RESUMO
The response to drugs of abuse is affected by expectation, which is modulated in part by dopamine (DA), which encodes for a reward prediction error. Here we assessed the effect of expectation on methylphenidate (MP)-induced striatal DA changes in 23 participants with an active cocaine use disorder (CUD) and 23 healthy controls (HC) using [11C]raclopride and PET both after placebo (PL) and after MP (0.5 mg/kg, i.v.). Brain dopamine D2 and D3 receptor availability (D2R: non-displaceable binding potential (BPND)) was measured under four conditions in randomized order: (1) expecting PL/receiving PL, (2) expecting PL/receiving MP, (3) expecting MP/receiving PL, and (4) expecting MP/receiving MP. Expecting MP increased pulse rate compared to expecting PL. Receiving MP decreased D2R in striatum compared to PL, indicating MP-induced striatal DA release, and this effect was significantly blunted in CUD versus HC consistent with prior findings of decreased striatal dopamine responses both in active and detoxified CUD. There was a group × challenge × expectation effect in caudate and midbrain, with expectation of MP increasing MP-induced DA release in HC but not in CUD, and expectation of PL showing a trend to increase MP-induced DA release in CUD but not in HC. These results are consistent with the role of DA in reward prediction error in the human brain: decreasing DA signaling when rewards are less than expected (blunted DA increases to MP in CUD) and increasing them when greater than expected (for PL in CUD reflecting conditioned responses to injection). Our findings also document disruption of the expectation of drug effects in dopamine signaling in participants with CUD compared to non-addicted individuals.
Assuntos
Encéfalo/metabolismo , Estimulantes do Sistema Nervoso Central/uso terapêutico , Transtornos Relacionados ao Uso de Cocaína/tratamento farmacológico , Dopamina/metabolismo , Metilfenidato/uso terapêutico , Recompensa , Adulto , Encéfalo/diagnóstico por imagem , Mapeamento Encefálico , Estudos de Casos e Controles , Estimulantes do Sistema Nervoso Central/farmacocinética , Transtornos Relacionados ao Uso de Cocaína/metabolismo , Transtornos Relacionados ao Uso de Cocaína/patologia , Feminino , Humanos , Masculino , Metilfenidato/farmacocinética , Pessoa de Meia-Idade , Tomografia por Emissão de Pósitrons , Racloprida/metabolismo , Receptores de Dopamina D2/metabolismo , Receptores de Dopamina D3/metabolismoRESUMO
The role of the protein kinase Akt1 in dopamine neurotransmission is well recognized and has been implicated in schizophrenia and psychosis. However, the extent to which variants in the AKT1 gene influence dopamine neurotransmission is not well understood. Here we investigated the effect of a newly characterized variant number tandem repeat (VNTR) polymorphism in AKT1 [major alleles: L- (eight repeats) and H- (nine repeats)] on striatal dopamine D2/D3 receptor (DRD2) availability and on dopamine release in healthy volunteers. We used PET and [11C]raclopride to assess baseline DRD2 availability in 91 participants. In 54 of these participants, we also measured intravenous methylphenidate-induced dopamine release to measure dopamine release. Dopamine release was quantified as the difference in specific binding of [11C]raclopride (nondisplaceable binding potential) between baseline values and values following methylphenidate injection. There was an effect of AKT1 genotype on DRD2 availability at baseline for the caudate (F(2,90) = 8.2, p = 0.001) and putamen (F(2,90) = 6.6, p = 0.002), but not the ventral striatum (p = 0.3). For the caudate and putamen, LL showed higher DRD2 availability than HH; HL were in between. There was also a significant effect of AKT1 genotype on dopamine increases in the ventral striatum (F(2,53) = 5.3, p = 0.009), with increases being stronger in HH > HL > LL. However, no dopamine increases were observed in the caudate (p = 0.1) or putamen (p = 0.8) following methylphenidate injection. Our results provide evidence that the AKT1 gene modulates both striatal DRD2 availability and dopamine release in the human brain, which could account for its association with schizophrenia and psychosis. The clinical relevance of the newly characterized AKT1 VNTR merits investigation.SIGNIFICANCE STATEMENT The AKT1 gene has been implicated in schizophrenia and psychosis. This association is likely to reflect modulation of dopamine signaling by Akt1 kinase since striatal dopamine hyperstimulation is associated with psychosis and schizophrenia. Here, using PET with [11C]raclopride, we identified in the AKT1 gene a new variable number tandem repeat (VNTR) marker associated with baseline striatal dopamine D2/D3 receptor availability and with methylphenidate-induced striatal dopamine increases in healthy volunteers. Our results confirm the involvement of the AKT1 gene in modulating striatal dopamine signaling in the human brain. Future studies are needed to assess the association of this new VNTR AKT1 variant in schizophrenia and drug-induced psychoses.
Assuntos
Corpo Estriado/metabolismo , Dopamina/biossíntese , Neurotransmissores/biossíntese , Polimorfismo de Nucleotídeo Único/genética , Proteínas Proto-Oncogênicas c-akt/fisiologia , Receptores Dopaminérgicos/metabolismo , Adulto , Disponibilidade Biológica , Feminino , Humanos , Masculino , Valores de Referência , Transmissão Sináptica/fisiologiaRESUMO
Monoamine oxidase (MAO) oxidizes amines from both endogenous and exogenous sources thereby regulating the concentration of neurotransmitter amines such as serotonin, norepinephrine, and dopamine as well as many xenobiotics. MAO inhibitor drugs are used in the treatment of Parkinson's disease and in depression stimulating the development of radiotracer tools to probe the role of MAO in normal human biology and in disease. Over the past 30 years since the first radiotracers were developed and the first positron emission tomography (PET) images of MAO in humans were carried out, PET studies of brain MAO in healthy volunteers and in patients have identified different variables that have contributed to different MAO levels in brain and in peripheral organs. MAO radiotracers and PET have also been used to study the current and developing MAO inhibitor drugs including the selection of doses for clinical trials. In this article, we describe the following: (1) the development of MAO radiotracers; (2) human studies including the relationship of brain MAO levels to genotype, personality, neurological, and psychiatric disorders; and (3) examples of the use of MAO radiotracers in drug research and development. We will conclude with outstanding needs to improve the radiotracers that are currently used and possible new applications.
Assuntos
Monoaminoxidase , Traçadores Radioativos , Radioquímica/métodos , Encéfalo/enzimologia , Descoberta de Drogas , Genótipo , Humanos , Monoaminoxidase/química , Monoaminoxidase/genética , Monoaminoxidase/metabolismoRESUMO
Selegiline (L-deprenyl) is a selective, irreversible inhibitor of monoamine oxidase B (MAO-B) at the conventional dose (10 mg/day oral) that is used in the treatment of Parkinson's disease. However, controlled studies have demonstrated antidepressant activity for high doses of oral selegiline and for transdermal selegiline suggesting that when plasma levels of selegiline are elevated, brain MAO-A might also be inhibited. Zydis selegiline (Zelapar) is an orally disintegrating formulation of selegiline, which is absorbed through the buccal mucosa producing higher plasma levels of selegiline and reduced amphetamine metabolites compared with equal doses of conventional selegiline. Although there is indirect evidence that Zydis selegiline at high doses loses its selectivity for MAO-B, there is no direct evidence that it also inhibits brain MAO-A in humans. We measured brain MAO-A in 18 healthy men after a 28-day treatment with Zydis selegiline (2.5, 5.0, or 10 mg/day) and in 3 subjects receiving the selegiline transdermal system (Emsam patch, 6 mg/day) using positron emission tomography and the MAO-A radiotracer [(11)C]clorgyline. We also measured dopamine transporter (DAT) availability in three subjects from the 10 mg group. The 10 mg Zydis selegiline dose significantly inhibited MAO-A (36.9±19.7%, range 11-70%, p<0.007)) but not DAT; and while Emsam also inhibited MAO-A (33.2±28.9 (range 9-68%) the difference did not reach significance (p=0.10)) presumably because of the small sample size. Our results provide the first direct evidence of brain MAO-A inhibition in humans by formulations of selegiline, which are currently postulated but not verified to target brain MAO-A in addition to MAO-B.
Assuntos
Encéfalo/efeitos dos fármacos , Encéfalo/enzimologia , Inibidores da Monoaminoxidase/farmacologia , Monoaminoxidase/metabolismo , Selegilina/farmacologia , Administração Cutânea , Administração Oral , Adolescente , Adulto , Encéfalo/metabolismo , Radioisótopos de Carbono/metabolismo , Clorgilina/metabolismo , Cocaína/metabolismo , Proteínas da Membrana Plasmática de Transporte de Dopamina/efeitos dos fármacos , Proteínas da Membrana Plasmática de Transporte de Dopamina/metabolismo , Relação Dose-Resposta a Droga , Neuroimagem Funcional , Humanos , Masculino , Inibidores da Monoaminoxidase/administração & dosagem , Tomografia por Emissão de Pósitrons , Selegilina/administração & dosagem , Adulto JovemRESUMO
OBJECTIVE: Dopamine mediates the rewarding effects of food that can lead to overeating and obesity, which then trigger metabolic neuroadaptations that further perpetuate excessive food consumption. We tested the hypothesis that the dopamine response to calorie intake (independent of palatability) in striatal brain regions is attenuated with increases in weight. METHOD: We used positron emission tomography with [11C]raclopride to measure dopamine changes triggered by calorie intake by contrasting the effects of an artificial sweetener (sucralose) devoid of calories to that of glucose to assess their association with body mass index (BMI) in nineteen healthy participants (BMI range 21-35). RESULTS: Neither the measured blood glucose concentrations prior to the sucralose and the glucose challenge days, nor the glucose concentrations following the glucose challenge vary as a function of BMI. In contrast the dopamine changes in ventral striatum (assessed as changes in non-displaceable binding potential of [11C]raclopride) triggered by calorie intake (contrast glucose - sucralose) were significantly correlated with BMI (râ=â0.68) indicating opposite responses in lean than in obese individuals. Specifically whereas in normal weight individuals (BMI <25) consumption of calories was associated with increases in dopamine in the ventral striatum in obese individuals it was associated with decreases in dopamine. CONCLUSION: These findings show reduced dopamine release in ventral striatum with calorie consumption in obese subjects, which might contribute to their excessive food intake to compensate for the deficit between the expected and the actual response to food consumption.
Assuntos
Índice de Massa Corporal , Dopamina/metabolismo , Ingestão de Energia , Glucose/farmacologia , Núcleo Accumbens/efeitos dos fármacos , Núcleo Accumbens/metabolismo , Adulto , Glucose/metabolismo , Voluntários Saudáveis , Humanos , Pessoa de Meia-Idade , Núcleo Accumbens/diagnóstico por imagem , Núcleo Accumbens/fisiologia , Tomografia por Emissão de Pósitrons , Edulcorantes/farmacologiaRESUMO
BACKGROUND: Gene polymorphisms that affect serotonin signaling modulate reactivity to salient stimuli and risk for emotional disturbances. Here, we hypothesized that these serotonin genes, which have been primarily explored in depressive disorders, could also have important implications for drug addiction, with the potential to reveal important insights into drug symptomatology, severity, and/or possible sequelae such as dysphoria. METHODS: Using an imaging genetics approach, the current study tested in 62 cocaine abusers and 57 healthy controls the separate and combined effects of variations in the serotonin transporter (5-HTTLPR) and monoamine oxidase A (MAOA) genes on processing of aversive information. Reactivity to standardized unpleasant images was indexed by a psychophysiological marker of stimulus salience (i.e., the late positive potential (LPP) component of the event-related potential) during passive picture viewing. Depressive symptomatology was assessed with the Beck Depression Inventory (BDI). RESULTS: Results showed that, independent of diagnosis, the highest unpleasant LPPs emerged in individuals with MAOA-Low and at least one 'Short' allele of 5-HTTLPR. Uniquely in the cocaine participants with these two risk variants, higher unpleasant LPPs correlated with higher BDI scores. CONCLUSIONS: Taken together, these results suggest that a multilocus genetic composite of monoamine signaling relates to depression symptomatology through brain function associated with the experience of negative emotions. This research lays the groundwork for future studies that can investigate clinical outcomes and/or pharmacogenetic therapies in drug addiction and potentially other psychopathologies of emotion dysregulation.
Assuntos
Monoaminas Biogênicas/metabolismo , Transtornos Relacionados ao Uso de Cocaína/genética , Transtornos Relacionados ao Uso de Cocaína/psicologia , Depressão/genética , Depressão/psicologia , Adulto , Transtornos Relacionados ao Uso de Cocaína/complicações , Depressão/complicações , Potenciais Evocados/fisiologia , Feminino , Genótipo , Humanos , Masculino , Pessoa de Meia-Idade , Monoaminoxidase/genética , Polimorfismo Genético/genética , Serotonina/fisiologia , Proteínas da Membrana Plasmática de Transporte de Serotonina/genéticaRESUMO
Functional polymorphisms in the dopamine transporter gene (DAT1 or SLC6A3) modulate responsiveness to salient stimuli, such that carriers of one 9R-allele of DAT1 (compared with homozygote carriers of the 10R-allele) show heightened reactivity to drug-related reinforcement in addiction. Here, using multimodal neuroimaging and behavioral dependent variables in 73 human cocaine-addicted individuals and 47 healthy controls, we hypothesized and found that cocaine-addicted carriers of a 9R-allele exhibited higher responses to drug cues, but only among individuals who had used cocaine within 72 h of the study as verified by positive cocaine urine screens (a state characterized by intense craving). Importantly, this responsiveness to drug cues was reliably preserved across multimodal imaging and behavioral probes: psychophysiological event-related potentials, self-report, simulated cocaine choice, and fMRI. Because drug cues contribute to relapse, our results identify the DAT1R 9R-allele as a vulnerability allele for relapse especially during early abstinence (e.g., detoxification).
Assuntos
Encéfalo/fisiopatologia , Transtornos Relacionados ao Uso de Cocaína/genética , Transtornos Relacionados ao Uso de Cocaína/fisiopatologia , Sinais (Psicologia) , Proteínas da Membrana Plasmática de Transporte de Dopamina/genética , Adulto , Alelos , Análise de Variância , Comportamento Aditivo/genética , Encéfalo/irrigação sanguínea , Encéfalo/patologia , Mapeamento Encefálico , Comportamento de Escolha , Cocaína/urina , Transtornos Relacionados ao Uso de Cocaína/patologia , Transtornos Relacionados ao Uso de Cocaína/urina , Eletroencefalografia , Emoções/fisiologia , Potenciais Evocados/fisiologia , Feminino , Humanos , Processamento de Imagem Assistida por Computador , Imageamento por Ressonância Magnética , Masculino , Pessoa de Meia-Idade , Testes Neuropsicológicos , Oxigênio/sangue , Estimulação Luminosa , Psicofísica , Reforço PsicológicoRESUMO
Aging is associated with reductions in brain glucose metabolism in some cortical and subcortical regions, but the rate of decrease varies significantly between individuals, likely reflecting genetic and environmental factors and their interactions. Here we test the hypothesis that the variant of the dopamine receptor D4 (DRD4) gene (VNTR in exon 3), which has been associated with novelty seeking and sensitivity to environmental stimuli (negative and positive) including the beneficial effects of physical activity on longevity, influence the effects of aging on the human brain. We used positron emission tomography (PET) and [(18)F]fluoro-D-glucose ((18)FDG) to measure brain glucose metabolism (marker of brain function) under baseline conditions (no stimulation) in 82 healthy individuals (age range 22-55 years). We determined their DRD4 genotype and found an interaction with age: individuals who did not carry the 7-repeat allele (7R-, nâ=â53) had a significant (p<0.0001) negative association between age and relative glucose metabolism (normalized to whole brain glucose metabolism) in frontal (râ=â-0.52), temporal (râ=â-0.51) and striatal regions (râ=â-0.47, p<0.001); such that older individuals had lower metabolism than younger ones. In contrast, for carriers of the 7R allele (7R+ nâ=â29), these correlations with age were not significant and they only showed a positive association with cerebellar glucose metabolism (râ=â+0.55; pâ=â0.002). Regression slopes of regional brain glucose metabolism with age differed significantly between the 7R+ and 7R- groups in cerebellum, inferior temporal cortex and striatum. These results provide evidence that the DRD4 genotype might modulate the associations between regional brain glucose metabolism and age and that the carriers of the 7R allele appear to be less sensitive to the effects of age on brain glucose metabolism.
Assuntos
Envelhecimento/genética , Encéfalo/metabolismo , Glucose/genética , Glucose/metabolismo , Polimorfismo Genético/genética , Receptores de Dopamina D4/genética , Adulto , Alelos , Éxons/genética , Feminino , Genótipo , Humanos , MasculinoRESUMO
Human brain function is mediated by biochemical processes, many of which can be visualized and quantified by positron emission tomography (PET). PET brain imaging of monoamine oxidase A (MAO A)-an enzyme metabolizing neurotransmitters-revealed that MAO A levels vary widely between healthy men and this variability was not explained by the common MAOA genotype (VNTR genotype), suggesting that environmental factors, through epigenetic modifications, may mediate it. Here, we analyzed MAOA methylation in white blood cells (by bisulphite conversion of genomic DNA and subsequent sequencing of cloned DNA products) and measured brain MAO A levels (using PET and [(11)C]clorgyline, a radiotracer with specificity for MAO A) in 34 healthy non-smoking male volunteers. We found significant interindividual differences in methylation status and methylation patterns of the core MAOA promoter. The VNTR genotype did not influence the methylation status of the gene or brain MAO A activity. In contrast, we found a robust association of the regional and CpG site-specific methylation of the core MAOA promoter with brain MAO A levels. These results suggest that the methylation status of the MAOA promoter (detected in white blood cells) can reliably predict the brain endophenotype. Therefore, the status of MAOA methylation observed in healthy males merits consideration as a variable contributing to interindividual differences in behavior.
Assuntos
Encéfalo/enzimologia , Metilação de DNA/genética , Epigênese Genética , Monoaminoxidase , Adulto , Clorgilina , Ilhas de CpG/genética , Etnicidade/genética , Estudos de Associação Genética , Humanos , Masculino , Repetições Minissatélites , Monoaminoxidase/sangue , Monoaminoxidase/genética , Monoaminoxidase/metabolismo , Fenótipo , Tomografia por Emissão de Pósitrons , Regiões Promotoras Genéticas/genéticaRESUMO
The dopamine transporter (DAT) is a principal regulator of dopaminergic neurotransmission and its gene (the SLC6A3) is a strong biological candidate gene for various behavioral- and neurological disorders. Intense investigation of the link between the SLC6A3 polymorphisms and behavioral phenotypes yielded inconsistent and even contradictory results. Reliance on objective brain phenotype measures, for example, those afforded by brain imaging, might critically improve detection of DAT genotype-phenotype association. Here, we tested the relationship between the DAT brain availability and the SLC6A3 genotypes using an aggregate sample of 95 healthy participants of several imaging studies. These studies employed positron emission tomography (PET) with [¹¹C]cocaine wherein the DAT availability was estimated as Bmax/Kd; while the genotype values were obtained on two repeat polymorphisms--3-UTR- and intron 8--VNTRs. The main findings are the following: 1) both polymorphisms analyzed as single genetic markers and in combination (haplotype) modulate DAT density in midbrain; 2) ethnic background and age influence the strength of these associations; and 3) age-related changes in DAT availability differ in the 3-UTR and intron 8--genotype groups.
Assuntos
Encéfalo/metabolismo , Adulto , Proteínas da Membrana Plasmática de Transporte de Dopamina/genética , Feminino , Genótipo , Haplótipos , Humanos , Masculino , Pessoa de Meia-Idade , Polimorfismo Genético/genética , Tomografia por Emissão de Pósitrons , Adulto JovemRESUMO
CONTEXT: Long-term cocaine use has been associated with structural deficits in brain regions having dopamine-receptive neurons. However, the concomitant use of other drugs and common genetic variability in monoamine regulation present additional structural variability. OBJECTIVE: To examine variations in gray matter volume (GMV) as a function of lifetime drug use and the genotype of the monoamine oxidase A gene, MAOA, in men with cocaine use disorders (CUD) and healthy male controls. DESIGN: Cross-sectional comparison. SETTING: Clinical Research Center at Brookhaven National Laboratory. PATIENTS: Forty individuals with CUD and 42 controls who underwent magnetic resonance imaging to assess GMV and were genotyped for the MAOA polymorphism (categorized as high- and low-repeat alleles). MAIN OUTCOME MEASURES: The impact of cocaine addiction on GMV, tested by (1) comparing the CUD group with controls, (2) testing diagnosis × MAOA interactions, and (3) correlating GMV with lifetime cocaine, alcohol, and cigarette smoking, and testing their unique contribution to GMV beyond other factors. RESULTS: (1) Individuals with CUD had reductions in GMV in the orbitofrontal, dorsolateral prefrontal, and temporal cortex and the hippocampus compared with controls. (2) The orbitofrontal cortex reductions were uniquely driven by CUD with low- MAOA genotype and by lifetime cocaine use. (3) The GMV in the dorsolateral prefrontal cortex and hippocampus was driven by lifetime alcohol use beyond the genotype and other pertinent variables. CONCLUSIONS: Long-term cocaine users with the low-repeat MAOA allele have enhanced sensitivity to gray matter loss, specifically in the orbitofrontal cortex, indicating that this genotype may exacerbate the deleterious effects of cocaine in the brain. In addition, long-term alcohol use is a major contributor to gray matter loss in the dorsolateral prefrontal cortex and hippocampus, and is likely to further impair executive function and learning in cocaine addiction.
Assuntos
Alelos , Encéfalo/efeitos dos fármacos , Encéfalo/patologia , Transtornos Relacionados ao Uso de Cocaína/genética , Cocaína/toxicidade , Inibidores da Captação de Dopamina/toxicidade , Lobo Frontal/efeitos dos fármacos , Lobo Frontal/patologia , Processamento de Imagem Assistida por Computador , Imageamento por Ressonância Magnética , Monoaminoxidase/genética , Polimorfismo Genético/genética , Adulto , Alcoolismo/genética , Alcoolismo/patologia , Alcoolismo/psicologia , Atrofia , Transtornos Relacionados ao Uso de Cocaína/patologia , Transtornos Relacionados ao Uso de Cocaína/psicologia , Dopamina/metabolismo , Predisposição Genética para Doença/genética , Genótipo , Hipocampo/efeitos dos fármacos , Hipocampo/patologia , Humanos , Masculino , Pessoa de Meia-Idade , Tamanho do Órgão/efeitos dos fármacos , Córtex Pré-Frontal/efeitos dos fármacos , Córtex Pré-Frontal/patologia , Fumar/genética , Fumar/psicologia , Tabagismo/genética , Tabagismo/patologia , Tabagismo/psicologiaRESUMO
BACKGROUND: Methamphetamine is one of the most toxic of the drugs of abuse, which may reflect its distribution and accumulation in the body. However no studies have measured methamphetamine's organ distribution in the human body. METHODS: Positron Emission Tomography (PET) was used in conjunction with [(11)C]d-methamphetamine to measure its whole-body distribution and bioavailability as assessed by peak uptake (% Dose/cc), rate of clearance (time to reach 50% peak-clearance) and accumulation (area under the curve) in healthy participants (9 Caucasians and 10 African Americans). RESULTS: Methamphetamine distributed through most organs. Highest uptake (whole organ) occurred in lungs (22% Dose; weight â¼1246 g), liver (23%; weight â¼1677 g) and intermediate in brain (10%; weight â¼1600 g). Kidneys also showed high uptake (per/cc basis) (7%; weight 305 g). Methamphetamine's clearance was fastest in heart and lungs (7-16 minutes), slowest in brain, liver and stomach (>75 minutes), and intermediate in kidneys, spleen and pancreas (22-50 minutes). Lung accumulation of [(11)C]d-methamphetamine was 30% higher for African Americans than Caucasians (p<0.05) but did not differ in other organs. CONCLUSIONS: The high accumulation of methamphetamine, a potent stimulant drug, in most body organs is likely to contribute to the medical complications associated with methamphetamine abuse. In particular, we speculate that methamphetamine's high pulmonary uptake could render this organ vulnerable to infections (tuberculosis) and pathology (pulmonary hypertension). Our preliminary findings of a higher lung accumulation of methamphetamine in African Americans than Caucasians merits further investigation and questions whether it could contribute to the infrequent use of methamphetamine among African Americans.
Assuntos
Metanfetamina/farmacocinética , Adulto , Negro ou Afro-Americano , Transtornos Relacionados ao Uso de Anfetaminas/complicações , Área Sob a Curva , Estudos de Casos e Controles , Estimulantes do Sistema Nervoso Central/farmacocinética , Humanos , Hipertensão Pulmonar/complicações , Hipertensão Pulmonar/etnologia , Processamento de Imagem Assistida por Computador , Masculino , Tomografia por Emissão de Pósitrons/métodos , Distribuição Tecidual , Tuberculose Pulmonar/complicações , Tuberculose Pulmonar/etnologia , População BrancaRESUMO
Human dopamine transporter gene (DAT1 or SLC6A3) has been associated with various brain-related diseases and behavioral traits and, as such, has been investigated intensely in experimental- and clinical-settings. However, the abundance of research data has not clarified the biological mechanism of DAT regulation; similarly, studies of DAT genotype-phenotype associations yielded inconsistent results. Hence, our understanding of the control of the DAT protein product is incomplete; having this knowledge is critical, since DAT plays the major role in the brain's dopaminergic circuitry. Accordingly, we reevaluated the genomic attributes of the SLC6A3 gene that might confer sensitivity to regulation, hypothesizing that its unique genomic characteristics might facilitate highly dynamic, region-specific DAT expression, so enabling multiple regulatory modes. Our comprehensive bioinformatic analyzes revealed very distinctive genomic characteristics of the SLC6A3, including high inter-individual variability of its sequence (897 SNPs, about 90 repeats and several CNVs spell out all abbreviations in abstract) and pronounced sensitivity to regulation by epigenetic mechanisms, as evident from the GC-bias composition (0.55) of the SLC6A3, and numerous intragenic CpG islands (27 CGIs). We propose that this unique combination of the genomic features and the regulatory attributes enables the differential expression of the DAT1 gene and fulfills seemingly contradictory demands to its regulation; that is, robustness of region-specific expression and functional dynamics.
Assuntos
Proteínas da Membrana Plasmática de Transporte de Dopamina/genética , Epigênese Genética , Humanos , Fenótipo , Polimorfismo de Nucleotídeo ÚnicoRESUMO
Monoamine oxidase A (MAO A) is an enzyme that catalyzes the oxidation of neurotransmitter amines. A functional polymorphism in the human MAOA gene (high- and low-MAOA) has been associated with distinct behavioral phenotypes. To investigate directly the biological mechanism whereby this polymorphism influences brain function, we recently measured the activity of the MAO A enzyme in healthy volunteers. When found no relationship between the individual's brain MAO A level and the MAOA genotype, we postulated that there are additional regulatory mechanisms that control the MAOA expression. Given that DNA methylation is linked to the regulation of gene expression, we hypothesized that epigenetic mechanisms factor into the MAOA expression. Our underplaying assumption was that the differences in an individual's genotype play a key role in the epigenetic potential of the MAOA locus and, consequently, determine the individual's level of MAO A activity in the brain. As a first step towards experimental validation of the hypothesis, we performed a comprehensive bioinformatic analysis aiming to interrogate genomic features and attributes of the MAOA locus that might modulate its epigenetic sensitivity. Major findings of our analysis are the following: (1) the extended MAOA regulatory region contains two CpG islands (CGIs), one of which overlaps with the canonical MAOA promoter and the other is located further upstream; both CGIs exhibit sensitivity to differential methylation. (2) The uVNTR's effect on the MAOA's transcriptional activity might have epigenetic nature: this polymorphic region resides within the MAOA's CGI and itself contains CpGs, thus, the number of repeating increments effectively changes the number of methylatable cytosines in the MAOA promoter. An array of in silico analyses (the nucleosome positioning, the physical properties of the local DNA, the clustering of transcription-factor binding sites) together with experimental data on histone modifications and Pol 2 sites and data from the RefSeq mRNA library suggest that the MAOA gene might have an alternative promoter. Based on our findings, we propose a regulatory mechanism for the human MAOA according to which the MAOA expression in vivo is executed by the generation of tissue-specific transcripts initiated from the alternative promoters (both CGI-associated) where transcriptional activation of a particular promoter is under epigenetic control.
Assuntos
Biologia Computacional/métodos , Metilação de DNA/genética , Loci Gênicos/genética , Monoaminoxidase/genética , Animais , Composição de Bases/genética , Sequência de Bases , Cromatina/metabolismo , Sequência Conservada/genética , Ilhas de CpG/genética , Genoma Humano/genética , Histonas/metabolismo , Humanos , Isoenzimas/genética , Isoenzimas/metabolismo , Dados de Sequência Molecular , Filogenia , Primatas/genética , Regiões Promotoras Genéticas/genética , Processamento de Proteína Pós-Traducional , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Transcrição GênicaRESUMO
Reversible inhibitors of monoamine oxidase-A (RIMA) inhibit the breakdown of three major neurotransmitters, serotonin, norepinephrine and dopamine, offering a multi-neurotransmitter strategy for the treatment of depression. CX157 (3-fluoro-7-(2,2,2-trifluoroethoxy)phenoxathiin-10,10-dioxide) is a RIMA, which is currently in development for the treatment of major depressive disorder. We examined the degree and reversibility of the inhibition of brain monoamine oxidase-A (MAO-A) and plasma CX157 levels at different times after oral dosing to establish a dosing paradigm for future clinical efficacy studies, and to determine whether plasma CX157 levels reflect the degree of brain MAO-A inhibition. Brain MAO-A levels were measured with positron emission tomography (PET) imaging and [(11)C]clorgyline in 15 normal men after oral dosing of CX157 (20-80 mg). PET imaging was conducted after single and repeated doses of CX157 over a 24-h time course. We found that 60 and 80 mg doses of CX157 produced a robust dose-related inhibition (47-72%) of [(11)C]clorgyline binding to brain MAO-A at 2 h after administration and that brain MAO-A recovered completely by 24 h post drug. Plasma CX157 concentration was highly correlated with the inhibition of brain MAO-A (EC(50): 19.3 ng/ml). Thus, CX157 is the first agent in the RIMA class with documented reversible inhibition of human brain MAO-A, supporting its classification as a RIMA, and the first RIMA with observed plasma levels that can serve as a biomarker for the degree of brain MAO-A inhibition. These data were used to establish the dosing regimen for a current clinical efficacy trial with CX157.
Assuntos
Encéfalo/efeitos dos fármacos , Encéfalo/enzimologia , Compostos Heterocíclicos/metabolismo , Compostos Heterocíclicos/farmacologia , Inibidores da Monoaminoxidase/metabolismo , Inibidores da Monoaminoxidase/farmacologia , Monoaminoxidase/metabolismo , Proteínas do Tecido Nervoso/antagonistas & inibidores , Adulto , Clorgilina/metabolismo , Compostos Heterocíclicos/química , Humanos , Masculino , Pessoa de Meia-Idade , Proteínas do Tecido Nervoso/metabolismo , Tomografia por Emissão de Pósitrons/métodos , Ligação Proteica/efeitos dos fármacos , Ligação Proteica/fisiologia , Adulto JovemRESUMO
INTRODUCTION: Histone deacetylases (HDACs) are enzymes involved in epigenetic modifications that shift the balance toward chromatin condensation and silencing of gene expression. Here, we evaluate the utility of 6-([(18)F]fluoroacetamido)-1-hexanoicanilide ([(18)F]FAHA) for positron emission tomography imaging of HDAC activity in the baboon brain. For this purpose, we assessed its in vivo biodistribution, sensitivity to HDAC inhibition, metabolic stability and the distribution of the putative metabolite [(18)F]fluoroacetate ([(18)F]FAC). METHODS: [(18)F]FAHA and its metabolite [(18)F]FAC were prepared, and their in vivo biodistribution and pharmacokinetics were determined in baboons. [(18)F]FAHA metabolism and its sensitivity to HDAC inhibition using suberanilohydroxamic acid (SAHA) were assessed in arterial plasma and by in vitro incubation studies. The chemical form of F-18 in rodent brain was assessed by ex vivo studies. Distribution volumes for [(18)F]FAHA in the brain were derived. RESULTS: [(18)F]FAHA was rapidly metabolized to [(18)F]FAC, and both labeled compounds entered the brain. [(18)F]FAHA exhibited regional differences in brain uptake and kinetics. In contrast, [(18)F]FAC showed little variation in regional brain uptake and kinetics. A kinetic analysis that takes into account the uptake of peripherally produced [(18)F]FAC indicated that SAHA inhibited binding of [(18)F]FAHA in the baboon brain dose-dependently. In vitro studies demonstrated SAHA-sensitive metabolism of [(18)F]FAHA to [(18)F]FAC within the cell and diffusion of [(18)F]FAC out of the cell. All radioactivity in brain homogenate from rodents was [(18)F]FAC at 7 min postinjection of [(18)F]FAHA. CONCLUSION: The rapid metabolism of [(18)F]FAHA to [(18)F]FAC in the periphery complicates the quantitative analysis of HDAC in the brain. However, dose-dependent blocking studies with SAHA and kinetic modeling indicated that a specific interaction of [(18)F]FAHA in the brain was observed. Validating the nature of this interaction as HDAC specific will require additional studies.
Assuntos
Anilidas , Encéfalo/diagnóstico por imagem , Encéfalo/enzimologia , Histona Desacetilases/metabolismo , Papio , Anilidas/sangue , Anilidas/metabolismo , Anilidas/farmacocinética , Animais , Artérias/metabolismo , Encéfalo/citologia , Encéfalo/metabolismo , Linhagem Celular Tumoral , Difusão , Inibidores Enzimáticos/farmacologia , Fluoracetatos/sangue , Fluoracetatos/metabolismo , Fluoracetatos/farmacocinética , Inibidores de Histona Desacetilases , Humanos , Ácidos Hidroxâmicos/farmacologia , Espaço Intracelular/metabolismo , Modelos Biológicos , Tomografia por Emissão de Pósitrons , Ratos , Distribuição Tecidual , VorinostatRESUMO
The genetic deletion of monoamine oxidase A (MAO A), an enzyme that breaks down the monoamine neurotransmitters norepinephrine, serotonin, and dopamine, produces aggressive phenotypes across species. Therefore, a common polymorphism in the MAO A gene (MAOA, Mendelian Inheritance in Men database number 309850, referred to as high or low based on transcription in non-neuronal cells) has been investigated in a number of externalizing behavioral and clinical phenotypes. These studies provide evidence linking the low MAOA genotype and violent behavior but only through interaction with severe environmental stressors during childhood. Here, we hypothesized that in healthy adult males the gene product of MAO A in the brain, rather than the gene per se, would be associated with regulating the concentration of brain amines involved in trait aggression. Brain MAO A activity was measured in vivo in healthy nonsmoking men with positron emission tomography using a radioligand specific for MAO A (clorgyline labeled with carbon 11). Trait aggression was measured with the multidimensional personality questionnaire (MPQ). Here we report for the first time that brain MAO A correlates inversely with the MPQ trait measure of aggression (but not with other personality traits) such that the lower the MAO A activity in cortical and subcortical brain regions, the higher the self-reported aggression (in both MAOA genotype groups) contributing to more than one-third of the variability. Because trait aggression is a measure used to predict antisocial behavior, these results underscore the relevance of MAO A as a neurochemical substrate of aberrant aggression.
Assuntos
Agressão/fisiologia , Encéfalo/enzimologia , Monoaminoxidase/metabolismo , Adulto , Agressão/psicologia , Aminas/metabolismo , Clorgilina , Humanos , Masculino , Inibidores da Monoaminoxidase , Concentração Osmolar , Personalidade , Tomografia por Emissão de Pósitrons , Valor Preditivo dos Testes , Inquéritos e Questionários , Distribuição TecidualRESUMO
Lysophosphatidic acid is an important lipid ligand regulating many aspects of cell function, including proliferation and migration. Operating via heterotrimeric G proteins to downstream effectors, lysophosphatidic acid was shown to regulate the function and trafficking of the G protein-coupled beta(2)-adrenergic receptor. C3 exotoxin, expression of dominant negative RhoA, and inhibition of c-Jun N-terminal kinase blocked the ability of lysophosphatidic acid to sequester the beta(2)-adrenergic receptor, whereas expression of constitutively active Galpha(13), p115RhoGEF, or RhoA mimicked lysophosphatidic acid (LPA) action, stimulating the internalization of the Galpha(s)-coupled beta(2)-adrenergic receptor. This study revealed a novel cross-talk exerted from the LPA/Galpha(13)/p115RhoGEF/RhoA pathway to the beta(2)-adrenergic receptor/Galpha(s)/adenylyl cyclase pathway, attenuating the ability of beta-adrenergic agonists to act following stimulation of cells by LPA as may occur during beta-adrenergic therapy of an inflammatory response.
Assuntos
Subunidades alfa G12-G13 de Proteínas de Ligação ao GTP/metabolismo , Fatores de Troca do Nucleotídeo Guanina/metabolismo , Lisofosfolipídeos/fisiologia , MAP Quinase Quinase 4/metabolismo , Receptores Adrenérgicos beta 2/metabolismo , Linhagem Celular Tumoral , Membrana Celular/metabolismo , Genes Dominantes , Humanos , Inflamação , Lisofosfolipídeos/metabolismo , Microscopia Confocal , Modelos Biológicos , Transporte Proteico , Fatores de Troca de Nucleotídeo Guanina Rho , Transdução de Sinais , Proteína rhoA de Ligação ao GTP/metabolismoRESUMO
Hormones that activate receptor tyrosine kinases have been shown to regulate G protein-coupled receptors, and herein we investigate the ability of IGF-I to regulate the beta(1)-adrenergic receptor. Treating Chinese hamster ovary cells in culture with IGF-I is shown to functionally antagonize the ability of expressed beta(1)-adrenergic receptors to accumulate intracellular cAMP in response to stimulation by the beta-adrenergic agonist Iso. The attenuation of beta(1)-adrenergic action was accompanied by internalization of beta(1)-adrenergic receptors in response to IGF-I. Inhibiting either phosphatidylinositol 3-kinase or the serine/threonine protein kinase Akt blocks the ability of IGF-I to antagonize and to internalize beta(1)-adrenergic receptors. Mutation of one potential Akt substrate site Ser412Ala, but not another Ser312Ala, of the beta(1)-adrenergic receptor abolishes the ability of IGF-I to functionally antagonize and to sequester the beta(1)-adrenergic receptor. We also tested the ability of IGF-I to regulate beta(1)-adrenergic receptors and their signaling in adult canine cardiac myocytes. IGF-I attenuates the ability of beta(1)-adrenergic receptors to accumulate intracellular cAMP in response to Iso and promotes internalization of beta(1)-adrenergic receptors in these cardiac myocytes.
Assuntos
Antagonistas de Receptores Adrenérgicos beta 1 , Fator de Crescimento Insulin-Like I/farmacologia , Receptores Adrenérgicos beta 1/metabolismo , Antagonistas Adrenérgicos/farmacologia , Agonistas Adrenérgicos beta/metabolismo , Agonistas Adrenérgicos beta/farmacologia , Animais , Células CHO , Células Cultivadas , Cricetinae , Cricetulus , Cães , Miócitos Cardíacos/efeitos dos fármacos , Miócitos Cardíacos/metabolismo , Proteína Oncogênica v-akt/fisiologia , Fosfatidilinositol 3-Quinases/fisiologia , Transdução de Sinais/efeitos dos fármacos , Fatores de TempoRESUMO
The AKAP gravin is a scaffold for protein kinases, phosphatases, and adaptor molecules obligate for resensitization and recycling of beta2-adrenergic receptors. Gravin binds to the receptor through well characterized protein-protein interactions. These interactions are facilitated approximately 1000-fold when gravin is anchored to the cytoplasmic leaflet of the plasma membrane. Although the N-terminal region (approximately 550 residues) is highly negatively charged and probably natively unfolded, it could anchor gravin to the inner leaflet through hydrophobic insertion of its N-terminal myristate and electrostatic binding of three short positively charged domains (PCDs). Loss of the site of N-myristoylation was found to affect neither AKAP macroscopic localization nor AKAP function. Synthetic peptides corresponding to PCD1-3 bound in vitro to unilamellar phospholipid vesicles with high affinity, a binding reversed by calmodulin in the presence of Ca2+. In vivo gravin localization is regulated by intracellular Ca2+, a function mapping to the N terminus of the protein harboring PCD1, PCD2, and PCD3. Mutation of any two PCDs eliminates membrane association of the non-myristoylated gravin, the sensitivity to Ca2+/calmodulin, and the ability of this scaffold to catalyze receptor resensitization and recycling.