systemic administration of the mitochondrial inhibitor rotenone (2 mg/kg/d, 7d, s.c.) induced a marked decrease in Hba-a2 and Hbb but not neuroglobin or cytoglobin mRNA in transcriptome analyses of nigral dopaminergic neurons. Quantitative RT-PCR confirmed the transcriptional downregulation of Hba-a2 and Hbb in nigral, striatal, and cortical neurons. Thus, hemoglobin chains are expressed in neurons and are regulated by treatments that affect mitochondria, opening up the possibility that they may play a novel role in neuronal function and response to injury.

Effect of centrophenoxine against rotenone-induced oxidative stress in an animal model of Parkinson's disease

Oxidative stress has been implicated in the etiology of Parkinson's disease (PD). The important biochemical features of PD, being profound deficit in dopamine (DA) content, reduced glutathione (GSH), and enhanced lipid peroxidation (LPO) in dopaminergic (DA-ergic) neurons resulting in oxidative stress, mitochondrial dysfunction and apoptosis. Rotenone-induced neurotoxicity is a well acknowledged preclinical model for studying PD in rodents as it produces selective DA-ergic neuronal degeneration. In our previous study, we have shown that chronic administration of rotenone to rats is able to produce motor dysfunction, which increases progressively with rotenone treatment and centrophenoxine (CPH) co-treatment is able to attenuate these motor defects. The present study was carried out to evaluate the antioxidant potential of CPH against rotenone-induced oxidative stress. Chronic administration of rotenone to SD rats resulted in marked oxidative damage in the midbrain region compared to other regions of the brain and CPH co-treatment successfully attenuated most of these changes. CPH significantly attenuated rotenone-induced depletion in DA, GSH and increase in LPO levels. In addition, the drug prevented the increase in nitric oxide (NO) and citrulline levels and also enhanced the activity of catalase and superoxide dismutase (SOD). Histological analysis carried out using hematoxylin and eosin staining has indicated severe damage to mid brain in comparison to cortex and cerebellum and this damage is attenuated by CPH co-treatment. Our results strongly indicate the possible therapeutic potential of centrophenoxine as an antioxidant in Parkinson's disease and other movement disorders where oxidative stress is a key player in the disease process.