Levodopa Research Articles

Mucuna pruriens Seed Extract Promotes Neurite Outgrowth via TEN-4 Dependent and Independent Mechanisms in NEURO2A Cells

Neurological diseases are one of the serious health hazards faced by mankind for decades. Neurite outgrowth is a key factor responsible for proper neuronal development. Any misplacement in the process could lead to neurological diseases like Alzheimer’s and Parkinson’s. Treatment with the available synthetic drugs imparts many difficulties to the patients due to the side effects. Compounds from natural sources can be considered as an effective replacement for this. Mucuna pruriens, used in traditional ayurvedic medicine, contains L-3,4-dihydroxy phenylalanine (L-DOPA) in its seeds, which possesses medicinal effects against neurological diseases. In this regard, seed extracts of M. pruriens originated from Thailand and India, were analyzed for their neuroprotective effects in Neuro2a cells. Hexane, ethyl acetate and ethanol extracts were found to be non-toxic to the viability of the cells. Ethanol extracts of M. pruriens of Thai origin (MTE), hexane extracts of M. pruriens of Indian origin (MIH) and ethyl acetate extracts of M. pruriens of Indian origin (MIEA) were able to induce neurite outgrowth in Neuro2a cells. Interestingly, both MTE and MIH induced neurite outgrowth dependent on Teneurin-4 (Ten-4) transmembrane protein whereas MIEA did the same independent of Ten-4, which was confirmed by real time PCR and gene silencing approach. The present study suggested that M. pruriens can be used as a potential drug in the treatment of neurological diseases as it can induce neurite outgrowth by multiple mechanisms, which will be of great use in the field of medicine.

Purification and characterization of phenoloxidase from the hemolymph of healthy and diseased Antheraea assamensis Helfer (Lepidoptera: Saturniidae): Effects of certain biological components and chemical agents on enzyme activity.

In the current study, a dimeric phenoloxidase (PO) from the hemolymph of healthy and diseased (pebrine infected) larvae of Antheraea assamensis Helfer was extracted and purified. The protein was subjected to purification using Sephacryl S-100 and CM Sepharose chromatography. The enzyme comprised of two subunits of ~76.8 and 76 kDa that showed PO activity in 6 mM l-3,4-dihydroxyphenylalanine (L-DOPA) and 8 mM catechol but not in hydroquinone. Optimum temperature for PO activity was 30°C in l-DOPA and 37°C in catechol. Optimum pH ranged from 6.8 to 7.0 in L-DOPA and 7.0-7.2 in catechol. Specific activity of the purified PO from healthy larvae was 53.9 µM/min per mg of protein per ml in L-DOPA and 50.77 µM/min per mg of protein per ml in catechol. Specific activity of PO from diseased larvae was 30.0 µM/min per mg of protein per ml in L-DOPA and 28.55 µM/min per mg of protein per ml in catechol. Purification fold was 3.27-4.21 for healthy and 2.38-2.56 for diseased fractions. The enzyme showedthe Michaelis constant (Km ) of 2.46-2.85 mM for healthy and diseased fractions in L-DOPA. In catechol Km of 9.23-17.71 mM was observed. Peptidoglycan was the best activator of purified PO from both healthy and diseased fractions. Interactions between controls and activators appeared statistically significant (F = 767.5; df = 3; P < 0.0001). Na+ , K+ , and Cu2+ increased, whereas Ca2+ , Zn2+ , Mg2+ , and Co2+ decreased PO activity. The overall interactions appeared highly significant (F = 217.0; df = 27; P < 0.0001). Kojic acid, dithiothreitol, thiourea, phenylthiourea, carbendazim, N-bromosuccinimide, N,N,N',N'-tetraacetic acid, and diethyldithiocarbamate inhibited PO activity.

Pharmacokinetic drug evaluation of CVT-301 for the treatment of Parkinson’s disease

ABSTRACTIntroduction: Levodopa (LD), in combination with a decarboxylase inhibitor, is a mainstay and the most effective therapeutic agent in the treatment of Parkinson’s disease (PD). Unfortunately, during chronic treatment with this agent, ON–OFF phenomena and dyskinesia appear. Despite the many medical treatment options available, unpredictable OFF episodes can still occur and be severe and disabling. A rescue therapy that provides a rapid and predictable ON response for patients with OFF periods would be of great value for such patients.Areas covered: CVT-301 is a self-administered dry powder aerosol inhaled formulation of LD that is being developed as a self-administered treatment for OFF periods. The PK profile of CVT-301, the efficacy, and the safety highlighted in randomized clinical trials will be reviewed.Expert opinion: CVT-301 may offer several potential advantages including increased systemic bioavailability through pulmonary absorption, rapid onset of action, avoidance of first-pass drug metabolism and less plasma-level variability.List of Abbreviations: PD: Parkinson’s disease; LD: Levodopa; CD: Carbidopa; AADC: aromatic L-amino acid decarboxylase; IR: immediate-release; FPD: fine particle dose; GI: gastrointestinal; PK: pharmacokinetic; CVs: coefficient of variation; UPDRS: Unified Parkinson’s Disease Rating Scale; AEs: adverse events; FEV: forced expiratory volume; FVC: forced vital capacity; DLCO: diffuse lung CO ; tmax: time to maximum concentration

Application of 18F-DOPA PET imaging in the diagnosis and assessment of early-stage Parkinson′s disease

Objective To investigate the clinical values of L-6-18F-fluoro-3, 4-dihydroxyphenylalanine (18F-DOPA) PET in the diagnosis and severity assessment of early-stage Parkinson′s disease (PD). Methods Thirty-eight patients (24 males, 14 females; age: 34-74 years) with early-stage PD (Hoehn-Yahr (H-Y) staging: 1-2) and 5 age-matched healthy volunteers (all males; age: 45-65 years) from July 2016 to March 2017 were included and underwent 18F-DOPA PET scan in this retrospective study. The striatal-to-occipital ratio (SOR) was calculated and compared between PD patients and healthy volunteers. The unified PD rating scale (UPDRS) Ⅲ score and H-Y staging were used to evaluate the clinical symptoms. Two-sample t test and Pearson correlation analysis were used. Results In the control group, 18F-DOPA SORs in bilateral putamen and caudate nucleus were 2.50±0.24 and 2.61±0.23, respectively. In PD group, the SORs of ipsilateral and contralateral putamen nucleus were 2.02±0.27 and 1.80±0.26 respectively, lower than those in the control group (t values: -4.006, -5.440, both P<0.01). The SORs of ipsilateral and contralateral caudate nucleus were 2.16±0.32 and 2.08±0.28 respectively, lower than those in the control group (t values: -2.990, -4.047, both P<0.01). The SORs of contralateral putamen and caudate nucleus were significantly lower than those of the ipsilateral striatum respectively (t values: -6.431, -3.837, both P<0.01). Furthermore, the SORs in the striatum (putamen and caudate nucleus) were negatively correlated with UPDRS Ⅲ score, H-Y staging, and duration of disease (r values: from -0.526 to -0.369, all P<0.05). Conclusions 18F-DOPA PET can reflect the changes in the striatum neurons, and it may be an important method in the diagnosis and assessment of early-stage PD patients. Key words: Parkinson disease; Positron-emission tomography; Tomography, X-ray computed; Levodopa; Fluorine radioisotopes

5-HT2A blockade for dyskinesia and psychosis in Parkinson’s disease: is there a limit to the efficacy of this approach? A study in the MPTP-lesioned marmoset and a literature mini-review

Virtually every patient affected by Parkinson's disease (PD) eventually requires treatment with L-3,4-dihydroxyphenylalanine (L-DOPA), which leads to complications such as dyskinesia and psychosis. Whereas blockade of serotonin 2A (5-HT2A) receptors appears to be an effective way to reduce both dyskinesia and psychosis, whether it has the potential to eliminate the two phenomena remains to be determined. In a previous study, we showed that highly selective 5-HT2A receptor blockade with EMD-281,014, at plasma levels comparable to those achieved in the clinic, reduced dyskinesia and psychosis-like behaviours (PLBs), in the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-lesioned marmoset. Here, we sought to determine whether further increasing the dose would result in greater therapeutic benefit or if maximal effectiveness was achieved at lower doses. Six MPTP-lesioned marmosets with stable dyskinesia and PLBs were administered EMD-281,014 (0.1, 1 and 10mg/kg) or vehicle in combination with L-DOPA and the effect on dyskinesia, PLBs and parkinsonism was assessed. Administration of EMD-281,014 (0.1, 1 and 10mg/kg) in combination with L-DOPA resulted in a significant reduction in the severity of dyskinesia, by up to 63%, 64% and 61% (each P < 0.001), when compared to L-DOPA/vehicle. Similarly, the addition of EMD-281,014 (0.1, 1 and 10mg/kg) to L-DOPA also significantly decreased the severity of PLBs, by up to 54%, 55% and 53% (each P < 0.001), when compared to L-DOPA/vehicle. Our results suggest that there might be a ceiling to the reduction of dyskinesia and psychosis that can be achieved through antagonism of 5-HT2A receptors.

PBF509, an Adenosine A2A Receptor Antagonist With Efficacy in Rodent Models of Movement Disorders.

Adenosine A2A receptor (A2AR) antagonists have emerged as complementary non-dopaminergic drugs to alleviate Parkinson’s disease (PD) symptomatology. Here, we characterize a novel non-xhantine non-furan A2AR antagonist, PBF509, as a potential pro-dopaminergic drug for PD management. First, PBF509 was shown to be a highly potent ligand at the human A2AR, since it antagonized A2AR agonist-mediated cAMP accumulation and impedance responses with KB values of 72.8 ± 17.4 and 8.2 ± 4.2 nM, respectively. Notably, these results validated our new A2AR-based label-free assay as a robust and sensitive approach to characterize A2AR ligands. Next, we evaluated the efficacy of PBF509 reversing motor impairments in several rat models of movement disorders, including catalepsy, tremor, and hemiparkinsonism. Thus, PBF509 (orally) antagonized haloperidol-mediated catalepsy, reduced pilocarpine-induced tremulous jaw movements and potentiated the number of contralateral rotations induced by L-3,4-dihydroxyphenylalanine (L-DOPA) in unilaterally 6-OHDA-lesioned rats. Moreover, PBF509 (3 mg/kg) inhibited L-DOPA-induced dyskinesia (LID), showing not only its efficacy on reversing parkinsonian motor impairments but also acting as antidyskinetic agent. Overall, here we describe a new orally selective A2AR antagonist with potential utility for PD treatment, and for some of the side effects associated to the current pharmacotherapy (i.e., dyskinesia).

Effects of yak excreta on soil organic carbon mineralization and microbial communities in alpine wetlands of southwest of China

Improving knowledge of how soil organic carbon (SOC) mineralization responds to excreta application is essential to better understand whether wetland carbon (C) pools will react to grazing. We investigated microbial activity and community structure in the different treatments of excreta addition experiments to examine how soil C mineralization responds to the excreta input in terms of microbial activities and compositions in wetland soils. The microcosms of mineralization incubation of excreta addition were established. The structure of the microbial community was described by the fatty acid composition of the phospholipids (PLFA). The methylumbelliferyl-linked substrates (MUB) and l-dihydroxyphenylalanine (L-DOPA) substrates were used to investigate the activities of β-glucosidase (BG), N-acetyl-glucosaminidase (NAG), acid phosphatase (AP), cellobiohydrolase (CBH), and phenol oxidase (PO). Excreta addition altered the cumulative C mineralization in swamp meadow (SM) and peatland (PL) soils, but SM was lower than PL. Excreta addition increased the biomass of individual PLFA and the fungi/bacteria ratio, suggesting that microbes are stimulated by nutrients and that the soil microbial community composition is modified by excreta inputs. The hydrolytic enzyme activities were higher in the PL soils than in the SM soils, but the trend was opposite for PO activity. The changes in pH, fungi, actinomycetes (ACT), AP, and CBH after yak fecal input significantly influenced the soil CO2 efflux. Our findings suggest that yak grazing could influence the rate of C cycling in wetland soils by influencing microbial communities, enzyme activities, and soil pH. This study suggest that the yak excreta addition increased cumulative C mineralization in SM and PL soils, and the effect of dung addition was more significant than urine addition. The effect of yak excreta addition on SOC mineralization was related with the soil pH, microorganism structure, and enzyme activity which modified by the excreta addition. Soil pH, fungi, AP, and CBH were positively correlated with SOC mineralization, but ACT was negatively correlated with SOC mineralization. In addition, the changes in C and N sources with yak excreta addition play an important role in altering microbial enzyme activities. The input of yak feces into wetlands because of grazing could increase SOC mineralization and thereby promote C emission.

A multiparametric analysis of the synergistic impact of anti-Parkinson's drugs on the fibrillation of human serum albumin

Protein aggregation have been associated with several human neurodegenerative diseases, such as Parkinson's and Alzheimer's diseases. There are several small molecules that can reduce aggregation of proteins. The present study aimed to test the hypothesis that the application of more than one inhibitor either simultaneously or consecutively may result in more efficient inhibition of protein aggregation. To this end, the anti-amyloidogenic behaviour of benserazide hydrochloride (BH) and levodopa (LD) individually and in combination (BH + LD) was investigated using various biophysical, microscopic, and computational techniques. BH, LD, and BH + LD treatments showed inhibitory effects on protein aggregation and had the ability to minimise the amyloid-induced cytotoxicity in human neuroblastoma cell line (SH-SY5Y). The two drugs in combination showed synergism (combination index, CI < 1) between them. These drugs also destabilised the preformed fibrils of human serum albumin (HSA). Our studies consistently showed that the BH + LD treatment showed highest efficacy towards inhibition and disaggregation of amyloid fibrils in comparison to treatment with BH and LD individually. Therefore, application of drugs in combination against fibrillogenesis may represent a new route for development of means for prevention or delaying of the aggregation-related diseases.

Effect of the selective 5-HT2A receptor antagonist EMD-281,014 on L-DOPA-induced abnormal involuntary movements in the 6-OHDA-lesioned rat.

L-3,4-Dihydroxyphenylalanine (L-DOPA) is the most effective therapy for motor symptoms of Parkinson's disease (PD); however, with repeated administration, as many as 94% of PD patients develop complications such as L-DOPA-induced dyskinesia. We previously demonstrated that EMD-281,014, a highly selective serotonin 2A (5-HT2A) receptor antagonist, reduces the severity of dyskinesia in the parkinsonian marmoset, without interfering with L-DOPA anti-parkinsonian benefit. Here, we assessed the effects of EMD-281,014 on L-DOPA-induced abnormal involuntary movements (AIMs) in the 6-hydroxydopamine (6-OHDA)-lesioned rat. We first determined the pharmacokinetic profile of EMD-281,014, to administer doses leading to clinically relevant plasma levels in the behavioural experiments. Dyskinetic 6-OHDA-lesioned rats were then administered EMD-281,014 (0.01, 0.03 and 0.1mg/kg) or vehicle in combination with L-DOPA and AIMs severity was evaluated. We also assessed the effect of EMD-281,014 on L-DOPA anti-parkinsonian action with the cylinder test. We found that the addition of EMD-281,014 (0.01, 0.03 and 0.1mg/kg) to L-DOPA did not reduce AIMs severity (P > 0.05), when compared to vehicle. EMD-281,014 did not compromise L-DOPA anti-parkinsonian action. Our results suggest that the highly selective 5-HT2A receptor antagonist EMD-281,014 is well-tolerated by parkinsonian rats, but does not attenuate L-DOPA-induced AIMs. Our results highlight differences between rodent and primate models of PD when it comes to determining the anti-dyskinetic action of 5-HT2A receptor antagonists.

Dopamine Synthesis by Non-Dopaminergic Neurons as an Effective Mechanism of Neuroplasticity

In addition to dopaminergic (DAergic) neurons, which possess all of the enzymes of dopamine synthesis (DA), there are neurons that express only one of the enzymes, tyrosine hydroxylase (TH) or aromatic L-amino acid decarboxylase (AAAD). These so-called monoenzymatic neurons are widely distributed in the brain and, in some areas, are even more numerous than dopaminergic (DAergic) neurons. Using in an vitro experimental approach that we developed it was first demonstrated that monoenzymatic neurons that contain complementary enzymes of DA synthesis, TH and AAAD, co-synthesize DA. L-3,4-dihydroxyphenylalanine (L-DOPA), which is synthesized from L-tyrosine in monoenzymatic TH-containing neurons, is transferred to monoenzymatic AAAD-containing neurons, where L-DOPA is converted to DA. We have also shown that cooperative synthesis of DA, although performed in some parts of the brain in the norm, is predominantly a manifestation of neuroplasticity in pathology. This additional source of DA synthesis contributes to compensation of the DA deficit, which occurs in neurodegenerative diseases such as hyperprolactinemia and Parkinson’s disease, whose pathogenesis is associated with degeneration of dopaminergic (DAergic) neurons. It is also possible that L-DOPA, which is secreted by monoenzymatic TH-containing neurons, plays the role of a neurotransmitter or neuromodulator and acts on target neurons through receptors to L-DOPA, DA, and norepinephrine. Thus, numerous non-dopaminergic monoenzymatic neurons, which are widely distributed in the brain, jointly synthesize DA, which is the most important mechanism of neuroplasticity; this compensates for the DA deficit during the degeneration of DAergic neurons.

One-step fabrication of bicompartmental microparticles as a dual drug delivery system for Parkinson’s disease management

Parkinson’s disease (PD) is a progressive neurodegenerative disorder primarily affecting elderly patients. In order to efficiently manage the symptoms of the disease, controlled and continuous supply of PD drugs [levodopa (LD) and carbidopa (CD)] would be highly beneficial. Currently available commercial tablets are not sufficiently capable of releasing PD drugs in a sustainable fashion. Hence, the purpose of this study was to engineer a carrier system to deliver more than one therapeutic to the target site with independent, sustainable release kinetics. To achieve that, we have designed and developed a set of disk-shaped microparticles composed of two distinct compartments, the first containing a relatively hydrophilic amorphous polymer, poly(lactide-co-glycolide) and the second composed entirely of hydrophobic semicrystalline polylactide. These bicompartmental microparticles with narrow particle size distribution (coefficient of variance ~ 6%) were fabricated by electrohydrodynamic co-jetting technique and characterized by optical microscopy, confocal laser scanning microscopy, SEM, DSC and Raman microscopy. The feasibility of the bicompartmentalized microparticles was confirmed by localizing two different hydrophilic PD drugs (LD and CD) in two different compartments with 4:1 ratio similar to commercially available tablets. This one-step particle fabrication method generated bicompartmental particles with high drug encapsulation efficiency (LD: > 97% and CD: > 68%). The resulting dual drug-loaded disk-shaped microparticles demonstrated simultaneous release of two independent PD drugs with similar release pattern (> 80% release of both drugs within 5 h). These compartmentalized microparticles can potentially act as dual drug delivery system to enable controlled, sustained release of a wide range of water soluble drugs including PD drugs.

Upgraded Methodology for the Development of Early Diagnosis of Parkinson's Disease Based on Searching Blood Markers in Patients and Experimental Models.

Numerous attempts to develop an early diagnosis of Parkinson's disease (PD) by searching biomarkers in biological fluids were unsuccessful. The drawback of this methodology is searching markers in patients at the clinical stage without guarantee that they are also characteristic of either preclinical stage or prodromal stage (preclinical-prodromal stage). We attempted to upgrade this methodology by selecting only markers that are found both in patients and in PD animal models. HPLC and RT-PCR were used to estimate the concentration of amino acids, catecholamines/metabolites in plasma and gene expression in lymphocytes in 36 untreated early-stage PD patients and 52 controls, and in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-treated mice at modeling the clinical ("symptomatic") stage and preclinical-prodromal ("presymptomatic") stage of PD. It was shown that among 13 blood markers found in patients, 7 markers are characteristic of parkinsonian symptomatic mice and 3 markers of both symptomatic and presymptomatic mice. According to our suggestion, the detection of the same marker in patients and symptomatic animals indicates adequate reproduction of pathogenesis along the corresponding metabolic pathway, whereas the detection of the same marker in presymptomatic animals indicates its specificity for preclinical-prodromal stage. This means that the minority of markers found in patients-decreased concentration of L-3,4-dihydroxyphenylalanine (L-DOPA) and dihydroxyphenylacetic acid (DOPAC) and increased dopamine D3 receptor gene expression-are specific for preclinical-prodromal stage and are suitable for early diagnosis of PD. Thus, we upgraded a current methodology for development of early diagnosis of PD by searching blood markers not only in patients but also in parkinsonian animals.

Heart rate variability and its influencing factors in patients with dementia with Lewy bodies and patients with Parkinson disease dementia

Objective To compare the difference in heart rate variability(HRV)between patients with dementia with Lewy bodies(DLB)and those with Parkinson disease dementia(PDD)and to identify the influencing factors. Methods We retrospectively enrolled 30 patients with DLB(DLB group)and 41 patients with PDD(PDD group)at the outpatient and inpatient services of our hospital from January 2010 to December 2017 in this study, and further recruited 119 normal elderly individuals to serve as the control group.HRV was recorded with 24 h dynamic electrocardiogram and compared between the DLB group and the PDD group.Time domain measures including standard deviation of all normal to normal(NN)intervals(SDNN)and square root of the mean squared differences of successive NN intervals(rMSSD)and frequency domain measures including total power(TP), low frequency(LF), and high frequency(HF)were analyzed with a customized program.The levodopa equivalent dose(LED), mini-mental state examination(MMSE), Montreal Cognitive Assessment(MoCA), Hoehn-Yahr stage(H-Y stage)and the unified Parkinson's disease rating scale Ⅲ(UPDRS Ⅲ), and the Alzheimer's disease Cooperative Study-Activities of Daily Living scale(ADCS-ADL)were assessed in DLB and PDD patients to investigate the influencing factors. Results SDNN, TP, and LF in the DLB group and the PDD group were significantly lower than in the control group(F=14.154, 4.742, 4.897, P 0.05). The DLB group and the PDD group did not differ in HF from the control group(P>0.05). There was no significant difference in any HRV indexes between DLB and PDD patients(P>0.05). Correlation analysis showed no correlation of HRV with cognitive level(MMSE, MoCA), motor disturbance degree(UPDRS Ⅲ, H-Yahr stage), daily living ability(ADCS-ADL), or dosage of anti-PD drugs(each P>0.05). Conclusions DLB and PDD patients present similar impairments in autonomic nervous system function, which are not associated with disease severity. Key words: Dementia; Lewy bodies; Parkinson disease; Heart rate variability

Nefazodone reduces dyskinesia, but not psychosis-like behaviours, in the parkinsonian marmoset.

Nefazodone is an anti-depressant that interacts with a wealth of pharmacological targets, including some that may exert anti-dyskinetic and anti-psychotic effects in Parkinson's disease (PD), notably serotonin 1A and 2A receptors. In this study, we sought to determine the effect of nefazodone on L-3,4-dihydroxyphenylalanine (L-DOPA)-induced dyskinesia and psychosis-like behaviours (PLBs) in the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-lesioned primate. Six common marmosets developed parkinsonism following administration of MPTP, after which they were treated chronically with L-DOPA to induce stable dyskinesia and PLBs. In behavioural experiments, nefazodone (0.01, 0.1 and 1mg/kg) or vehicle was administered in combination with L-DOPA and its effects on dyskinesia, PLBs and parkinsonian disability were assessed. The addition of nefazodone 0.01, 0.1 and 1mg/kg to L-DOPA reduced the severity of peak dose dyskinesia by ≈ 21%, ≈ 39% and ≈ 42% (all P < 0.05), while it did not have any significant effect on PLBs, when compared to L-DOPA/vehicle. Parkinsonian disability was not affected by any dose of nefazodone. Our results suggest that nefazodone may be effective to alleviate L-DOPA-induced dyskinesia in PD, while it may not exert any beneficial effect on dopaminergic psychosis.

Survey on sleep disorders in patients with Parkinson disease

Objective To survey the prevalence and distribution of sleep disorders in patients with Parkinson disease (PD) and to analyze the influencing factors. Methods The prevalence and distribution of sleep disorders were surveyed with Parkinson Disease Sleep Scale (PDSS) among 206 PD patients. The association of sleep disorders with age, course of disease, cognitive function, motor function, depression, and the equivalent dose of levodopa (LED) was analyzed. Results The overall PDSS score in 206 patients was (116.9±21.4). The three most frequent items of sleep disorders were the overall sleep quality(181/206, 87.9%), difficulty in maintaining sleep(160/206, 77.7%)and nocturnal enuresis(151/206, 73.3%); the three least frequent items were early awaking(87/206, 42.2%), urinary incontinence(56/206, 27.2%)and hallucination(44/206, 21.4%). The three items with the lowest average scores were nocturnal enuresis(6.9±3.1), difficulty in maintaining of sleep(7.1±2.7)and overall sleep quality(7.1±2.0); three items with the highest average scores were audiovisual illusion(9.3±1.8), incontinence caused by motion disability(9.0±2.1) and early awaking with upper and lower limb pain(8.7±2.1). PD patients were divided into group 1 [Hoehn-Yahr(HY F=3.27, P=0.04) between group 1 and group 3. The linear regression analysis showed that the HAMD and the LED scores had the greatest influence on PDSS score (R2=0.142, 0.196). Conclusion PD patients have a variety of sleep symptoms. The treatment of large doses of dopamine and depression contribute to the occurrence of PD sleep disorders. Key words: Parkinson disease; Sleep disorders; Depression; Questionnaires

Formulation of levodopa containing dry powder for nasal delivery applying the quality-by-design approach

The aim of this work was to carry out preliminary experiments for preparation of levodopa (LEVO)-containing intranasal powder. The experiments were designed according to the Quality by Design (QbD) concept. Based on prior risk assessment, LEVO and chitosan (CH) or sodium hyaluronate (HA) as mucoadhesive matrix formers were co-milled using planetary ball mill to prepare microparticles as drug delivery systems. The rotation speed, the milling time and the drug-additive ratio were evaluated to be the most relevant milling factors - as a result of the initial risk assessment; which were set according to a factorial design. The effects of critical process parameters and excipients were investigated on the particle size and surface characteristics of products, and on the crystallinity, in vitro dissolution and permeability of LEVO. Milling in the presence of higher amount of HA resulted in smaller average particle size of powders (D50 = 13.068 μm) and higher initial dissolution and permeation of LEVO compared to CH-containing formulations (D50 = 21.667 μm).

Electrodeposited Gold on Carbon-Fiber Microelectrodes for Enhancing Amperometric Detection of Dopamine Release from Pheochromocytoma Cells.

Exocytosis is an ultrafast cellular process which facilitates neuron-neuron communication in the brain. Microelectrode electrochemistry has been an essential tool for measuring fast exocytosis events with high temporal resolution and high sensitivity. Due to carbon fiber's irreproducible and inhomogeneous surface conditions, however, it is often desirable to develop simple and reproducible modification schemes to enhance a microelectrode's analytical performance for single-cell analysis. Here we present carbon-fiber microelectrodes (CFEs) modified with a thin film of electrodeposited gold for the detection of exocytosis from rat pheochromocytoma cells (PC12), a model cell line for neurosecretion. These new probes are made by a novel voltage-pulsing deposition procedure and demonstrate improved electron-transfer characteristics for catecholamine oxidation, and their fabrication is tractable for many different probe designs. When we applied the probes to the detection of catecholamine release, we found that they outperformed unmodified CFEs. Further, the improved performance was conserved at cells incubated with L-DOPA (l-3,4-dihydroxyphenylalanine), a precursor to dopamine that increases the quantal size of the release events. Future use of this method may allow nanoelectrodes to be modified for highly sensitive detection of exocytosis from chemical synapses.

Acupuncture Alleviates Levodopa-Induced Dyskinesia via Melanin-Concentrating Hormone in Pitx3-Deficient aphakia and 6-Hydroxydopamine-Lesioned Mice.

Although L-3,4-dihydroxyphenylalanine (L-DOPA) is currently the most effective medication for treating Parkinson's disease (PD) motor symptoms, its prolonged administration causes several adverse effects, including dyskinesia. To identify the mechanisms underlying the effects of acupuncture on L-DOPA-induced dyskinesia (LID), antidyskinetic effects of acupuncture were investigated in two mouse models of PD. Acupuncture stimulation at GB34 alleviated abnormal involuntary movements (AIMs) in Pitx3-deficient aphakia mice (ak/ak) following L-DOPA administration and these effects were reproduced in 6-hydroxydopamine (6-OHDA)-lesioned mice with LID. A transcriptome analysis of the hypothalamus revealed pro-melanin-concentrating hormone (Pmch) gene was highly expressed in acupuncture-treated mouse from ak/ak model of LID as well as 6-OHDA model of LID. Acupuncture combined with the administration of MCH receptor antagonist did not have any beneficial effects on dyskinesia in L-DOPA-injected ak/ak mice, but the intranasal administration of MCH attenuated LID to the same degree as acupuncture in both ak/ak and 6-OHDA mice with LID. A gene expression profile with a hierarchical clustering analysis of the dyskinesia-induced ak/ak mouse brain revealed an association between the mechanisms underlying acupuncture and MCH. Additionally, altered striatal responses to L-DOPA injection were observed after prolonged acupuncture and MCH treatments, which suggests that these treatment modalities influenced the compensatory mechanisms of LID. In summary, present study demonstrated that acupuncture decreased LID via hypothalamic MCH using L-DOPA-administered ak/ak and 6-OHDA mouse models and that MCH administration resulted in novel antidyskinetic effects in these models. Thus, acupuncture and MCH might be valuable therapeutic candidates for PD patients suffering from LID.

Analysis on the clinical, neuroimaging and pathological characteristics in patients with corticobasal syndrome

Objective To investigate the clinical, neuroimaging and pathological features of patients with corticobasal syndrome (CBS). Methods Three CBS patients underwent neuropsychological assessment, MRI scan, 18F-FDG PET scan, and 11C-PiB PET scan, and the test results were analyzed. Results All three patients showed poor response to levodopa treatment and asymmetric dyskinesia including rigidity, tremor, dystonia and abnormalities in posture and gait.One patient showed apraxia and three patients presented with cognitive impairment.The crania MRI demonstrated mild cerebral atrophy which was slightly more severe in the contralateral side of the initially affected limb.The (18)F-FDG PET/CT scan revealed asymmetric decreased metabolism in the frontal, parietal, temporal, and insular lobe, as well as in basal ganglia.The results of (11)C-PiB) PET showed that amyloid beta-peptide (A Beta) deposition in the cortex was positive in one patient, and negative in the other two patients. Conclusions Corticobasal syndrome is characterized by asymmetric dyskinesia and cognitive impairment, and often associated with apraxia, cortical sensory deficits, and alien limb phenomena.The MRI and FDG PET are helpful for CBS diagnosis, and the PiB PET facilitates the pathological diagnosis. Key words: Pathology; Neuroimaging; Corticobasal syndrome

Human tyrosine hydroxylase in Parkinson's disease and in related disorders.

Parkinson's disease (PD) is an aging-related movement disorder mainly caused by a deficiency of neurotransmitter dopamine (DA) in the striatum of the brain and is considered to be due to progressive degeneration of nigro-striatal DA neurons. Most PD is sporadic without family history (sPD), and there are only a few percent of cases of young-onset familial PD (fPD, PARKs) with the chromosomal locations and the genes identified. Tyrosine hydroxylase (TH), tetrahydrobiopterin (BH4)-dependent and iron-containing monooxygenase, catalyzes the conversion of L-tyrosine to L-3,4-dihydroxyphenylalanine (L-DOPA), which is the initial and rate-limiting step in the biosynthesis of catecholamines (DA, noradrenaline, and adrenaline). PD affects specifically TH-containing catecholamine neurons. The most marked neurodegeneration in patients with DA deficiency is observed in the nigro-striatal DA neurons, which contain abundant TH. Accordingly, TH has been speculated to play some important roles in the pathophysiology in PD. However, this decrease in TH is thought to be secondary due to neurodegeneration of DA neurons caused by some as yet unidentified genetic and environmental factors, and thus, TH deficiency may not play a direct role in PD. This manuscript provides an overview of the role of human TH in the pathophysiology of PD, covering the following aspects: (1) structures of the gene and protein of human TH in relation to PD; (2) similarity and dissimilarity between the phenotypes of aging-related sPD and those of young-onset fPD or DOPA-responsive dystonia due to DA deficiency in the striatum with decreased TH activity caused by mutations in either the TH gene or GTP cyclohydrolase I (GCH1) gene; and (3) genetic variants of the TH gene (polymorphisms, rare variants, and mutations) in PD, as discovered recently by advanced genome analysis.