Network analysis of gene expression in mice provides new evidence of involvement of the mTOR pathway in antipsychotic-induced extrapyramidal symptoms.

To identify potential candidate genes for future pharmacogenetic studies of antipsychotic (AP)-induced extrapyramidal symptoms (EPS), we used gene expression arrays to analyze changes induced by risperidone in mice strains with different susceptibility to EPS. We proposed a systems biology analytical approach that combined the identification of gene co-expression modules related to AP treatment, the construction of protein-protein interaction networks with genes included in identified modules and finally, gene set enrichment analysis of constructed networks. In response to risperidone, mice strain with susceptibility to develop EPS showed downregulation of genes involved in the mammalian target of rapamycin (mTOR) pathway and biological processes related to this pathway. Moreover, we also showed differences in the phosphorylation pattern of the ribosomal protein S6 (rpS6), which is a major downstream effector of mTOR. The present study provides new evidence of the involvement of the mTOR pathway in AP-induced EPS and offers new and valuable markers for pharmacogenetic studies.

A genetic dissection of antipsychotic induced movement disorders.

BACKGROUND: Antipsychotic medications (APM) are the first line pharmacological treatment for psychotic disorders and other behavioral disorders. Nevertheless, their use causes a number of side effects, including extrapyramidal symptoms (EPS). EPS decrease the efficacy of the antipsychotic treatments by causing poorer compliance to the treatment, stigma and a poorer quality of life for patients. Genetic studies hold the potential to unravel the molecular underpinnings of the EPS induced by APM but results are not conclusive and are far to be used in clinical practice despite decades of research. A more sophisticated selection of the list of genetic mutations explaining the genetic variance of EPS induced by APM could help in the definition of a personalized treatments for patients. Moreover, it would increase the quality of the current treatments with APM. METHODS: We reviewed the literature searching for the genetic association studies focused on dystonia, parkinsonism, akathisia and tardive dyskinesia. Moreover, we reviewed the current biological knowledge of the APM induced side effects. Finally, we provide a reasoned list of candidate genes and their genetic variations, with the aim of identifying a list of candidates for APM induced EPS genetic investigations. RESULTS: Variations located within PIK3CA (phosphoinositide-3- kinase, catalytic, alpha polypeptide), PLA2G4A (phospholipase A2, group IVA, cytosolic, calcium-dependent), PRKCA (protein kinase C, alpha), PRKACG (Phosphatidylinositol-4,5-bisphosphate 3-kinase 110 kDa catalytic subunit gamma), ERK-1 (extracellular signalregulated kinase 1 (MAPK3)), ERK-2 (extracellular signal-regulated kinase 2 (MAPK1)), GNAS (guanine nucleotide binding protein (G protein), alpha stimulating activity polypeptide 1), PLCB1 (phospholipase C, beta 1 (phosphoinositide-specific)) and ITPR1 (inositol 1,4,5-triphosphate receptor type 1) were found to be relevant for APM induced EPS. Some of the genes are classical candidates for this kind of research, others were never investigated. For each of these genes we provide a list of variations that balances the limitations of multitesting with the advantages of the tagging approach. CONCLUSIONS: We undertook a review of the literature about the APM induced EPM to provide some rational genetic candidates to be tested in further genetic investigations.

MPTP intoxication in mice: a useful model of Leigh syndrome to study mitochondrial diseases in childhood.

The basal ganglia, which are interconnected in the striato-nigral dopaminergic network, are affected in several childhood diseases including Leigh syndrome (LS). LS is the most common mitochondrial disorder affecting children and usually arise from inhibition of the respiratory chain. This vulnerability is attributed to a particular susceptibility to energetic stress, with mitochondrial inhibition as a common pathogenic pathway. In this study we developed a LS model for neuroprotection trials in mice by using the complex I inhibitor MPTP. We first verified that MPTP significantly inhibits the mitochondrial complex I in the brain (p = 0.018). This model also reproduced the biochemical and pathological features of LS: MPTP increased plasmatic lactate levels (p = 0.023) and triggered basal ganglia degeneration, as evaluated through dopamine transporter (DAT) autoradiography, tyrosine hydroxylase (TH) immunohistochemistry, and dopamine dosage. Striatal DAT levels were markedly decreased after MPTP treatment (p = 0.003). TH immunoreactivity was reduced in the striatum and substantia nigra (p = 0.005), and striatal dopamine was significantly reduced (p < 0.01). Taken together, these results confirm that acute MPTP intoxication in young mice provides a reproducible pharmacological paradigm of LS, thus opening new avenues for neuroprotection research.

Increased hexosaminidase activity in antipsychotic-induced extrapyramidal side effects: possible association with higher occurrence in bipolar disorder patients.

Dystonic movements and Parkinsonism are frequently seen in gangliosidoses and these conditions have been reported to modify dopaminergic plasticity. We investigated whether the activity of hexosaminidase, a type-two ganglioside (GM2) degrading enzyme, correlates with drug-induced extrapyramidal system (EPS) side effects in psychiatric patients. We compared hexosaminidase activity in the lymphocytes of 29 EPS-positive patients, 13 EPS-negative patients, and 30 healthy volunteers. The activities of A and B isoforms of hexosaminidase were higher in EPS-positive patients than EPS-negative patients and healthy controls. Multivariate analysis suggested an interaction with increased B isoform activity and EPS side effects in female bipolar disorder patients. Higher levels of hexosaminidase enzyme activity may explain the frequent occurrence of antipsychotic-induced extrapyramidal side effects in mood disorder patients.

Cyp2d6*3, *4, *5 and *6 polymorphisms and antipsychotic-induced extrapyramidal side-effects in patients receiving antipsychotic therapy.

1. The aim of the present study was to examine the relationship between CYP2D6 polymorphisms and the risk of antipsychotic (AP)-induced extrapyramidal symptoms (EPS) in patients receiving AP treatment. The allele status for CYP2D6*3, CYP2D6*4, CYP2D6*5 and CYP2D6*6 was determined in 267 patients receiving AP therapy. Seventy-nine cases presenting with EPS (Simpson-Angus Scale 3) and 188 controls without EPS (Simpson-Angus > 3) took part in the study. 2. We found a non-significant over-representation of poor metaboliser genotypes among cases, but a significant association between the mutant homozygous genotype for CYP2D6*4 (odds ratio (OR) 4.1, 95% confidence interval (CI) 1.01-16, permutated P value 0.01) and the heterozygous genotype for CYP2D6*6 (OR 5.4, 95% CI 1.13-18, permutated P value 0.003) and the risk of suffering EPS. 3. These results suggest that the CYP2D6 genotype may be a contributory factor in the development of EPS in patients undergoing AP therapy.

Aicardi-Goutières syndrome (AGS).

In 1984, Jean Aicardi and Françoise Goutières described 8 children showing both severe brain atrophy and chronic cerebrospinal fluid lymphocytosis, with basal ganglia calcification in at least one member of each affected family. The course was rapid to death or a vegetative outcome. Aicardi and Goutières correctly predicted that the disorder would be genetic, but emphasised that "some features, especially the pleocytosis, may erroneously suggest an inflammatory condition". The increased interferon-alpha in affected children (Pierre Lebon, Paris) mimicked congenital viral infection, but the associated chilblains (pernio) pointed to lupus erythematosus and an autoimmune mechanism. Genetic research led by Yanick Crow has clarified these puzzling relationships in Aicardi-Goutières syndrome, a syndrome that now includes conditions previously known as microcephaly-intracranial calcification syndrome, pseudo-TORCH and Cree encephalitis. At the time of writing, Crow's team has discovered that over 80% of families with Aicardi-Goutières syndrome have mutations in one of four nuclease genes, the exonuclease TREX1 and the genes for all three subunits of the ribonuclease H2 enzyme complex. Aicardi-Goutières syndrome is both genetically and phenotypically heterogeneous, with a range of severity from life-threatening perinatal illness to mild late infancy onset. All infants of whatever genotype have increased interferon-alpha in the first year of life and this appears to be the final common pathway that links Aicardi-Goutières syndrome, congenital virus infection and systemic lupus erythematosus.

[Imaging of brain acetylcholinesterase activity in dementias and extrapyramidal disorders].

Carbon-11 labeled N-methylpiperidin-4-yl acetate ([11C]MP4A) and carbon-11 labeled N-methylpiperidin-4-yl propionate ([11C]MP4P) are acetylcholine analogues and have been successfully used for measurement of brain acetylcholinesterase (AChE) activity in vivo in humans. In Alzheimer's disease (AD), there is a significant loss of AChE activity in the cerebral cortex in association with mental decline. The reduction of AChE activity in the cerebral cortex is more remarkable in early-onset AD than late-onset AD. There is mild but significant reduction of AChE activity in the cerebral cortex, even in the early stage of Parkinson's disease (PD) without dementia. There is remarkable reduction of AChE activity in the entire cerebral cortex in both PD with dementia and dementia with Lewy bodies (DLB). In two patients with frontotemporal dementia with parkinsonism linked to chromosome 17, there was prominent reduction of AChE activity in the cerebral cortex and thalamus. In 12 patients with progressive supranuclear palsy, there was profound reduction of AChE activity in the thalamus but not in the cerebral cortex. In corticobasal degeneration, there is symmetrical loss of AChE activity in the sensori-motor cortex.

Clinical and biochemical characteristics in patients with a high mutant load of the mitochondrial T8993G/C mutations.

We retrospectively analyzed the clinical, histological, and biochemical data of 11 children, five of which carried the maternally-inherited mitochondrial T8993C and six carrying the T8993G point mutations in the ATP synthase 6 gene. The percentage of heteroplasmy was 95% or higher in muscle and in blood. All patients had an early clinical presentation with muscle hypotonia, severe extrapyramidal dysfunction and Leigh disease demonstrated by the cranial MRI. A slower clinical progression and more frequent sensory-neuronal involvement were noted in the patients carrying the T8993C mutation in a high mutation load in muscle and blood. No histological abnormality was found. In 9 out of 11 patients a decreased ATP production was detected, and complex V activity was deficient in all children. The activities of the respiratory enzyme complexes II and IV were normal, whereas an associated combined complex I and III deficiency were present in two patients. No obvious difference was found between the biochemical parameters of the two patient groups harboring different mutations in the same gene. No correlation was found between the degree of complex V enzyme deficiency and the severity of the phenotype. We confirmed an impaired assembly/stability of complex V in our patients. This is the first report of decreased activity and impaired assembly/stability of complex V in patients with T8993C mutations measured in muscle tissue.

Expression of 8-oxoguanine DNA glycosylase (OGG1) in Parkinson's disease and related neurodegenerative disorders.

Oxidative stress including DNA oxidation is implicated in Parkinson's disease (PD). We postulated that DNA repair enzymes such as 8-oxoguanosine DNA glycosylase (OGG1) are involved in the PD process. We performed immunohistochemical and biochemical studies on brains of patients with PD and those of patients with progressive supranuclear palsy (PSP) and corticobasal degeneration (CBD) as disease controls, and control subjects. We found higher expression levels of mitochondrial isoforms of OGG1 enzymes in the substantia nigra (SN) in cases of PD. Furthermore, Western blot analysis revealed high OGG1 levels in the SN of the patients with PD. Our results indicate the importance of oxidative stress within the susceptible lesions in the pathogenesis of PD.

Glial overexpression of heme oxygenase-1: a histochemical marker for early stages of striatal damage.

The level of heme oxygenase-1 (HO-1) in the normal striatum is below the limit of immunodetection. However, HO-1 is overexpressed in both neural and non-neural cells in response to a wide range of lesions. We induced different types of lesions affecting the striatal cells or the main striatal afferent systems in rats to investigate if overexpression of HO-1 could be a useful histochemical marker of striatal damage. Thirty-six hours after intrastriatal or intraventricular injection of excitotoxins that affect striatal neurons (ibotenic acid) or of neurotoxins that affect striatal dopaminergic (6-hydroxydopamine) or serotonergic (5,7-dihydroxytriptamine) afferent terminals, or after surgical lesioning of cortico-striatal projections, there was intense induction of striatal HO-1 immunoreactivity (HO-1-ir). Double immunolabeling revealed that the HO-1-ir was located in glial cells. After intrastriatal injection of ibotenic acid, a central zone of neuronal degeneration contained numerous round and pseudopodic HO-1-ir cells, and was surrounded by a ring of HO-1-ir cells, most of which were immunoreactive for astroglial markers. Intraventricular injection of neurotoxins induced astroglial HO-1-ir cells which were more evenly distributed throughout the lesioned or denervated areas. HO-1-ir microglial cells were also observed in areas subjected to mechanical damage. The HO-1-ir was markedly lower or absent 1 week after lesion, and even more so 3 weeks after, although some HO-1-ir cells were still observed after intrastriatal injection of ibotenic acid or surgical corticostriatal deafferentation. The results indicate that determination of glial HO-1-ir is a useful histochemical marker for early stages of striatal damage.

Adverse effects of dopamine potentiation by long-term treatment with selegiline.

A patient with triosephosphate isomerase (TPI) deficiency exhibited worsening of abnormal involuntary movements of the dystonic type and developed psychiatric symptoms while on selegiline. When selegiline was stopped after 9 years of treatment, abnormal involuntary movements improved to pretreatment level and psychiatric behaviour returned to normal. Monoamine oxidase-B platelet activity was low in this patient.

Neonatal dopa-responsive extrapyramidal syndrome in twins with recessive GTPCH deficiency.

The authors report two twin sisters, age 15 years, with recessive GTP cyclohydrolase deficiency, who presented with neonatal onset of rigidity, tremor, and dystonia but with no other symptoms suggestive of a diffuse CNS involvement. The plasma phenylalanine levels were normal. Treatment with L-dopa/carbidopa, started at age 1 year, was associated with sustained recovery from all neurologic signs. The patients were homozygous for a new recessive mutation in the GHI gene.

Antipsychotic-induced extrapyramidal syndromes and cytochrome P450 2D6 genotype: a case-control study.

To study the association between polymorphism of the cytochrome P450 2D6 gene (CYP2D6) and the risk of antipsychotic-induced extrapyramidal syndromes, as measured by the use of antiparkinsonian medication. Data for this case-control study were obtained from a psychiatric hospital where newly admitted patients are routinely screened for several CYP2D6 mutant alleles. Cases were patients prescribed antiparkinsonian medication during oral antipsychotic drug treatment in the period September 1994 to August 2000. They were divided into those using an antipsychotic drug the metabolic elimination of which depends on the activity of the CYP2D6 enzyme ('CYP2D6-dependent') and those using other antipsychotic drugs. We formed a control group of antipsychotic drug users for both case groups using a matching ratio of 3 : 1 (controls : cases). Control patients were matched on whether or not their prescribed antipsychotic drug was CYP2D6-dependent. Odds ratios for patients who were slow metabolizers versus patients who were extensive metabolizers were calculated using conditional logistic regression and were adjusted for age, gender, dose and other potential confounding factors. We identified 77 case patients who were prescribed a CYP2D6-dependent antipsychotic drug and 54 case patients who were prescribed non CYP2D6-dependent antipsychotic drugs. Among the case- and control-patients using a CYP2D6-dependent antipsychotic drug, the poor metabolizers were more than four times more likely to start with antiparkinsonian medication than the extensive metabolizers (odds ratio 4.44; 95% confidence interval 1.11-17.68). An increased risk was not observed for patients using non CYP2D6-dependent antipsychotic drugs (odds ratio 1.20; 95% confidence interval 0.21-6.79). Genetically impaired CYP2D6 activity can increase the risk of antipsychotic-induced extrapyrimidal syndromes. Poor metabolizers should have their antipsychotic drug dosage reduced when the metabolism of the prescribed drug depends on CYP2D6 activity or should receive an antipsychotic drug that is not CYP2D6-dependent.

Hypothalamic digoxin related membrane Na+-K+ ATPase inhibition and familial basal ganglia calcification.

The isoprenoid pathway produces three key metabolites-digoxin (membrane sodium-potassium ATPase inhibitor and regulator of intracellular calcium-magnesium ratios), dolichol (regulator of N-glycosylation of proteins) and ubiquinone (free radical scavenger). The pathway was assessed in a rare and specific type of familial basal ganglia calcification described. The family had a coexistence of basal ganglia calcification (six out of 10 cases), schizophrenia, Parkinson's disease, Alzheimer's disease, rheumatoid arthritis, systemic tumours and syndrome X and were all right hemispheric dominant. The isoprenoid pathway was also studied for comparison in right hemispheric dominant, bihemispheric dominant and left hemispheric dominant individuals. The isoprenoid pathway was upregulated with increased digoxin synthesis in familial basal ganglia calcification. Membrane sodium-potassium ATPase inhibition can lead on to increase in intracellular calcium and calcification of the basal ganglia. There was increase in tryptophan catabolites and a reduction in tyrosine catabolites. There was also an increase in dolichol and glycoconjugate levels with reduced lysosomal stability in these patients. The ubiquinone levels were low and free radical levels increased. The cholesterol-phospholipid ratio was increased and glycoconjugate level of the RBC membrane reduced in these group of patients. No significance difference was noted in family members with and without basal ganglia calcification. This findings were correlated with the pathogenesis of syndrome X, immune mediated diseases, degenerations, tumours and psychiatric disorders noted in the familial basal ganglia calcification described. The biochemical patterns obtained in familial basal ganglia calcification correlated with those in right hemispheric dominance.

Dopamine transporter and nitric oxide synthase in hypoxic-ischemic brain.

Changes in dopamine transporter and neuronal nitric oxide synthase (nNOS) were investigated by immunohistochemistry in 18 cases of hypoxic-ischemic basal ganglia necrosis. Neuropil dopamine transporter immunostaining in the striatum was increased in seven cases, with relatively mild basal ganglia necrosis, and decreased in four cases, with marked basal ganglia necrosis, compared with age-matched control subjects. Correspondingly, some striatal neurons had increased immunoreactivity to dopamine transporter in the cases of increased immunostaining in the neuropil. nNOS-positive neurons did not obviously change in cases of basal ganglia necrosis within 2 days after birth and then decreased or were not detectable in cases of basal ganglia necrosis at more than 3 days after birth. The results suggest that the synthesis of dopamine transporter is up-regulated in relatively mild basal ganglia necrosis to compensate for the uptake of increased dopamine, that this compensative ability is lost in marked basal ganglia necrosis, and that nNOS-containing neurons in the striatum are relatively resistant to hypoxic ischemia. We speculate that glutamate excitotoxicity mediated by glutamate receptors 1, 2/3, and 4 and excessive dopaminergic excitatory activity may play important roles in hypoxic-ischemic basal ganglia necrosis and that nNOS does not contribute to that condition.

Enhanced association of mutant triosephosphate isomerase to red cell membranes and to brain microtubules.

In a Hungarian family with triosephosphate isomerase (TPI; D-glyceraldehyde-3-phosphate keto-isomerase, EC 5.3.1.1) deficiency, two germ-line identical, but phenotypically differing compound heterozygote brothers (one of them with neurological disorder) have been identified with the same very low (<5%) TPI activity and 20- or 40-fold higher erythrocyte dihydroxyacetone phosphate levels as compared with normal controls. Our present studies with purified TPI and hemolysates revealed the binding of TPI, and the binding of human wild-type and mutant TPIs in hemolysate, to the red cell membrane, and the interference of binding with other hemolysate proteins. The binding of the mutant TPI is enhanced as compared with the wild-type enzyme. The increased binding is influenced by both the altered structure of the mutant and the changes in the red cell membrane. Compared with binding of glyceraldehyde-3-phosphate dehydrogenase, the isomerase binding is much less sensitive to ionic strength or blocking of the N-terminal tail of the band-3 transmembrane protein. The binding of TPIs to the membrane decreases the isomerase activity, resulting in extremely high dihydroxyacetone phosphate levels in deficient cells. In cell-free brain extract, tubulin copolymerizes with TPI and with other cytosolic proteins forming highly decorated microtubules as shown by immunoblot analysis with anti-TPI antibody and by electron microscopic images. The efficacy order of TPI binding to microtubules is propositus > brother without neurological disorder > normal control. This distinct microcompartmentation of mutant proteins may be relevant in the development of the neurodegenerative process in TPI deficiency and in other, more common neurological diseases.

Guanidinoacetate methyltransferase deficiency: a newly recognized inborn error of creatine biosynthesis.

In an infant with progressive, severe extrapyramidal movement disorder and extremely. low urinary creatinine excretion, in vivo proton magnetic resonance spectroscopy of the brain showed a depletion of creatine and an accumulation of guanidinoacetate, the immediate precursor of creatine. The suggested defect in creatine biosynthesis at the level of guanidinoacetate methyltransferase was confirmed by the demonstration of defective activity of this enzyme in liver tissue and by identification of the underlying genetic defect. Creatine substitution by means of oral creatine monohydrate at high dosage (4-8 g per day) resulted in a striking improvement of the extrapyramidal movement disorder, normalisation of abnormal slow background activity in the EEG, and disappearance of bilateral abnormal signal intensities in the globus pallidus. The low urinary creatine excretion normalized and brain creatine and creatine phosphate, as measured by in vivo magnetic resonance spectroscopy, increased significantly. Guanidinoacetate methyltransferase deficiency is a new, treatable inborn error of metabolism. Screening methods and non-invasive diagnosis of the enzyme defect are needed for the early detection and treatment of patients with this effect.

CYP 2D6 PM phenotype hypothesis of antidepressant extrapyramidal side-effects.

Extrapyramidal symptoms occur as side-effects of neuroleptics. For many years, case reports of such side-effects, linked to antidepressant treatments, have been published, but this phenomenon is not well known. Tricyclic and serotonergic antidepressants are both involved. The authors present an hypothesis which provides one possible neurobiochemical explanation for the aetiology of these side-effects. The proposed explanation is related to the inhibition of the CYP 2D6 isoenzyme by antidepressants (or neuroleptics) that may be involved in the genesis of the observed extrapyramidal side-effects.

Aciduria glutárica tipo I: una encefalopatía metabólica extrapiramidal / Type I glutaric aciduria: an extrapyramidal metabolic encephalopathy

Se describen tres niños, uno varón, de 4, 6 y 2 años de edad, afectados de aciduria glutárica tipo I. Su desarrollo fue normal hasta la segunda mitad del primer año de vida, cuando sufrieron alteración de conciencia y convulsiones, seguidas de pérdida de las habilidades adquiridas, distonía y movimientos anormales. La tomografía axial y resonancia nuclear magnética de cerebro mostraron atrofia frontotemporal y de los núcleos caudados y putámenes. Habíagran cantidad de ácidos glutárico, 3-hidroxiglutárico y glutacónico en orina y actividad disminuida de la enzima glutaril CoA deshidrogenasa en cultivos de fibroblastos de los tres niños, confirmándose así el diagnóstico de esta afección metabólica