laser-microdissection-based-protocol-for-lc-msms-analysis-proteomic

Laser Microdissection-Based Protocol for the LC-MS/MS Analysis of the Proteomic Profile of Neuromelanin Granules

Neuromelanin is a black-brownish pigment, present in so-called neuromelanin granules (NMGs) in dopaminergic neurons of the substantia nigra pars compacta. ...

University Hospital of Wuerzburg, University of Wuerzburg

This article briefly reviews findings from studies on neuromelanin (NM)-bound ferric iron, which provide unique insights into the physiological functions of NM and possible pathophysiological mechanisms underlying dopaminergic neuronal cell death in Parkinson's disease (PD). NM is considered an endogenous iron-binding molecule of pigmented neurons and is believed to play a physiological role in intraneuronal iron homeostasis. In PD, where nigral iron levels are increased, saturation of high-affinity iron-binding sites on NM may overwhelm the protective capacity of this molecule, leading instead to an increase in redox-active iron, and subsequent cellular damage both in vitro and in vivo. Available data also suggest that the iron released from NM affects the ubiquitin-proteasome system in mitochondria, leading to the failure to clear proteins such as alpha-synuclein and to the development of abnormal alpha-synuclein-immunopositive Lewy bodies that contribute to dopaminergic nerve cell death in PD. NM-bound ferric iron mimics certain characteristic features of the human disease in vitro and in vivo (face validity), in conformity with the theoretical rationale for PD (construct validity) and predicts aspects of PD behaviour and neurobiology (predictive validity) that makes it a valid experimental model with which to study the mechanisms of dopaminergic neurodegeneration in PD.

Neuromelanin-bound ferric iron as an experimental model of dopaminergic neurodegeneration in Parkinson's disease.

Sarizotan, a 5-HT(1A) agonist with additional affinity for D(3) and D(4) receptors, has been demonstrated to have anti-dyskinetic effects. The mechanism by which these effects occur is not clear. Using unilateral 6-hydroxydopamine-lesioned rats that received chronic intraperitoneal (ip) administration of L: -3,4-dihydroxyphenylalanine (L: -DOPA) we investigated the involvement of D(3) and 5-HT(1A) receptors in the effects of sarizotan on contraversive circling and abnormal involuntary movements (AIMs). Before sensitization by chronic L: -DOPA treatment (12.5 with 3.25 mg/kg benserazide ip, twice daily for 21 days), no effect of the selective D(3) agonist, PD128907 (1 or 3 mg/kg ip), or the selective D(3) antagonist, GR103691 (0.5 or 1.5 mg/kg ip), was observed. Treatment with sarizotan (1 or 5 mg/kg ip) dose-dependently inhibited the L: -DOPA-induced contraversive turning and AIMs. In co-treatment with the 5-HT(1A) antagonist, WAY100635 (1 mg/kg ip), sarizotan failed to affect this behaviour, confirming the prominent 5-HT(1A) receptor-mediated mechanism of action. In the presence of PD128907 (3 mg/kg ip), the effects of sarizotan on contraversive turning, locomotive dyskinesia and axial dystonia, but not on orolingual and forelimb dyskinesia, were blocked. On its own, PD128907 had no effect on the behavioural effects of L: -DOPA except that it tended to reduce orolingual and forelimb dyskinesia. GR103691 had no effect on its own or in combination with sarizotan. These data identify an involvement of D(3) receptors in the action of sarizotan on some, but not all L: -DOPA-induced motor side effects. This selective involvement is in contrast to the more general involvement of 5-HT(1A) receptors in the anti-dyskinetic effects of sarizotan.

Role of dopamine D3 and serotonin 5-HT 1A receptors in L: -DOPA-induced dyskinesias and effects of sarizotan in the 6-hydroxydopamine-lesioned rat model of Parkinson's disease.

A central problem in the treatment of Parkinson's disease (PD) is the development of motor disturbances like L: -DOPA-induced dyskinesia (LID) after long-term treatment. Preclinical and clinical studies demonstrated that serotonin 5-HT(1A) receptor agonists attenuate this disabling motor side effect. The aim of this study was to investigate the ability of flibanserin compared to buspirone to attenuate L: -DOPA-sensitized contraversive circling in hemiparkinsonian rats, which is an animal model of LID. Both drugs have a preferential affinity for the serotonin 5-HT(1A) receptors. Buspirone was in comparison because it was expected to have an effect in this model. Unilaterally 6-hydroxydopamine lesioned rats were treated twice daily intraperitoneally (ip) with L: -DOPA methylester (12.5 mg/kg) and benserazide (3.25 mg/kg) for 21 days (on days 1, 3, 5, 8, 11, 14, 17 and 21). On day 24, L: -DOPA-sensitized rats were treated ip 5 min prior to administration of L: -DOPA methyl ester and benserazide with either saline (controls), 2.5, 5 and 10 mg/kg buspirone or flibanserin. Acute administration of both flibanserin and buspirone, dose dependently, attenuated the increased contraversive circling. An almost complete inhibition of the turning response was observed at 5 mg/kg buspirone and 10 mg/kg flibanserin. The current preclinical findings further implicate the 5-HT(1A) receptor as a promising therapeutic target for the reduction of LID and predict a potential efficacy of flibanserin in the treatment of LID in PD.

Flibanserin attenuates L: -DOPA-sensitized contraversive circling in the unilaterally 6-hydroxydopamine-lesioned rat model of Parkinson's disease.

Reliable biomarkers that can be used for early diagnosis and tracking disease progression are the cornerstone of the development of disease-modifying treatments for Parkinson's disease (PD). The German Society of Experimental and Clinical Neurotherapeutics (GESENT) has convened a Working Group to review the current status of proposed biomarkers of neurodegeneration according to the following criteria and to develop a consensus statement on biomarker candidates for evaluation of disease-modifying therapeutics in PD. The criteria proposed are that the biomarker should be linked to fundamental features of PD neuropathology and mechanisms underlying neurodegeneration in PD, should be correlated to disease progression assessed by clinical rating scales, should monitor the actual disease status, should be pre-clinically validated, and confirmed by at least two independent studies conducted by qualified investigators with the results published in peer-reviewed journals. To date, available data have not yet revealed one reliable biomarker to detect early neurodegeneration in PD and to detect and monitor effects of drug candidates on the disease process, but some promising biomarker candidates, such as antibodies against neuromelanin, pathological forms of α-synuclein, DJ-1, and patterns of gene expression, metabolomic and protein profiling exist. Almost all of the biomarker candidates were not investigated in relation to effects of treatment, validated in experimental models of PD and confirmed in independent studies.

Biomarker candidates of neurodegeneration in Parkinson's disease for the evaluation of disease-modifying therapeutics.

Piribedil is a non-ergoline, dopamine D(2)/D(3) receptor agonist with α(2) adrenoceptor antagonist properties that has been used in the treatment of Parkinson's disease (PD). Noradrenergic neurotransmission may be involved in the pathogenesis of dyskinesias induced by chronic treatment with L-DOPA (3,4-dihydroxyphenylalanine, levodopa), but its role in the in vivo action of piribedil or on different subclasses of abnormal involuntary movements (AIMs) remains unclear. The aims of this study were therefore (1) to investigate the anti-dyskinetic effects of piribedil on L-DOPA-induced contralateral turning behaviour, locomotive dyskinesias (LD), axial dystonia (AD), orolingual dyskinesia (OD) and forelimb dyskinesia (FD) and (2) to compare these effects to the α(2) adrenoceptor antagonist, idazoxan, or the α(2) adrenoceptor agonist, clonidine. Rats were unilaterally lesioned with 6-hydroxydopamine (6-OHDA) and injected intraperitoneally twice daily with L-DOPA methylester (12.5 mg/kg) and benserazide (3.25 mg/kg). After 3 weeks, the effects of piribedil (5, 15, 40 mg/kg), clonidine (0.15 mg/kg), idazoxan (10 mg/kg) and combinations of these drugs were scored during 2 h. Pre-treatment with 5 and 40 mg/kg, but not 15 mg/kg, of piribedil reduced turning behaviour and AD, OD and FD, but piribedil increased LD at the 40 mg/kg doses compared to the L-DOPA group. Idazoxan induced similar effects as piribedil (40 mg/kg), except that it had no effect on LD. Idazoxan blocked the effect of piribedil on AD and FD. Clonidine reduced all AIMs except OD, possibly because of its sedative effect. Clonidine blocked the effect of piribedil on AD, OD and FD. These data suggest a differential involvement of α(2) adrenergic receptors in the action of piribedil on different subclasses of L-DOPA-induced dyskinesias.

The effect of piribedil on L-DOPA-induced dyskinesias in a rat model of Parkinson's disease: differential role of α(2) adrenergic mechanisms.

A major concern regarding psychostimulant medication (amphetamine and methylphenidate) in the treatment of children and adolescents with attention deficit/hyperactivity disorder (ADHD) are the potential adverse effects to the developing brain, particularly in regard to dopaminergic brain function. The present review focuses on the pharmacology of these psychostimulants, their mode of action in the human brain and their potential neurotoxic effects to the developing brain in animals, particularly concerning DA brain function. The potential clinical significance of these findings for the treatment of ADHD in children and adolescents is discussed. Studies on sensitization to psychostimulants' rewarding effects, which is a process expected to increase the risk of substance abuse in humans, are not included. The available findings in non-human primates support the notion that the administration of amphetamine and methylphenidate with procedures simulating clinical treatment conditions does not lead to long-term adverse effects in regard to development, neurobiology or behaviour as related to the central dopaminergic system.

Is the treatment with psychostimulants in children and adolescents with attention deficit hyperactivity disorder harmful for the dopaminergic system?

Annual report 2015.

What are the benefits of methylphenidate as a treatment for children and adolescents with attention-deficit/hyperactivity disorder?

Editors must be vigilant to guarantee the quality and credibility of published scientific work.

Special edition on the occasion of Jan K. Buitelaar's 65th anniversary.

Misconceptions about ADHD stigmatize affected people, reduce credibility of providers, and prevent/delay treatment. To challenge misconceptions, we curated findings with strong evidence base.

Article	Year
Neuromelanin-bound ferric iron as an experimental model of dopaminergic neurodegeneration in Parkinson's disease. Parkinsonism & related disorders\| PubMed ID: 18585086	2008
Role of dopamine D3 and serotonin 5-HT 1A receptors in L: -DOPA-induced dyskinesias and effects of sarizotan in the 6-hydroxydopamine-lesioned rat model of Parkinson's disease. Journal of neural transmission (Vienna, Austria : 1996)\| PubMed ID: 21253782	2011
Flibanserin attenuates L: -DOPA-sensitized contraversive circling in the unilaterally 6-hydroxydopamine-lesioned rat model of Parkinson's disease. Journal of neural transmission (Vienna, Austria : 1996)\| PubMed ID: 21274579	2011
Biomarker candidates of neurodegeneration in Parkinson's disease for the evaluation of disease-modifying therapeutics. Journal of neural transmission (Vienna, Austria : 1996)\| PubMed ID: 21755462	2012
The effect of piribedil on L-DOPA-induced dyskinesias in a rat model of Parkinson's disease: differential role of α(2) adrenergic mechanisms. Journal of neural transmission (Vienna, Austria : 1996)\| PubMed ID: 22592937	2013
Is the treatment with psychostimulants in children and adolescents with attention deficit hyperactivity disorder harmful for the dopaminergic system? Attention deficit and hyperactivity disorders\| PubMed ID: 23605387	2013
Annual report 2015. Attention deficit and hyperactivity disorders\| PubMed ID: 26895449	2016
What are the benefits of methylphenidate as a treatment for children and adolescents with attention-deficit/hyperactivity disorder? Attention deficit and hyperactivity disorders\| PubMed ID: 28168407	2017
Editors must be vigilant to guarantee the quality and credibility of published scientific work. Attention deficit and hyperactivity disorders\| PubMed ID: 30328576	2018
Special edition on the occasion of Jan K. Buitelaar's 65th anniversary. Attention deficit and hyperactivity disorders\| PubMed ID: 30758776	2019
The World Federation of ADHD International Consensus Statement: 208 Evidence-based conclusions about the disorder. Neuroscience and biobehavioral reviews\| PubMed ID: 33549739	2021

Manfred Gerlach

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