Chronic High-frequency Stimulation of the Subthalamic Nucleus Induces a Sustained Inhibition of Serotonergic System via Loss of Cell Phenotype

Deep brain stimulation (DBS) of the subthalamic nucleus (STN) has become a standard treatment in Parkinson’s disease (PD). However, in a considerable number of patients debilitating psychiatric side-effects occur. Recent research has revealed that external stimuli can alter the neurotransmitters’ homeostasis in neurons, which is known as “neurotransmitter respecification”. Herein, we addressed if neurotransmitter respecification could be a mechanism by which DBS suppresses the serotonergic function in the dorsal raphe nucleus (DRN) leading to mood changes. We infused transgenic 5-HT-Cre (ePet-cre) mice with AAV viruses to achieve targeted expression of eYFP and the genetically encoded calcium indicator GCaMP6s in the DRN prior to methyl-4phenyl-1,2,3,6-tetrahydropyridine (MPTP) treatment. Mice received bilateral DBS electrodes in the STN and an optic fiber in the DRN for Ca2+ photometry. MPTP treated mice demonstrated behavioral and histological PD phenotype, whereas all STN-DBS animals exhibited an increased immobility time in the forced swim test, reduced Ca2+ activity, and loss of TPH2 expression in the DRN. Given the prominent role of Ca2+ transients in mediating neurotransmitter respecification, these results suggest a chronic loss of serotonergic phenotype in the DRN following STN-DBS. These findings indicate that loss of 5-HT cell phenotype may underlie the unwanted depressive symptoms following STN-DBS.

A Mouse Model of Nigrostriatal Dopaminergic Axonal Degeneration As a Tool for Testing Neuroprotectors

Degeneration of nigrostriatal dopaminergic neurons in Parkinsons disease begins from the axonal terminals in the striatum and, then, in retrograde fashion, progresses to the cell bodies in the substantia nigra. Investigation of the dynamics of axonal terminal degeneration may help in the identification of new targets for neuroprotective treatment and be used as a tool for testing potential drugs. We have shown that the degeneration rate of dopaminergic axonal terminals changes over time, and that the striatal dopamine concentration is the most sensitive parameter to the action of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). This model was validated using neuroprotectors with well-known mechanisms of action: the dopamine transporter inhibitor nomifensine and SEMAX peptide that stimulates the secretion of endogenous neurotrophic factors or acts as an antioxidant. Nomifensine was shown to almost completely protect dopaminergic fibers from the toxic effect of MPTP and maintain the striatal dopamine concentration at the control level. However, SEMAX, slightly but reliably, increased striatal dopamine when administered before MPTP treatment, which indicates that it is more effective as an inductor of endogenous neurotrophic factor secretion rather than as an antioxidant.

Synaptic potentiation of anterior cingulate cortex contributes to chronic pain of Parkinson’s disease

Parkinson’s disease (PD) is a multi-system neurodegenerative disorder. Patients with PD often suffer chronic pain. In the present study, we investigated motor, sensory and emotional changes in three different PD mice models. We found that 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-treatment caused significant changes in all measurements. Mechanical hypersensitivity of PD model induced by MPTP peaked at 3 days and persisted for at least 14 days. Using Fos transgenic mice, we found that neurons in the anterior cingulate cortex (ACC) were activated after MPTP treatment. Inhibiting ACC by bilateral microinjection of muscimol significantly reduced mechanical hypersensitivity and anxiety-like responses. By contrast, MPTP induced motor deficit was not affected, indicating ACC activity is mostly responsible for sensory and emotional changes. We also investigated excitatory synaptic transmission and plasticity using brain slices of MPTP treated animals. While L-LTP was blocked or significantly reduced. E-LTP was not significantly affected in slices of MPTP treated animals. LTD induced by repetitive stimulation was not affected. Furthermore, we found that paired-pulse facilitation and spontaneous release of glutamate were also altered in MPTP treated animals, suggesting presynaptic enhancement of excitatory transmission in PD. Our results suggest that ACC synaptic transmission is enhanced in the animal model of PD, and cortical excitation may play important roles in PD related pain and anxiety.

Cardiac tyrosine hydroxylase activation and MB-COMT in dyskinetic monkeys

The impact of age-associated disorders is increasing as the life expectancy of the population increments. Cardiovascular diseases and neurodegenerative disorders, such as Parkinson’s disease, have the highest social and economic burden and increasing evidence show interrelations between them. Particularly, dysfunction of the cardiovascular nervous system is part of the dysautonomic symptoms of Parkinson’s disease, although more studies are needed to elucidate the role of cardiac function on it. We analyzed the dopaminergic system in the nigrostriatal pathway of Parkinsonian and dyskinetic monkeys and the expression of some key proteins in the metabolism and synthesis of catecholamines in the heart: total and phosphorylated (phospho) tyrosine hydroxylase (TH), and membrane (MB) and soluble (S) isoforms of catechol-O-methyl transferase (COMT). The dopaminergic system was significantly depleted in all MPTP-intoxicated monkeys. MPTP- and MPTP + L-DOPA-treated animals also showed a decrease in total TH expression in both right (RV) and left ventricle (LV). We found a significant increase of phospho-TH in both groups (MPTP and MPTP + L-DOPA) in the LV, while this increase was only observed in MPTP-treated monkeys in the RV. MB-COMT analysis showed a very significant increase of this isoform in the LV of MPTP- and MPTP + L-DOPA-treated animals, with no significant differences in S-COMT levels. These data suggest that MB-COMT is the main isoform implicated in the cardiac noradrenergic changes observed after MPTP treatment, suggesting an increase in noradrenaline (NA) metabolism. Moreover, the increase of TH activity indicates that cardiac noradrenergic neurons still respond despite MPTP treatment.

Acute MPTP Treatment Impairs Dendritic Spine Density in the Mouse Hippocampus

Among the animal models of Parkinson’s disease (PD), the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-lesioned mouse model has shown both dopaminergic (DA) damage and related motor control defects, as observed in patients with PD. Recent studies have suggested that the DA system interacts with the synaptic plasticity of the hippocampus in PD. However, little is known about how alterations in the hippocampal structural plasticity are affected by the DA damage in MPTP-lesioned models. In the present study, we investigated alterations in dendritic complexity and spine density in the mouse hippocampus following acute MPTP treatment (22 mg/kg, intraperitoneally, four times/day, 2-h intervals). We confirmed that acute MPTP treatment significantly decreased initial motor function and persistently reduced the number of tyrosine hydroxylase-positive DA neurons in the substantia nigra. Golgi staining showed that acute MPTP treatment significantly reduced the spine density of neuronal dendrites in the cornu ammonis 1 (CA1) apical/basal and dentate gyrus (DG) subregions of the mouse hippocampus at 8 and 16 days after treatment, although it did not affect dendritic complexity (e.g., number of crossing dendrites, total dendritic length, and branch points per neuron) in both CA1 and DG subregions at all time points after treatment. Therefore, the present study provides anatomical evidence that acute MPTP treatment affects synaptic structure in the hippocampus during the late phase after acute MPTP treatment in mice, independent of any changes in the dendritic arborization of hippocampal neurons. These findings offer data for the ability of the acute MPTP-lesioned mouse model to replicate the non-nigrostriatal lesions of clinical PD.

Tanshinone IIA protects against dopaminergic neuron degeneration via regulation of DJ-1 and Nrf2/HO-1 pathways in a rodent model of Parkinson’s disease

Purpose: To study the potential neuroprotective effects of tanshinone IIA, a diterpene quinone, in an experimental model of 1-methyl-4-phenyl-1, 2, 3, 6-tetrahydropyridine (MPTP)-induced Parkinson disease (PD). Methods: Mice (C57BL/6) were administered freshly-prepared MPTP at a dose of 20 mg/kg body weight intraperitoneally, 4 times at 2-h intervals, to induce PD. Doses of 12.5, 25 and 50 mg/kg tanshinone IIA were administered to the mice as treatments for PD. Pole and Rota-rod tests were carried out to assess muscular coordination and bradykinesia. Protein expressions, reactive oxygen species (ROS) and malonaldehyde and other parameters were evaluated. Results: Tanshinone IIA at doses of 12.5, 25 and 50 mg/kg reduced deficits in muscular coordination and improved learning ability of MPTP-treated mice. It also reduced loss of tyrosine hydroxylase (TH)- positive neurons following MPTP-induction. Tanshinone IIA regulated apoptotic pathway proteins, i.e., Bax and Bcl-2, and inhibited the translocation of Cyt C to the mitochondria. Oxidative stress induced by MPTP was significantly inhibited by tanshinone IIA via up-regulation of DJ-1/Nrf2 /HO-1 expression and reduction of ROS and MDA levels. Brain tissue total glutathione content was increased by tanshinone IIA treatment. Conclusion: Tanshinone IIA effectively improves antioxidant status and reduces neuronal loss following MPTP treatment. These results indicate that tanshinone IIA exerts protective effects in MPTPinduced PD in mice. Thus, tanshinone IIA has a good potential for use as a therapy for PD.

Impaired Mitochondrial functions and Energy Metabolism in MPTP-induced Model of Parkinson’s Disease in Mice: Comparison of Strains and Dose Regimens

Parkinson’s disease is a multiplexed disease involving diverse symptoms and progression rate. Heterogenous diseases need an efficient animal model to enhance the understanding of the underlying mechanism. MPTP (1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine), a neurotoxin, has been widely used to replicate the pathophysiology of PD in rodents, but its effect on energy metabolic perturbation is limited. Moreover, susceptibility to different dosage regime of MPTP varies among mice strains. Thus, herein the present study compares the effect of acute and sub-acute MPTP dosage regimes on mitochondrial functions in terms of mitochondrial respiratory chain enzymes, mitochondrial swelling and membrane potential in C57BL/6 and Balb/c mice. In addition, activities of enzymes involved in energy metabolic pathways were also studied along with behaviour and neurochemical alterations. The results showed that acute dose of MPTP in C57BL/6 mice had more profound effect on the enzyme activities of electron transport chain complexes. Further, the activity of MAO-A and MAO-B was increased following acute and sub-acute MPTP administration in C57BL/6 mice. However, no significant change was observed in Balb/c strain. Acute MPTP treatment resulted in decreased mitochondrial membrane potential along with swelling of mitochondria in C57BL/6 mice. In addition, perturbations were observed in hexokinase and pyruvate dehydrogenase of glycolysis pathway and citrate synthase, aconitase, isocitrate dehydrogenase and fumarase of TCA cycle. Moreover, acute MPTP led to pronounced depletion in neostriatal dopamine levels in C57BL/6 than in Balb/c mice. Behavioral tests such as open field, Narrow beam walk test and footprint test showed that locomotor activity was drastically reduced as an acute effect of MPTP in C57BL/6 mice strain. Therefore, these results consistently showed that acute MPTP treatment in C57BL/6 strain had severe mitochondrial dysfunctions, perturbed energy metabolic pathways, altered neurotransmission and motor defects as compared to Balb/c strain. Thus, the findings suggest that the dose and strain of mice need to consider for pre-clinical studies targeting mitochondrial dysfunctions in MPTP-induced Parkinson’s disease model.

Cardiac Tyrosine Hydroxylase Activation and Mb-Comt in Dyskinetic Monkeys

The impact of age-associated disorders is increasing as the life expectancy of the population increments. Cardiovascular diseases and neurodegenerative disorders, such as Parkinson’s disease, have the highest social and economic burden and increasing evidence show interrelations between them. Particularly, dysfunction of the cardiovascular nervous system is part of the dysautonomic symptoms of Parkinson’s disease, although more studies are needed to elucidate the role of cardiac function on it. We analyzed the dopaminergic system in the nigrostriatal pathway of Parkinsonian and dyskinetic monkeys and the expression of some key proteins in the metabolism and synthesis of catecholamines in the heart: total and phosphorylated (phospho) tyrosine hydroxylase (TH), and membrane (MB) and soluble (S) isoforms of catechol-O-methyl transferase (COMT). The dopaminergic system was significantly depleted in all MPTP-intoxicated monkeys. MPTP- and MPTP+L-DOPA-treated animals also showed a decrease in total TH expression in both right (RV) and left ventricle (LV). We found a significant increase of phospho-TH in both groups (MPTP and MPTP+L-DOPA) in the LV, while this increase was only observed in MPTP-treated monkeys in the RV. MB-COMT analysis showed a very significant increase of this isoform in the LV of MPTP- and MPTP+L-DOPA-treated animals, with no significant differences in S-COMT levels. These data suggest that MB-COMT is the main isoform implicated in the cardiac noradrenergic changes observed after MPTP treatment, suggesting an increase in NA metabolism. Moreover, the increase of TH activity indicates that cardiac noradrenergic neurons still respond despite MPTP treatment.

Pretreatment with Human Lactoferrin Had a Positive Effect on the Dynamics of Mouse Nigrostriatal System Recovery after Acute MPTP Exposure

We studied the effect of human lactoferrin (hLf) on degenerative changes in the nigrostriatal system and associated behavioral deficits in the animal model of Parkinson disease. Nigrostriatal dopaminergic injury was induced by single administration of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP; 40 mg/kg) to five-month-old C57Bl/6 mice. Behavioral disturbances were assessed in the open field and rotarod tests and by the stride length analysis. Structural deficits were assessed by the counts of tyrosine hydroxylase (TH)-immunoreactive neurons in the substantia nigra and optical density (OD) of TH-immunolabeled fibers in the striatum. Acute MPTP treatment induced long-term behavioral deficit and degenerative changes in the nigrostriatal system. Pretreatment with hLf prevented body weight loss and promoted recovery of motor functions and exploratory behavior. Importantly, OD of TH-positive fibers in the striatum of mice treated with hLf almost returned to normal, and the number of TH-positive cells in the substantia nigra significantly increased on day 28. These results indicate that hLf produces a neuroprotective effect and probably stimulates neuroregeneration under conditions of MPTP toxicity in our model. A relationship between behavioral deficits and nigrostriatal system disturbances at delayed terms after MPTP administration was found.

Cardiac Tyrosine Hydroxylase Activation and MB-COMT in Dyskinetic Monkeys

Background: The impact of age-associated disorders is increasing as the life expectancy of the population increments. Cardiovascular diseases and neurodegenerative disorders, such as Parkinson’s disease, have the highest social and economic burden and increasing evidence show interrelations between them. Particularly, dysfunction of the cardiovascular nervous system is part of the dysautonomic symptoms of Parkinson’s disease, although more studies are needed to elucidate the role of cardiac function on it . Methods: We analyzed the dopaminergic system in the nigrostriatal pathway of Parkinsonian and dyskinetic monkeys and the expression of some key proteins in the metabolism and synthesis of catecholamines in the heart: total and phosphorylated (phospho) tyrosine hydroxylase (TH), and membrane (MB) and soluble (S) isoforms of catechol-O-methyl transferase (COMT). Results: The number of dopaminergic neurons in the Substantia Nigra pars compacta and the optical density of TH+ fibers and dopamine transporter in the striatum were significantly decreased in all MPTP-intoxicated monkeys. MPTP- and MPTP+L-DOPA-treated animals also showed a decrease in total TH expression in both right (RV) and left ventricle (LV). We found a significant increase of phospho-TH in both groups (MPTP and MPTP+L-DOPA) in the LV, while this increase was only observed in MPTP-treated monkeys in the RV. MB-COMT analysis showed a very significant increase of this isoform in the LV of MPTP- and MPTP+L-DOPA-treated animals. However, we found no significant differences in S-COMT levels. These data suggest that MB-COMT is the main isoform implicated in the cardiac noradrenergic changes observed after MPTP treatment, suggesting an increase in NA metabolism. Moreover, the increase of TH activity indicates that cardiac noradrenergic neurons still respond despite MPTP treatment. Conclusions: These results could help to elucidate the possible role of alterations in the catecholaminergic system that can contribute to noradrenergic deficiency in the hearts of PD patients. Therefore, this information might be relevant to clinical field, contributing to the therapeutic design of the disease.