Prof. Manfred Göthert
Institute of Pharmacology and Toxicology
University of Bonn, Bonn (Germany)


Cannabinoid research has been stimulated by the identification and the molecular and functional characterization of two classes of cannabinoid (CB) receptors, termed CB1 and CB2, as well as by the discovery of endogenous CB receptor ligands such as anandamide. The CB1 receptor occurs in nervous tissue whereas the CB2 receptor is expressed by immune cells.
Evidence has accumulated in recent years that the effects of cannabinoids in the nervous system are mediated not only by CB1 receptors but that, in addition, ligand- and voltage-gated ion channels are targets of endogenous and exogenous cannabinoid receptor agonists. In view of a common functional role of cannabinoid receptor agonists and 5-HT3 receptor antagonists in the control of emesis, putative actions of cannabinoids at this serotonin receptor became particularly interesting. The 5-HT3 receptor is exceptional among the serotonin receptor types in that it is a ligand-gated ion channel. Such channels are generally known to be endowed with allosteric modulatory sites. To check the possibility of an allosteric modulation of 5-HT3 receptors by cannabinoids, excised outside-out patches from HEK293 cells stably transfected with the human (h) 5-HT3A receptor cDNA were used to determine the effects of cannabinoid receptor ligands on the 5-HT-induced current using the patch clamp technique. The 5-HT-induced current was inhibited by cannabinoid receptor agonists such as anandamide, delta9-tetrahydrocannabinol (THC) and WIN55,212-2. The potency of anandamide in inhibiting 5-HT-induced current was even higher than its affinity for CB1 and CB2 receptors in radioligand binding studies. WIN55,212-2 caused a noncompetitive inhibition of the 5-HT-induced current. The inhibition was stereoselective since WIN55,212-3, an enantiomer of WIN55,212-2, did not affect the 5-HT-induced current. The CB1 receptor antagonist rimonabant (SR141716A) failed to counteract the inhibitory effect of WIN55,212-2, thus excluding an involvement of CB1 receptors. The latter possibility was also ruled out by radioligand binding studies on membranes of the HEK293 cells.
Taken together, these data indicate that cannabinoids stereoselectively inhibit currents through recombinant h5-HT3A receptors independently of cannabinoid receptors. Probably the cannabinoids act allosterically at a modulatory site of the h5-HT3A receptor. When considering the high potency of the endogenous agonist anandamide at this site, the latter may be tonically activated by anandamide and, hence, anandamide may play a physiological role by mediating a regulatory effect on the functional state of the 5-HT3 receptor. In line with the data mentioned so far, WIN55,212-2 inhibited 5-HT-induced current through native 5-HT3 receptor channels of N1E-115 cells composed of 5-HT3A and 5-HT3B subunits. A cannabinoid receptor-independent inhibition by anandamide was also found in Xenopus oocytes transfected with the cDNA of the mouse 5-HT3 receptor.
In addition, evidence has been presented that cannabinoid-induced allosteric modulation of 5-HT3 receptor function is operative in mammalian tissue (rat nodose ganglion) and in whole animals in vivo, in particular in anaesthetized rats pretreated with the CB1 receptor antagonist rimonabant. In such animals, activation of the 5-HT3 receptors on cardiac afferent vagal nerves by bolus injection of phenylbiguanide induced the Bezold-Jarisch reflex, i.e. a decrease in heart rate. This effect was inhibited by WIN55,212-2 but not by WIN55,212-3.
As already mentioned, THC shares the antiemetic effect of 5-HT3 receptor antagonists. Furthermore, it should be noted in the context of the analgesic effect of THC that 5-HT3 receptors occur at high density in regions which are involved in the control of pain. Accordingly it is conceivable that the allosteric site via which cannabinoids modulate 5-HT3 receptor function may become a target of new analgesic and antiemetic drugs.

Impaired DNA repair systems and neurodegenerative diseases

Prof. Maurizio Memo
Department of Biomedical Sciences and Biotechnologies
University of Brescia, Medical School, Brescia (Italy)

Preservation of genomic stability is an essential biological function. Cells engage very efficiently mechanisms involving DNA surveillance/repair proteins that work to maintaining inherited nucleotide sequence of genomic DNA over time. After DNA damage, that can arise during duplication or after genotoxic stimuli, cells activate intracellular pathways which are able to recognize the damage, to arrest cell cycle, to recruit DNA repair factors, to repair the damage or induce apoptosis. This definitely relevant process is finalized to prevent the generation and the persistence of impaired cells which may ultimately be detrimental to the organism. Very little is known about the role of DNA damage sensors and repair factors in terminally differentiated, not proliferating cells, like neurons.
It is well recognized that mutation of genes related with DNA damage repair are associated with specific cancer-prone syndromes. Interestingly, many human pathological conditions with genetic defects in DNA damage responses are also characterized by neurological deficits. These neurological deficits can manifest themselves during many stages of development, suggesting an important role for DNA repair during the development and maintenance of the brain.
Here I summarize recent data underlying the contribution to neurodegeneration of at least two transcription factors known to be involved in DNA damage sensing and repairing: the tumour suppressor gene p53 and the component of the DNA repair system MSH2. Both proteins participate in the cancer prevention machinery for the body as well as in the neurodegenerative process. Moreover, they interact with each others to orchestrate DNA repair functions.
Historically, one of the most well known connections between abnormalities of the DNA damage response and neurodegeneration has been the human syndrome of ataxia telangiectasia. However, other syndromes associated with defective DNA damage response also include neurological symptoms as a primary feature of their phenotypes. This argues that defects in the repair of, or response to, DNA damage impact significantly on brain function. We speculate that the list of neuropathologies associated with an impairment of the DNA damage repair system could be even larger including chronic and progressive neurodegenerative diseases like Alzheimer’s disease (AD). In this regard, quantitative and histopathological markers of oxidative DNA damage have been found the brain of AD patients suggesting that the neuronal machinery devoted to restore damaged DNA might be impaired in AD brain. Future studies focussing on these and other aspects of -amyloid neurotoxicity will be important for the molecular description of specific DNA processing and repair mechanisms in brain as well as to define whether cancer and neurodegenerative disease share common genetic risk factors for the development and progression of the disease.

 

 

Has the cholinergic deficit still a key role in alzheimer’s disease?

Prof. Giancarlo Pepeu
Department of Pharmacology
University of Florence, Florence (Italy)

Since the seminal finding that a selective cholinergic neuron loss occurs in Alzheimer’s Disease (AD), the cholinergic hypofunction has been considered the main pathogenetic mechanism of the memory impairment which characterizes the disease. The correction of the cholinergic hypofunction has been, and still is, the main aim of AD therapy, and the cholinesterase inhibitors (ChEI) tacrine, donepezil, rivastigmine, and galantamine are approved for the treatment of AD worldwide. However, ChEIs produce small improvements in cognition and activities of daily living in patients with mild to moderate AD. A recent study (AD2000 Collaborative Group) concluded that “donepezil is not cost effective, with benefits below minimally relevant thresholds” raising the question whether the public health services should pay for this therapy. We can ask why the cholinergic hypofunction correction results in such small improvements of the cognitive functions which are believed to be subserved by the cholinergic system.
It has been shown that choline acetyltransferase (ChAT) and acetylcholinesterase (AChE) in brains of patients with different AD severity, were significantly lower, in comparison with controls, only in subjects with severe dementia, and even a ChAT elevation in the frontal cortex and hippocampus of patients affected by Mild Cognitive Impairment was detected According to these findings, the cholinergic hypofunction in early stages of AD is doubtful or minimal.
We may also ask whether ChEI standard doses bring about a sufficient activation of the brain cholinergic system. In animal experiments, ChEI administration results in ChE inhibition and an increase in extracellular acetylcholine (ACh). We may expect similar effect in man. By measuring ChE inhibition in living human brain by PET, a 20% inhibition, accompanied by changes in executive and attentional but not primary memory functions, after 12 weeks of donepezil therapy was demonstrated. Although a direct relationship between ChE inhibition and ACh levels was never demonstrated, from the animal data it may be assumed that a 20% inhibition may lead to a modest increase in ACh level. However, there is evidence that if high ACh levels are indeed needed for attention and encoding, low ACh levels set dynamics for consolidation. Therefore, it should be clarified whether in AD patients the persistent increase in ACh levels induced by ChEI may impair new information consolidation.
The role of the cholinergic system in cognitive processes has been studied in man mostly by measuring the changes induced by blocking muscarinic ACh receptors with scopolamine. Scopolamine disrupts anterograde memory but spars remote memory and may impair attention. To which extent these deficits are comparable to those observed in AD patients is still matter of debate.
In conclusion, a cholinergic hypofunction can be clearly demonstrated only in severe AD. In these patients the cholinergic hypofunction is compounded with deficits in other neurotransmitter systems involved in learning and memory. Moreover the cholinergic system subserves only some cognitive functions. Therefore, on theoretical ground, only modest cognitive improvements are to be expected in most AD patients. Whether ChEIs may delay AD progression by acting on other pathogenetic mechanisms is still an open question.

 

 

Pharmacogenomics of AD

Prof. Sebastiano Cavallaro
Institute of Neurological Sciences
Italian National Research Council, Catania (Italy)

Alzheimer’s disease is characterized by the extracellular deposits of b-amyloid (Ab), a 42 amino acid peptide cleaved from the large transmembrane protein amyloid precursor protein (APP).
Familial-linked Alzheimer’s disease has been linked to apolipoprotein E4 polymorphism and mutations in APP, presenilin-1, and presenilin-2, all of which are associated with increases in total Ab or selective increases in Ab 1–42. Both in vitro and in vivo studies have reported the toxic effects of Ab or Ab peptide fragments. These studies suggest an important role for Ab in the pathogenesis of Alzheimer’s disease. Exposure of neurons to Ab in particular, induces a form of programmed cell death known as apoptosis. To elucidate the multigenic program underlying this type of cell death, we have used genome-scale screening by oligonucleotide microarrays. The genes implicated offer a genomic view of the changes that accompany neuronal apoptosis and yield new insights into the molecular basis underlying it. Knowledge of the mechanisms and pathways that determine apoptosis and are aberrant in neurodegenerative diseases will pave the way to new and effective therapeutic approaches.

 

 

Apoptosis and neurogenesis in Alzheimer’s disease

Prof. Agata Copani
Department of Pharmaceutical Sciences
University of Catania, Catania (Italy)

Alzheimer’s disease (AD) is the most common neurodegenerative disorder affecting the elderly. Brain regions involved in learning and memory are reduced in size as the result of synaptic loss and apoptotic death of neurons. Postmortem analysis of the brain must show a sufficient number of plaques and tangle to allow the definitive diagnosis of AD. Plaques are extracellular deposits of ß-amyloid protein (Aß), whereas tangles are intracellular aggregates of hyperphosphorylated tau protein.
The puzzle has always been that the pathway from plaques and tangles to neuronal death is not clear. It is now becoming clear that the path proceeds through the reactivation of the cell cycle in neurons. The role played by Aß both as a trigger for cell-cycle related neuronal death and differentiating cue for neuroprogenitor cells will be discussed.

 

 

New therapeutic options in AD

Prof. Stefano Govoni
Department of Experimental and Applied Pharmacology
University of Pavia, Pavia (Italy)

Thirty years ago, neurochemical studies showed a cholinergic deficit as cause of AD, providing the rationale for the pharmacological interventions that have been developed and introduced as therapy in mid 90’s. Before the development of the last generation of cholinesterase inhibitors, the clinical interest on cholinergic drugs has been tempered by the low clinical efficacy or excessive side effects of many of the developed compounds. Over the past 15 years, extensive research has been focused and made progresses on the study of the early processes that lead to the disease; in fact, the aetiology and pathophysiological mechanisms that lead to the death of the neurons in AD have been identified by the characterization of neuritic plaques and neurofibrillary tangles (typical lesions of AD). In particular, it has been demonstrated that the major proteinaceous component of senile plaques is the b-amyloid (Ab) peptide, derived from a larger membrane spanning b-amyloid precursor protein (bAPP). Several studies have indicated Ab formation as an early event in AD and have favoured the hypothesis that, during the development of the disease, an alterated APP metabolism increases the production of Ab, resulting in a neurotoxic effect. Interestingly enough the collected information lead to the notion that APP metabolism may be amenable to pharmacological modulation and may be controlled by classical neurotransmitters, and by drugs inhibiting acetylcholinesterase activity. During the 90’s, improved basic knowledge and better techniques of research allowed the development of new AChE inhibitors, which have been approved for the specific use in AD and have renewed the attention on the connections between ACh and AD. Nowadays some molecules (tacrine, donepezil, rivastigmine and galantamine) have accumulated a significative amount of clinical data about their efficacy, stimulating new experimental and clinical research on the role of cholinergic system in AD, including, as already mentioned, their effect on APP metabolism or Ab deposition. Particularly intriguing are the findings suggesting that Ab can be directly neurotoxic and can increase neuronal vulnerability to a variety of insults including excitotoxicity, glucose deprivation and free radicals. The neurotoxic activity of Ab is still under investigation and it is believed to be correlated with its ability to form aggregates. These notions have promoted the development of new compounds aimed to inhibit Ab formation and aggregation, including the use of immunization techniques against Ab. Within this contest some experience has been accumulated regarding the “vaccine” against beta-amyloid with mixed results that do not allow, at this time, to draw firm conclusions on the efficacy of the treatment, not taking into considerations that serious side effects prevented the full completion of the ongoing clinical trials. These results also did not provide the proof of concept evidence that cleaning up beta-amyloid deposits halts disease progression, an information that up today relies only on animal data. As far as the involvement of classic neurotransmitters in AD pathology glutamate has recently gained attention as possible contributor to the pathology, chronically acting as excitatory neurotoxin in parallel or in cascade with beta amyloid. The approval of memantine (MEM), a reversible voltage dependent antagonist of glutamate NMDA receptors, for AD treatment and its clinical effects in severe AD have further supported this field of investigation. Moreover several preclinical data support an interaction between beta amyloid and glutamate.

In consideration of the different mechanism of action MEM and Acetylcholinesterase Inhibitors (AChEI) have been combined in the hope to obtain at least an additive action. The first trials demonstrated that the co administration of MEM and AChEI has no pharmacokinetic interactions and does not give origin to an excess of side effects while resulting in significantly better outcomes than placebo on measures of cognition, activities of daily living, global outcome, and behavior. However, some fundamental questions whether the combined effect is symptomatic or acts on the progression of the neurodegenerative process and how long the benefits can last have yet to be answered as well as the possibility to extend this therapy to other dementias. During the time these developments had place several other pharmacotherapeutic options have been tried including antinflammatory agents, antioxidants, estrogens, statins and others without clear-cut evidence, at this time, but data are in progress, for a sufficient activity to justify their prescription to AD patients. New therapies are not just behind the corner, but the general picture holds the promise for future better, perhaps combined, therapies for AD.

 

 

Neurotrasmitter alterations and new therapeutic options

Prof. Ezio Giacobini
Department of Rehabilitation and Geriatrics
University of Geneva, Medical School, Geneva (Switzerland)

Various forms of pharmacological treatment are being tested clinically in an effort to slow down or block the conversion of MCI (Mild Cognitive Impairment) to AD (Alzheimer Disease). The results of three recent major studies have not demonstrated a significant effect in delaying diagnosis to AD. Experimental findings on animal models and clinical data with long-term treatment suggest that ChEI (cholinesterase inhibitors) in addition to symptomatic benefit might have a delaying effect on AD progress (Giacobini, 2000). This hypothesis is being tested clinically with new compounds. The indication for ChEI have been extended beyond AD to include other types of dementia.

Preventive approaches being investigated include anti-inflammatories (rofecoxib, 1200 pats,3yrs), anti-oxidants (vit .E) + ChEI (donepezil) , nootropics (piracetam, 675 pats., 1yr), AMPA receptor agonists (ampakines) and cholesterol–lowering drugs (statins). These studies have shown that presently, we don’t have access to a pharmacological treatment able to prevent AD. A more realistic preventive approach seems to try to reduce the risk of vascular dementia and indirectly also of AD.

Data from the first vaccination study (Nitsch et al. 2004, Hock et al. 2004) with pre-aggregated A-beta-42 suggest that patients who generated amyloid plaque immunoreactivity over one year period showed significantly slower rate of decline of cognitive functions and improvement in activities of daily living. These preliminary results support the view that targeting A-beta with immunization could be of benefit to early cases of AD. This approach needs more clinical trials with modified immunotherapies to avoid severe adverse effects.

 

 

The taupathies

Prof. Maria Grazia Spillantini
Brain Repair Centre and Department of Clinical Neurosciences,
University of Cambridge, Cambridge (UK)

Several neurodegenerative diseases of the brain are characterized by the presence of intracellular protein inclusions. In diseases such as Alzheimer’s disease, Pick’s disease, corticobasal degeneration and progressive supranuclear palsy the protein that aggregates intracellularly is the microtubule associated protein tau and the latter three disorders constitute the sporadic tauopathies.

Hereditary tauopathies are familial syndromes clinically characterized by frontotemporal dementia and often associated with parkinsonism, oculomotor disturbances or motoneuron signs, they are grouped under the name of frontotemporal dementia and parkinsonism linked to chromosome 17 (FTDP-17). Neuropathologically, tau deposits are found either in neurones or in both neurones and glia and are present in multiple areas of the CNS with a prevalence in the neocortex, hypothalamus, midbrain, pons and spinal cord. The cellular lesions may be indistinguishable from those of Alzheimer’s disease and sporadic tauopathies. FTDP-17 is caused by exonic or intronic mutations in Tau, the gene coding for the microtubule-associated protein tau.

Adult human tau exists as six isoforms generated by alternative splicing. The repeat domain in the carboxy-terminal half, encoded by exons 9 to 12, constitutes the microtubule binding site of the protein. Alternative splicing of exon 10, which encodes a 31-amino-acid repeat, gives rise to isoforms with three or four binding domains.

Mutations have been found in exons1, 9, 10, 11, 12, 13 and in the intron following exon 10, in the splice-donor site. Mutations in exon 10 or at the exon 10-intron boundary lead to a pathology in neurones and glia that consists of twisted ribbons predominantly composed of four-repeat isoforms and similar to that found in progressive supranuclear palsy and corticobasal degeneration. Mutations in exons 9, 12 and 13 lead to a predominantly neuronal pathology without a significant glial component. Some mutations in exon 12 and 13 lead to the formation of paired helical and straight filaments having a morphology indistinguishable from that of the filaments seen in Alzheimer's neurofibrillary tangles and containing all six isoforms. By contrast, a mutation in exon 9 and another in exon 13 causes intraneuronal inclusions, filament morphology and a pattern of tau bands resembling Pick’s disease.

The variability of the clinicopathologic phenotypes, may be the result of differing molecular mechanisms. Splice site mutations and some exon 10 mutations alter the splicing mechanism, resulting in the production of more four-repeat tau isoforms. Other mutations in exon 10 only affect the four-repeat isoforms, while mutations in exons1, 9, 11, 12 and 13 affect all isoforms; all these mutations disrupt tau-microtubule interactions and some facilitate tau protein aggregation. The study of tau pathology in families with tau mutations is starting to shed light on the function of tau aggregates in neurodegeneration and has given the possibility to produce transgenic mice where to investigate the development of tau pathology. These animals constitute an animal model where drugs can be tested to prevent tau aggregation.

 

 

Not only amnesia

Prof. Eugenio Aguglia
Department of Psychiatry,School of Medicine
University of Trieste, Trieste (Italy)

Mild Cognitive impairment, an intermediate state between normal aging and dementia, is characterized by acquoired cognitive deficits, without significant decline in functional activities of daily living. Studies conducted on MCI have introduced new concept regarding the possible distinctions beetween normal and pathological aging of the brain. Neuroimaging and genetic testing have aided in the identification of individuals at increased risk of dementia. Recently published results of the Current Concepts in MCI conference suggested subclassifications for MCI: MCI-amnestic, MCI-multiple domains slightly impaired, MCI- single non memory domain; based on recognized heterogeneity in the use of the term.

Greater consensus is needed to standardize definitions and research metodology for MCI, so as to make future studies more comparable and more useful for designing effective treatment strategies.

 

 

Pharmacological interactions in the treatment of AD

Prof. Riccardo Torta
Psycho-Oncology Department of Neurosciences and Oncology
University of Turin, Turin (Italy)  

Patients with Alzheimer’ disease often present numerous comorbidities and receive treatment with multiple medications. Drug interactions are possible, both concerning pharmacokinetic and pharmacodynamic aspects. Although pharmacokinetic interactions have been extensively discussed, pharmacodynamic interactions may play a greater role in the significance of drug interactions.

This presentation deals with the more frequent combination, and their pharmacological and clinical interactions, among antidepressants, antipsychotics and cholinesterase inhibitors in patients with Alzheimer’s disease.

 

 

Off label use of antipsychotics

Prof. Stuart A. Montgomery
Imperial College of Science, Technology and Medicine
University of London, London (UK)

Typical and atypical antipsychotics have traditionally been used outside of the range of the simple indication of schizophrenia. The licences for haloperidol, for example, included psychosis and aggression, which was interpreted loosely to include what is now called schizophrenia, mania, autism, conduct disorder and the treatment of agitation in a large number of conditions such as depression, suicide risk and dementia.

The CPMP substantially narrowed the range of treatment indications for atypical antipsychotics compared with their traditional use by abandoning the use of the indication antipsychotic and restricting licensing to only the indications investigated in placebo controlled studies. This has had the unfortunate effect of leaving the older antipsychotics, which are generally more toxic than newer drugs, with a wider range of indications, often without adequate data, than the safer atypical antipsychotics.

Risperidone was licensed by individual countries in the EU whereas olanzapine was licensed centrally and it is clear that the central procedure is more restrictive and the national procedure more variable and erratic. Atypicals are now licensed nationally for schizophrenia, schizoaffective psychosis, negative schizophrenia, affective or depressive symptoms of schizophrenia, and for suicide prevention in schizophrenia; those licensed centrally are only for the indications schizophrenia and affective symptoms of schizophrenia. Quetiapine, licensed nationally throughout the EU, remains unlicensed in France emphasising the inconsistency.

The atypical amisulpiride is licensed for the treatment of dysthymia or depression in four countries only and the combination of olanzapine and fluoxetine is licensed centrally for the treatment of resistant depression and olanzapine for bipolar depression. Risperidone has good placebo controlled evidence of efficacy in the treatment of conduct disorder and autism and is licensed in some countries. However in those countries where the licences were not granted haloperidol remains in use despite the poor quality of the data.

All of the atypicals now have evidence for the treatment of mania with or without concomitant depression but only olanzapine has a licence for long term treatment.

There is good placebo controlled evidence for the efficacy of risperidone, olanzapine, aripiprazole and quetiapine for the treatment of the behavioural and psychological symptoms of dementia (BPSD) and the licence was granted to risperidone and olanzapine. However the FDA and the EMEA have issued a warning that all atypical antipsychotics have an increased risk of mortality due to heart-related events or infection in dementia. No such warning applies to typical antipsychotics. The risk benefit of treatment in this group of difficult to treat terminal dements has not been fully assessed (Brit Med J 2005).

In general the licensing decisions in relation to atypical antipsychotics in Europe appear erratic, variable and contradictory, with little attempt to use expert groups to help to improve the consistency in the application of licensing decisions in line with the evidence. The controls on atypicals are far more stringent than on typicals without apparently taking account of relative risk and this leads to a dependence on the older more dangerous drugs.

Sudeep et al (2005) Brit Med J 330 445-448

 

 

Dopamine and central circuitry

Prof. Micaela Morelli
Department of Toxicology and Centre of Excellence for Neurobiology of Dependence
University of Cagliari, Cagliari (Italy)

The organization, cellular and molecular features, as well as the functional correlates of the circuits which utilize dopamine (DA) as neurotransmitter represent one of the most fertile fields of investigation in neuroscience. Interest in these circuits and their regulation has been and is stimulated especially by their involvement in neurological and psychiatric diseases, besides their role in motor and cognitive functions, and in the motivational aspects of behavior.
The dopaminergic neurons of the midbrain are distributed in a continuum across a number of anatomical structures . On the basis of their cytoarchitectonic features, the main dopaminergic cell groups are located in the substantia nigra (SN), in the ventral tegmental area medial to the SN, and in the retrorubral area caudal and dorsal to the SN. SN compacta dopaminergic neurons (SNc) extends in the SN reticulata (SNr) and in the SN pars lateralis (SNl).
The striatum, which is the principal target of dopaminergic SNc neurons, receives two additional channels of information: one from the cortex the other from the thalamic projection, derived mainly from the intralaminar nuclei of the thalamus. In the striatum, neural information is processed and conveyed into a series of loops, one of these loops is represented by the strionigral pathway reciprocated by the nigrostriatal pathway. The other main striatal output is towards the globus pallidus, which in turn projects to the SNr.
The physiological actions of DA are mediated by at least five different G protein-coupled receptor subtypes which are classified into D1-like family (D1 and D5) and D2-like family (D2, D3 and D4). The functions of D1 and D2 receptors and partially D3 have been characterized through behavioral and biochemical studies, whereas the lack of highly selective compounds for D4 and D5 have hampered the clarification of their functions.
DA plays a prominent role in basal ganglia functioning and several drugs acting at DA receptor level are used in the therapy of schizophrenia and Parkinson’s disease (PD).
DA functions are modulated by several neurotransmitters, among them adenosine is attracting much attention since adenosine A2A receptors are coupled to dopamine D2 receptors on striatal neurons where they modulate DA functions.
The current therapy of PD is based on the use of the dopamine precursor L-DOPA or direct dopamine receptor agonists. These drug treatments, however, after few years become less effective and produce several side effects, the most disabilitating being dyskinesia. Recent evidences obtained in animal models of PD and preliminary clinical trials, indicate that adenosine A2A receptor antagonists might represent a new valuable therapeutic tool for the treatment of PD. Therefore, modulation of DA functions, besides direct actions on the DA system may represent a new approach to the treatment of diseases directly correlated to a DA disfunction.

 

 

Environmental Factors in Parkinson’s Disease

Prof. Roberto Maggio
Department of Neurosciences
University of Pisa, Pisa (Italy)

Thanks to the discovery of MPTP (1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine), it is now clear that, by targeting the nigrostriatal system, neurotoxicants can reproduce the neurochemical and pathological features of idiopathic parkinsonism. The sequence of toxic events triggered by MPTP has also provided us with intriguing clues concerning mechanisms of toxicant selectivity and nigrostriatal vulnerability. Relevant examples are (i) the role of the plasma membrane dopamine transporter in facilitating the access of potentially toxic species into dopaminergic neurons; (ii) the vulnerability of the nigrostriatal system to failure of mitochondrial energy metabolism; and (iii) the contribution of inflammatory processes to tissue lesioning. Epidemiological and experimental data suggest the potential involvement of specific agents as neurotoxicants (e.g. pesticides) or neuroprotective compounds (e.g. tobacco products) in the pathogenesis of nigrostriatal degeneration, further supporting a relationship between the environment and Parkinson's disease. A likely scenario that emerges from our current knowledge is that neurodegeneration results from multiple events and interactive mechanisms. These may include (i) the synergistic action of endogenous and exogenous toxins (e.g. the ability of the pesticide diethyldithiocarbamate to promote the toxicity of other compounds); (ii) the interactions of toxic agents with endogenous elements (e.g. the protein alpha-synuclein); (iii) the tissue response to an initial toxic insult; and, last but not least, (iv) the effects of environmental factors on the background of genetic predisposition and aging.

 

 

Extatory synaptic organization in neurodegenrative disease

Prof. Monica Di Luca
Centre of Excellence on Neurodegenerative Diseases and Department of Pharmacological Sciences,
University of Milan, Milan (Italy)

Molecular pathogenesis of neurodegenerative diseases such as Alzheimer Disease (AD) and Parkinson disease (PD) is the consequence of a complex interplay of several crossing pathways. In the case of AD, accumulating evidences support the amyloid cascade as a pivotal player in AD pathogenesis. Nevertheless, discussion is still ongoing on whether plaque formation can be the telltale of events occurring earlier in the course of the disease, including synaptic changes. The glutamatergic circuitry has been shown implicated among the others in the earlier phases of AD, and it is reflected as a loss of synaptic plasticity. In this study, we evaluate the interaction between ADAM10, a candidate for alpha-secretase, and synapse-associated protein 97 (SAP97). Confocal microscopy shows that SAP97 and ADAM10 display a high co-localisation pattern in transfected COS-7 cells as well as in hippocampal neurons. Moreover, both SAP97 and ADAM10 are enriched at the postsynaptic density. Our results show that SAP97 directly interacts with ADAM10, as demonstrated in co-immuno precipitation experiments from postsynaptic densities, purified from mouse brain tissue. Furthermore, pull down assays reveal that SAP97 interacts with ADAM10 through its SH3 domain, which recognizes proline rich motifs in ADAM10 cytoplasmic tail. These results demonstrate an interaction between ADAM10 and SAP97, which may have a functional implication for the regulation of alpha-secretase trafficking/activity. In PD, the NMDA receptor complex represents a molecular key element in the pathogenesis of long-term synaptic changes in PD. Here we show that NMDA-NR1 subunit and PSD-95 protein levels are selectively reduced in the post-synaptic density (PSD) of dopamine (DA) denervated striata. These effects are accompanied by an increase in striatal levels of alpha Ca2+/Calmodulin-dependent protein kinase II (alphaCaMKII) autophosphorylation along with a higher recruitment of activated alphaCaMKII to the regulatory NMDA-receptor NR2A/B subunits. While normalizing alphaCaMKII autophosphorylation levels as well as assembly and anchoring of the kinase to the NMDA receptor complex, the intrastriatal administration of the CaMKII inhibitors KN-93 and Ant-AIP-II is able to reverse both the alterations in corticostriatal synaptic plasticity and the deficits in spontaneous motor behaviour that are found in an animal model of PD. The same beneficial effects are produced by a regimen of L-DOPA treatment that is able to normalize alphaCaMKII autophosphorylation. These data indicate that an abnormal alphaCaMKII autophosphorylation plays a causal role in the alterations of striatal plasticity and motor behaviour that follow DA denervation.

Acknowledge support from the EC contract number LSHM-CT-2004-511995 (synScaff).

 

 

Basic mechanisms of striatal synaptic plasticity

Prof. Antonio Pisani
Department of Neuroscience
University of Rome “Tor Vergata” and Fondazione Santa Lucia, I.R.C.C.S., Rome (Italy)

Long-term changes occurring at excitatory corticostriatal synapses are critically involved in motor learning. Cortico-striatal glutamatergic afferents form asymmetrical synapses primarily with dendritic spines and dendritic shafts. Spiny neurons represent also the major synaptic target of the dopaminergic inputs to the striatum from midbrain areas. Dopaminergic terminals make symmetrical synaptic contact with necks of spines that also receive input, at their head, from terminals which form asymmetrical synaptic specializations. We have shown that repetitive stimulation of the corticostriatal pathway can cause either a long-lasting increase or an enduring decrease in synaptic strength, respectively referred to as long-term potentiation (LTP), and long-term depression, both requiring a complex sequence of biochemical events. Essential features of LTD are: co-activation of D1 and D2 dopamine receptors, activation of both a-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) and metabotropic glutamate (mGlu) receptors, whereas NMDA receptors are exclusively involved in LTP. Interestingly, after its induction, LTP can be reversed to control levels by a low-frequency stimulation protocol, an active process defined synaptic depotentiation. In Parkinson's disease (PD), the thalamo-cortical feedback loop is hypoactive. In the 6-hydroxydopamine rat model of PD, striatal synaptic plasticity is impaired, resembling the abnormality in thalamo-cortical circuitry, and chronic treatment with L-dopa is able to restore it. Of interest, a number of L-dopa-treated animals developed involuntary movements, resembling human dyskinesias. Strikingly, electrophysiological recordings from the dyskinetic group of rats demonstrated a selective impairment of synaptic depotentiation, compared to animals not showing involuntary movements. This involves a complex sequence of events, requiring the selective activation of D1 dopamine receptors.

These data support the relevance of striatal plasticity in the control of motor function, and suggest that its impairment has a crucial role in the pathogenesis both of PD and L-dopa-induced motor complications.

 

 

Genetic basis of Parkinson’s disease

Prof. Olaf Riess
Department of Medical Genetics
University of Tübingen, Tübingen (Germany)

Recent progress in the characterization of Parkinson’s disease (PD) patients provides increasing evidence for a genetic basis of the disease. Linkage analysis in some large families with autosomal dominant inheritance of the disease allowed the identification of several gene loci (PARK1, PARK3, PARK4, PARK5, and PAKR8) with subsequent characterization of important disease genes, a-synuclein (PARK1 and 4), UCHL1 (PARK5) and very recently of LRRK2 (PARK8). In other families with early onset PD and autosomal recessive inheritance additional gene loci were identified (PARK2, PARK6, PARK7) and mutations in the respective genes characterized (parkin, PINK1, and DJ1). Functional studies implicated that the PARK2 gene product, parkin, encodes a ubiquitin ligase. Based on the fact that parkin ubiquitinates a-synuclein a model of altered protein degradation as the cause for PD has been proposed. This model currently stimulates the genetic analysis of other candidate genes and led to the identification of a mutation in the ubiquitin C-terminal hydrolase L1 gene (UCH-L1). The identification of rare mutations in PD patients in potentially involved gene products of this pathway as in 14-3-3 genes, periphilin or in the a-synuclein interacting protein synphilin 1 is in particular intriguing. These studies are complemented by the first successful whole genome mapping approaches with indications for linkage on chromosome 5q, 8p, and 17q, respectively. Clinically it becomes obvious that the phenotype is broader than previously anticipated. It is now an obvious demand to identify the link between the different altered proteins and the mechanisms leading to cell death of dopaminergic neurons primarily in the Substantia nigra. Cellular and animal models have been generated to serve as a valuable tool to explore the underlying mechanism.

 

 

Clinical aspects of Parkinson´s disease

Prof. Thomas Klockgether
Clinic of Neurology
University of Bonn, Bonn (Germany)

Parkinsonism is a clinical syndrome characterized by akinesia, muscular rigidity and resting tremor. The most frequent cause of parkinsonism is Parkinson´s disease (PD). Progressive loss of substantia nigra neurons along with occurrence of Lewy bodies are considered essential neuropathological features of PD. Recent neuropathological studies suggest that nigral degeneration is only part of a more extended brain degeneration that starts in the medulla oblongata and then spreads to the mesencephalon and cerebral cortex. Correspondingly, the clinical symptoms occurring in PD go far beyond parkinsonism. Depending on the disease stage, autonomic dysfunction, olfactory disturbances, depression, and dementia are frequently encountered in PD. These neuropathological and clinical observations have major implications for future research in PD. In particular, it will be essential to analyze which properties the neuronal cell types involved in PD have in common that might make them susceptible to degeneration.
As a consequence of nigrostriatal degeneration, there is severe loss of dopamine in the striatum. Current medical therapy with levodopa and dopamine receptor agonists aims to compensate for striatal dopamine loss. While this therapy is very effective in ameliorating the motor symptoms of PD, it does not influence the non-motor symptoms which are due to brain degeneration beyond the nigrostriatal system. Development of effective therapies for these symptoms is a major future challenge.

 

 

Psychiatric aspects of Parkinson’s Disease

Prof. Donatella Marazziti
Department of Neurosciences
University of Pisa, Pisa (Italy)

Parkinson’s disease (PD) may be often accompanied by psychiatric disorders which, sometimes, occur some years before the onset of extra-pyramidal symptoms. The most common complication is depression. Neuropsychiatric sequelae include sub-cortical dementia, dysthymia, anxiety disorders, obsessive-compulsive symptoms, sleep disorders, and sexual disorders. Panic attacks are particularly common.

Depression
Parkinson himself in his studies described as, along with the motor impairment, his patients frequently showed unhappiness, discouragement, melancholy. Subsequently this phenomenon has received increasing attention and the frequency of depressive symptoms in PD patients has been reported in different studies with a range between 20 and 90%. This wide range may be addressed to the great symtoms overlapping between PD and depression. For a psychiatric diagnosis of depression, in addition to mood changes, symptoms such as psychomotor slowing, facial expression, voice tone and neuro-vegetative alterations are commonly considered. However, most of these symptoms may be also referred to PD itself and/or to the effects of PD treatments and this may constitute a difficulty for an appropriate differential diagnosis, especially in case of mild depression. Accumulating evidences suggest that depression in PD imay be secondary to the underlying neuro-anatomical degeneration rather than simply a reaction to the psychosocial stress and disability. The incidence of depression is correlated with changes in central serotonergic function and neurodegeneratron of specific cortical and sub-cortical pathways (McDonald 2003), but also to alterations in lower frontal lobe area.

A majority of PD patients suffer from mild to moderate depression; suicidal risk is rare and not correlated to the severity of the illness and/or the antiparkinsonian treatments. PD patients with depression seem to represent a subgroup of PD population with a positive family history of psychiatric disorders, more early onset of motor disability, more rapid progression of motor symptoms (Starkstein 1990).

The presence of depression would be associated with a more severe and rapid cognitive decline (Huber 1988).

Anxiety Disorders
Up to 40% of patients with PD experience clinically significant anxiety which may be a psychological reaction to the stress of the illness or may be related to the neurochemical changes of the disease itself. Generalized anxiety disorder, panic disorder, social phobia, phobic disorder, agoraphobia, obsessive-compulsive disorder and anxiety disorder not otherwise specified have all been diagnosed in patients with PD (Mensa 1993). The prevalence of panic attacks plus depression in PD’s patients is higher than in the general population. Norepinephrine, serotonin, dopamine and gamma amino butyric acid have been implicated in the pathophysiology of anxiety (Nutt 1992) and abnormalities of these neurotransmitter systems have been shown in PD patients (Mayeux 1986).

Anxiety in PD patients could involve a dopaminergic deficit directly or could be due to interactions between dopaminergic deficits and the variable deficits in norepinephrine and serotonin that are known to occur in PD. Both the ventral tegmental area and the locus coeruleus (which give rise to mesolimbic and noradrenergic pathways respectively) show significant neuronal loss in PD. Dopamine inhibits the rate of firing of the locus coeruleus and the loss of dopaminergic inhibition could explain the high prevalence of anxiety disorders in patients with PD (Iruela 1992).

A functional polymorphism in the promoter region of the serotonin transporter gene has recently been linked to anxiety. Menza et al. (1999) reported that patients with PD who carried the short allele of the serotonin transporter scored significantly higher than non-carriers on anxiety scales.

The temporal relationship between panic attacks and off periods has led to the hypothesis that panic attacks may be related to decreased brain levo-dopa levels. Anxiety fluctuations may be an important component of levo-dopa induced fluctuations (Maricle 1995), while showing that anxiety levels decrease and motor performance improved during levodopa infusion.

It has been reported that the degree of comorbidity between anxiety and depression in patients with PD is higher, as compared to that found in patients with other neurological disorders (Copeland 1992).

Obsessive-compulsive disorder (OCD)
Several structural and functional neuroimaging studies have shown that OCD is related to dysfunction of the basal ganglia (Robinson D. Arch Psych 1995). Lesions in the basal ganglia may provoke obsessive-compulsive symptoms similar to idiopathic obsessive-compulsive disorder. Patients with PD show several dysfunctions of fronto-basal ganglia circuitry. Alterations in the limbic circuitry may also be responsible for the occurrence of obsessive-compulsive traits in patients with PD. Tomer et al. (1993) described a relation between obsessive-compulsive symptomatology and severity of motor impairment in PD. Others (Alegret M et al 2001, Lang et al. 1998) showed that obsessive-compulsive symptoms appeared late during the disease progression in patients with idiopathic PD. This suggests that the emergence of obsessive symptoms could be directly related to the subset of neurochemical changes taking place at the level of the basal ganglia circuitry as the disease progresses. The functional disturbances produced by degeneration of the nigro-striatal pathway could influence the striato-frontal circuitry in the advanced stages of PD.

Psychosis
Psychotic symptoms are frequent complications of PD and affect between 15 and 40% of patients (Fenelon 2000). The most common are represented by visual hallucinations, which range from benign visual hallucination with insight to florid hallucinations and delusions which can be associated with disruptive behavour. The development of psychosis in idiopathic PD is virtually always drug-induced, but factors such as older age and cognitive impairment are also relevant in its occurrence (Holroyd 2001). For example in a community based sample, patients with dementia had higher rates of psychosis (40%) relative to those without dementia (15%) (Aarsland 1999). Marsh et al. (2004) reported that PD patients with psychosis have a higher rate of comorbid psychiatric disorders, especially affective syndromes and this additional psychopathology is associated with greater cognitive impairment and caregiver distress. Affective disturbance, including depression, anxiety and apathetic syndromes are common in PD patients with psychosis. Onset of affective disturbances preceded or co-occurred with the onset of psychotic symptoms (Marsh et al. 2004). Sleep-wake disturbances, which are common in depression and anxiety, could predispose to development of psychosis (Juncos 1999). Imbalance between dopaminergic and serotonergic systems or overstimulation of serotonergic receptor by dopaminergic medications may be relevant when there are comorbid affective disorders, which are known to be associated with serotonergic deficits (McDonald 2003). Higher depressive symptoms scores in PD patients with psychosis were associated with an apomorphine-induced growth hormone response, an index of central dopaminergic sensitivity (Mellers 1995). Thus, efficacy and tolerability of atypical antipsychotic agents in PD-related psychosis may be related to their potent anti-serotonergic properties, but reduced dopamine D2 receptor blockade (Zoldan 1996). Dopaminergic antiparkinsonian medications and secondary dopamine receptors hypersensitivity and overstimulation in mesocortical and mesolimbic regions are most commonly implicated in the pathophysiology of psychosis in PD (Graham 1997). However, development of psychosis is not directly related to the dose of dopamino-mimetic medications or levo-dopa plasma levels (Goetz 1998).

Since cognitive deficits in PD are associated with cholinergic deficits (Nakano 1984) and the most consistent clinical correlate of psychosis is cognitive impairment, cholinergic deficits are also likely to play a role in the development of psychosis. Therapeutically, cholinesterase inhibitors in PD patients are reported to reduce psychosis (Reading 2001) and, to improve cognitive impairments in PD (Leroi 2004).

 

 

Diagnosis and treatment of dementia associated with PD

Prof. Marco Onofrj
Department of Oncology and Neurosciences
University “G. D'Annunzio”, Chieti (Italy)

Parkinson’s Disease Dementia (PDD) has recently been debated, because its classification as an entity akin to Lewy Bodies Dementia (LBD) appears yet controversial. Specific clinical aspects of LBD are defined by Consensus Criteria, which consist of cognitive decline accompanied by cognitive fluctuations, visual hallucinations, parkinsonism, neuroleptic hypersensitivity, frequent syncopes and falls. Not listed by the Consensus is the relevant occurrence of REM sleep behaviour disorder (RBD) and SOREM with cataplexy.
Parkinson’s Disease (PD) is frequently accompanied by cognitive alterations (90%), more frequent in the visuo-spatial and executive domain, but the prevalence of dementia in patients above 65 years of age is reported, variably, around 15-40%.
Few studies described the characteristics of PDD, showing that visuo-spatial alterations and disesecutive symptoms are a common feature of PDD and LBD, but the occurrence of cognitive fluctuations is described only in one study; the incidence of syncopes has not been studied, neuroleptic hypersensitivity is only anecdotally described, scintigraphy studies are lacking. RBD occurs in PD yet comparative studies of its incidence in LBD and PDD are lacking. Visual hallucinations are a common feature of PD, LBD, PDD, but their expression is probably dependent on the different therapeutic approaches in the three diseases.
Despite the evident necessity for further studies on the clinical expression of PDD, there is already sufficient evidence, obtained in a numerically adequate double blind study, that the acetyl-butirryl- cholinesterase inhibitor Rivastigmine can be of remarkable benefit in PDD, by improving cognitive and neuropsychiatric symptoms including hallucinations. An adequate sized study on Donepezil is also in its final path.
In the rivastigmine study, benefits were observed in all secondary efficacy measures.
Significant decline was observed in the placebo group and this decline was more marked in the hallucinators. Hallucinations appear to predict greater therapeutic benefit from rivastigmine treatment in PDD.

References
McKeith IG, et al. Neurology. Consensus guidelines for the clinical and pathologic diagnosis of dementia with Lewy bodies (DLB): report of the consortium on DLB international workshop. 1996 Nov;47(5):1113-24.
Emre M, et al Rivastigmine for dementia associated with Parkinson's disease. N Engl J Med. 2004 Dec 9;351(24):2509-18.
Onofrj M, Thomas A, Iacono D, Luciano AL, Di Iorio A. The effects of a cholinesterase inhibitor are prominent in patients with fluctuating cognition: a part 3 study of the main mechanism of cholinesterase inhibitors in dementia. Clin Neuropharmacol 2003 Sep-Oct;26(5):239-51.
Onofrj M, Thomas A.. Severe worsening of parkinsonism in Lewy body dementia due to donepezil. Neurology 2003 Nov 25;61(10):1452.
Onofrj M Thomas A Acute Akinesia. Neurology 2005 Apr.6 in press
Onofrj M, Thomas A Albani G, Mauro A, Bulla D, Bonanni L A double crash system underlies the occurrence of Visual Hallucinations. J. Neurol. Sci.2005 in press

 

 

New horizons in the treatment of Parkinson’s disease

Prof. Paolo Del Dotto
Department of Neurology
Versilia Hospital, Lucca (Italy)

Many of the motoric features that define Parkinson’s disease (PD) result primarily from the loss of dopaminergic neurons of the substantia nigra. Levodopa remains to this day the most powerful symptomatic drug for the treatment of this condition. However, motor complications of chronic levodopa treatment have emerged as a major limitations of this therapy. Basic research has focused on better understanding of the mechanism of motor complications and how to prevent them. Slowing or delaying the progression of the disease with neuroprotective therapies may delay the need for levodopa. In the past few years, novel insight into the pathogenetic mechanisms of neurodegeneration in PD have been provided by the discoveries of genes responsible for rare monogenic parkinsonoan syndromes. Compelling evidence is accumulating, suggesting that the products of several of these genes can interact with environmental toxins and intervene in molecular pathways controlling the functional integrity of mitochondria and proteasome-ubiquitin system, leading to protein aggregation and production of Lewy bodies. Increased oxidative stress, apoptosis, excitotoxicity, and inflammation are also part of these processes that ultimately result in neurodegeneration. Drugs that are now under both preclinical and clinical scrutiny as neuroprotectant include molecules that combine one or more of the following properties: (1) monoamine oxidase inhibition (rasagiline, safinamide); (2) mitochondrial enhancement (coenzyme Q10, creatine); (3) antiapoptotic activity; (4) anti-inflammatory activity; (5) protein aggregation inhibition; (6) nitric oxide production; 7) neurotrophic activitiy.
In advanced Parkinson's disease, the combination of disease progression and levodopa therapy leads to the development of motor problems complicating the therapeutic response, known as motor response complications, particularly wearing off, on off and peak-dose dyskinesias. The nonphysiologic pulsatile stimulation of striatal dopamine receptors, produced by the currently available dopaminergic drugs, may trigger a dysregulation of many neurotransmitter systems within the basal ganglia, mainly localized on medium spiny striatal neurons. These include alterations of glutamatergic, serotonergic, adrenergic receptors and adenosine A2A receptors. Novel strategies for pharmacological intervention with nondopaminergic treatments hold the promise of providing effective control or reversal of motor response complications. Of particular interest are NMDA or AMPA antagonists (talampanel), or drugs acting on 5-hydroxytryptamine subtype 2A (sarizotan), alpha2-adrenergic (fipamezole), adenosine A2A (istradefylline) and cannabinoid CB1 receptors. Future strategies may also target pre- and postsynaptic components that regulate firing pattern of basal ganglia neurons, such as synaptic vesicle proteins, nonsynaptic gap junction communication mechanisms, or signal transduction systems that modulate the phosphorylation state of glutamatergic receptors.
Finally, there is great enthusiasm over the perspectives deriving from cell therapies in PD, that tend to replace the lack of dopamine in striatum through the nigral or striatal implant of different type of cells producing dopamine (fetal dopamine neurons, stem cells, spheramines). Double-blind controlled trials of transplanted dopamine neurons did not show significant benefit; moreover this technique was complicated by emergence of off-medication dyskinesia. Spheramine comprises retinal pigmented epithelial cells, adherent to spheroids, which release levodopa. A double-blind controlled trial to confirm the efficacy and safety of striatal spheramine implant is underway.

 

 

Advanced phase of PD: treatment of motor complications and psychosis

Prof. Letterio Morgante
Department of Neurosciences
University of Messina, Messina (Italy)

Although levodopa continues to be the most effective drug for relieving the motor symptoms associated with Parkinson’s disease (PD), multiple complications arise from long-term therapy, such as motor fluctuations and dyskinesias. Research has focused on levodopa-sparing drugs, including MAO inhibitors and dopamine agonists that provide not only symptomatic effects but also may exert some disease-modifying effect. While levodopa may be more effective than dopamine agonists in ameliorating motor symptoms, dopamine agonists are efficient to control mild symptoms in the early stages of the disease and in certain cases may be administered as a monotherapy for many years. Furthermore, the early treatment of PD with dopamine agonists, reduces the risk of developing the long-term therapy motor complications.
Non-motor features of PD include depression, dementia, nocturnal disturbances and chronic pain, all of which contribute to lowering patients’ quality of life. This presentation will cover some of these aspects with a major emphasis to depression.
The prevalence rates of depression in PD disease have been reported to be as high as 40%; depressive symptoms may precede the occurrence of motor symptoms in approximately 30% of patients. Despite the fact that depression seems to be a relevant aspect of PD, there are only a few randomized controlled studies designed to evaluate the efficacy of antidepressants in PD, all of which enrolled a small number of patients. Furthermore, several potential problems are associated with the evaluation of therapeutic effects of antidepressants in PD, including the use of rating scales not validated for depression in PD and the large placebo effect in depression trials. Selective serotonin reuptake inhibitors and tricyclic antidepressants are used widely in practice. Recent evidence indicates a possible role of the dopamine agonist, pramipexole, in reducing depressive symptoms in PD.

 

 

Neurologic and psychiatric profile of MS

Arturo Reggio
MS Centre
University of Catania, Catania (Italy)

Multiple Sclerosis (MS) is a demyelinating disease of the Central Nervous System (CNS), affecting young adulds. The prevalence of MS ranges from 30 to 200 cases and the incidence from 1 to 8 cases for 100.000 inhabitans.
It is well known that MS is caused by an autoimmune mechanism which leads to acute relapses with progressive accumulation of disability due the ongoing degenerative process. During MS several functional systems can be affected: pyramidal, cerebellar, visual, sphincteral, brainstem, sensory systems. Also mental functions can be involved with cognitive and emotional impairment. Neuropsychiatric dysfunctions in MS have been object of hystorical descriptions by Cruveilhier, Vulpian and Charcot who described mental changes, memory difficulties, indifference, blunting of emotions, pathological laughing and crying and classic forms of mental disease. In the last decades psychometric tests clarified the differences between cognitive impairment and affective changes such as depression, euphoria, anosognosia, , bipolar disorder, antisocial behaviour.
Aim of this talk will be to highlight the possible causes and the features of emotional functions in MS.

 

 

Genetics of demielinizating diseases

Prof. Luigi Grimaldi
Foundation Institute San Raffaele, G. Giglio Hospital
Cefalù (Italy)

Multiple sclerosis (MS) is the most common chronic inflammatory neurological disease. Despite major advances the aetiology of this disease it is still not completely understood. Susceptibility to MS is determined by both inherited and non-inherited factors. The importance of genetic factors is demonstrated by the increased risk of disease in relatives of MS patients. Generally speaking, the models that best reproduce the observed values have multiple loci with strongly synergistic interaction and autosomal dominant inheritance. At least six loci are required, with no upper limit on the number of loci. Models with genetic heterogeneity, where only a fraction of the risk loci are required for disease, are possible. In models with large numbers of loci the "abnormal" alleles conferring risk of disease are the most common allele. A variety of genetic models with multiple genes, dominant inheritance, and synergistic interaction between risk genes are consistent with the observed familial recurrence rates in MS. The search for individual loci is in progress. The genetic association of susceptibility to MS with loci outside the MHC has been reconfirmed. Evidence of parent-of-origin and seasonal effects on disease susceptibility add further complexity to the genetics of MS. The search for MS susceptibility genes continues using the candidate-gene approach as well as large-scale single-nucleotide-polymorphism association studies and novel cross-species synteny analysis. Genome-wide expression profiling using microarrays produced numerous therapeutic targets and is progressing towards profiling of rare cells. In summary, the complexity of MS is clearly reflected in the latest findings using global profiling methods. Nevertheless, these new technologies are confirming some of the basic aspects of the disease pathophysiology, i.e. its polygenicity, the central role of neuroinflammation and the emerging neurodenegerative processes. These data are primarily the results of genomic approaches, yet promising attempts are also made using proteomics and metabolomics.

 

 

Tissue damage and clinical outcome in MS

Prof. Carlo Pozzilli
Department of Neurological Science
University of Rome “La Sapienza”, Rome (Italy)

In the last decade remarkable progress has been done in the understanding of Multiple Sclerosis (MS). A key role has been played by the introduction of Magnetic Resonance Imaging (MRI) technique in the diagnostic process as well as in the definition of the natural history of the disease.
Many short-term studies focus on measuring the number and volume of gadolinium (Gd) enhancing lesions on T1-weighted images (T1 WI) as a sensitive detector of acute inflammatory activity, while longer-term studies assess the volume of hyperintense lesions on T2-weighted images (T2 WI) as a measure of more permanent tissue damage. Several studies on the natural history of multiple sclerosis have consistently shown that the presence of enhancing lesions portends an increased risk of clinical relapses and forecasts subclinical demyelination in untreated patients with CIS and RRMS. Additionally, in patients with clinically isolated syndrome (CIS) suggestive of MS the lesion burden on T2-weighted MRI sequences at baseline –i.e. at the time of the clinical presentation- predicts the clinical course of the disease and the extent of disability. However, hyperintense lesions on T2-weighted images provide a nonspecific measure of the overall extent of macroscopic tissue injury. Increased signal intensity on these scans in large measure reflects increased tissue water. The abnormal tissue water may be associated with different pathological substrates, ranging from edema and inflammation to irreversible demyelination and axonal loss and, consequently, could represent different clinical outcomes.
Specific MRI techniques have recently been developed that offer the prospect of selectively monitoring specific aspects of the pathological process. Examples include T1 hypointense lesion volume quantification, MR spectroscopy, magnetisation transfer imaging and measures of both brain and spinal cord atrophy . T1 hypointense lesions (“black holes”) provide in vivo evidence of axonal loss and they are strongly correlated with clinical disability in MS. Among patients with MS, men had a significant lower number of Gd-enhancing lesions but higher proportion of lesions that developed into T1 hypointense lesions compared to women. These data suggest that male patients with MS are prone to develop less inflammatory lesions but more destructive lesions than female patients. Factors influencing development of T1 hypointense lesions are partially unknown. Hypointense lesions may develop as a result of the extent and severity of the inflammatory processes or as consequence of exhaustion of repair mechanisms (i.e. remyelination). An influence of sex hormonal balance on disease activity and a specific role of estrogen and testosterone in modulating the development of tissue damage has been suggested on the basis of the relationship between serum levels of sex hormones and MRI features.

 

 

Present and future therapeutic options in Multiple Sclerosis (MS)

Prof. Giovanni Savettieri
Department of Neurology Ophthalmology, Otolaryngology and Psychiatry
University of Palermo, Palermo (Italy)

The major objectives of treatment in MS are: 1) reduce of relapse rate; 2) prevent disability due to relapses; 3) prevent disability due to progression of disease; 4) provide symptomatic treatment .
The reduction of relapses rate has been faced by treatment with interferons. This therapy is able to reduce annual relapse rate by 30-37% with respect to placebo treated group. Three other agents are used to reduce relapse frequency: glatiramer acetate, azathioprine and mitoxantrone. It has been demonstrated that glatiramer acetate reduce the frequency of annual relapse by 25%. Concerning azathioprine the amount of the trial data are insufficient to reach conclusions. Because of its toxicity, the use of mitoxantrone is limited.
The disability attributable to relapses is currently managed by the treatment with corticosteroids. The use of Interferons in the prevention of disability due to disease progression (that corresponds to the treatment of secondary progressive form of MS) is controversial and still debated.
At present other therapeutic procedure direct to reduce disease activity are under investigation. One of the approach is concerning the combination of established drugs (i.e. interferons and cyclophosphamide), other are direct to manipulate the interaction between Myelin Basic Protein (MBP), T cells, and antigen presenting cells. Other treatments under investigation through clinical trials are direct to interfere with the entry of activated T cells into the brain (i.e. monoclonal antibodies against integrins).

 

 

Are lipid-lowering agents the neuroprotective drugs of the future?

Prof. Regis Bordet
Department of Pharmacology, School of Medicine
University of Lille, Lille (France)

Statins and fibrates have been yet demonstrated their ability to prevent both cardio-vascular events and stroke. While this preventive effect has been initially related to their lipid-lowering effects, in particular hypocholesterolemic effect, analysis of primary or secondary prevention trials suggest that preventive effects could be partly independent from effects on lipid disorders. Thus, pleïtropic effects, such as vascular, anti-inflammatory or anti-oxidants effects, has been recently described. These cellular protective effects could have therapeutic interest to induce neuroprotection because of activation of cytoprotective proteins or regulation of deleterious molecular pathways involved in neuronal death. In stroke, statins or fibrates have been demonstrated experimentally, and more recently clinically, to induce a preventive or acute neuroprotection related to their pleïotropic anti-inflammatory and antioxidant properties. Beyond pharmacological effects in cerebrovascular diseases, these lipid-lowering agents have been also studied in neurodegenerative diseases. Experimental and clinical data support hypothesis that this therapeutic strategy could be also effective in neurodegenerative diseases such as Parkinson’s disease or Alzheimer’s disease. Moreover, recent data suggest also potential effects on neuroplasticity suggesting that this class of drugs could be promising for treatment of several cerebral diseases.

 

 

Proapoptotic cytokines in Alzheimer’s disease

Prof. Giuseppina Cantarella
Department of Experimental and Clinical Pharmacology
University of Catania, Catania (Italy)

Alzheimer’s disease (AD), is a high social impact disorder characterized by prominent, progressive neuronal loss. As pathogenesis of AD has not been, so far, entirely elucidated, currently available AD therapy is insufficient for the control of the illness.
Among a number of pathogenetic factors candidate promoters of AD, a crucial role is today recognized to amyloid-b, an anomalous peptide molecule synthesized in substantial amounts in the brain of individuals affected such disorder. Although most mechanisms related to formation of deposit of Ab in the brain are still poorly understood, an array of data suggests that inflammation may effectively fuel progression of AD-related pathological processes.
In this line, proinflammatory/proapoptotic cytokines detected in the brain of AD patients have been regarded as pivotal mediators of local inflammatory phenomena. However, while certain cytokines are normally expressed in the central nervous system (CNS), others, such as, for example, tumor necrosis factor related apoptosis inducing ligand (TRAIL), a molecule belonging to the TNF superfamily, are not detectable in the normal brain tissue. On the other hand, these are expressed in the pathological brain, as immunohistochemical data demonstrate that TRAIL-like immunoreactivity is localized in either neuronal and glial cells in the vicinity of Ab plaques in the human AD CNS. Gene array experiments, showing increased expression of TRAIL in human neurons treated with Ab, corroborate the hypothesis that TRAIL plays a role as a mediator of the Ab detrimental effects upon neurons. Finally, TRAIL displays intrinsic, caspase-dependent toxicity on neural cells and, moreover, its effects eventually encompass inflammatory switch on of glial cells.
It is noteworthy that in vitro immunoneutralization of TRAIL results in complete and prolonged abrogation of neurotoxic effects of Ab. Such Ab-TRAIL neurotoxic network supplies a rationale for an AD drug therapy benefitting of a contribution by specific TRAIL receptor antagonists.
The bulk of data is suggestive that the proapoptotic cytokine TRAIL and related molecules are redundantly contributing to Ab dependent neurotoxicity, involving different CNS cell types.
Finally, on the basis of actual data, the proinflammatory/proapoptotic cytokine molecular machinery could be regarded as a potential relevant target for development and application of efficacious, innovative AD drug therapy.

 

 

Immune response mediators and cells in Amyotrophic Lateral Sclerosis

Prof. Tiziana Mennini
Laboratory for Receptor Pharmacology
Negri Institute for Pharmacological Research, Milan (Italy)

In the last few years the possible contribution of TNF-alpha (TNF) in inducing motoneuron death in human amyotrophic lateral sclerosis (ALS) and in murine models of ALS has been widely investigated. An increasing body of evidence seems to confirm the hypothesis that neuroglial activation might produce an additive source of stress to injured motoneurons.

Animal models of ALS provide a useful tool to study the time-course of immune response in relation to motoneuron degeneration. Among these, the wobbler mouse is one of the most studied, although the gene mutation associated to the wobbler motoneuron disease is still unknown. This is a hereditary, autosomal, recessive disease with a multisystemic clinical pattern. Wobbler mice show rapid and selective vulnerability of cervical motoneurons, whose number is reduced by about 65% from the 3rd to the 12th week of age. Denervation, axonal swelling, microtubular aggregation and reduction in the number and diameter of myelinated fibers are typical features of this axonopathy. Affected animals show a focal atrophy of forelegs, motor impairment and severe muscular weakness.

In order to elucidate the correlation between neuroinflammation and motoneuron death, we studied the possible contribution of TNF on the degenerative process occurring in the cervical spinal cord of wobbler mice. Immunohystochemical experiments, were performed to ascertain if microglia, TNF and TNFR1 were overexpressed in the cervical spinal cord of 4 week-old wobbler mice. In addition, since the coupling TNF/TNFR1 often leads to the activation of an extrinsic apoptotic cell death pathway,we observed whether caspase -8 and -3 and DNA fragmentation (INSEL staining) were increased in degenerating motoneurons of wobbler mice.

A marked increase of activated microglial cells, particularly close to degenerating areas, and a clear expression of TNF and TNFR1 were observed in the cervical region of early symptomatic wobbler mice. However, neither significant increase in active caspase -8 and -3 positive cells nor evidence of DNA fragmentation in nuclei of affected motoneurons were observed.

In conclusion immunohystochemical and pharmacological results suggest that the activation of TNF /TNFR1-induced pathways can be responsible for the amplification of degenerative process occurring in the cervical motoneurons of wobbler mice. However, although the activation of TNFR1-associated pathways are in general correlated to a process of extrinsic apoptosis, no evidence of apoptosis emerged from our study. Future experiments aimed at distinguishing which selective pathway (the FADD-DED-PROCASAPASE 8 or the RIP-TRAF2-JNK/NFkb/AKT) is activated by TNF-/TNFR1 coupling are in progress to understand the actual role of TNF in motoneurons death.