RESUMEN
The subcutaneous and systemic injection of serotonin reduces cutaneous and visceral pain thresholds and increases responses to noxious stimuli. Different subtypes of 5-hydroxytryptamine (5-HT) receptors are suggested to be associated with different types of pain responses. Here we show that serotonin also inhibits catechol O-methyltransferase (COMT), an enzyme that contributes to modultion the perception of pain, via non-competitive binding to the site bound by catechol substrates with a binding affinity comparable to the binding affinity of catechol itself (K(i) = 44 µM). Using computational modeling, biochemical tests and cellular assays we show that serotonin actively competes with the methyl donor S-adenosyl-L-methionine (SAM) within the catalytic site. Binding of serotonin to the catalytic site inhibits the access of SAM, thus preventing methylation of COMT substrates. The results of in vivo animal studies show that serotonin-induced pain hypersensitivity in mice is reduced by either SAM pretreatment or by the combined administration of selective antagonists for ß(2)- and ß(3)-adrenergic receptors, which have been previously shown to mediate COMT-dependent pain signaling. Our results suggest that inhibition of COMT via serotonin binding contributes to pain hypersensitivity, providing additional strategies for the treatment of clinical pain conditions.
Asunto(s)
Inhibidores de Catecol O-Metiltransferasa , Umbral del Dolor/efectos de los fármacos , Serotonina/farmacología , Antagonistas Adrenérgicos beta/farmacología , Animales , Catecol O-Metiltransferasa/metabolismo , Femenino , Masculino , Ratones , Umbral del Dolor/fisiología , Unión Proteica/efectos de los fármacos , S-Adenosilmetionina/farmacología , Serotonina/química , Serotonina/metabolismoRESUMEN
Core symptoms of autism include deficits in social interaction, impaired communication, and restricted, repetitive behaviors. The repetitive behavior domain encompasses abnormal motoric stereotypy, an inflexible insistence on sameness, and resistance to change. In recent years, many genetic mouse models of autism and related disorders have been developed, based on candidate genes for disease susceptibility. The present studies are part of an ongoing initiative to develop appropriate behavioral tasks for the evaluation of mouse models relevant to autism. We have previously reported profiles for sociability, preference for social novelty, and resistance to changes in a learned pattern of behavior, as well as other functional domains, for 10 inbred mouse strains of divergent genetic backgrounds. The present studies extend this multi-component behavioral characterization to several additional strains: C58/J, NOD/LtJ, NZB/B1NJ, PL/J, SJL/J, SWR/J, and the wild-derived PERA/EiJ. C58/J, NOD/LtJ, NZB/B1NJ, SJL/J, and PERA/EiJ demonstrated low sociability, measured by time spent in proximity to an unfamiliar conspecific, with 30-60% of mice from these strains showing social avoidance. In the Morris water maze, NZB/B1NJ had a persistent bias for the quadrant where the hidden platform was located during acquisition, even after 9 days of reversal training. A particularly interesting profile was found for C58/J, which had low social preference, poor performance in the T-maze, and overt motoric stereotypy. Overall, this set of tasks and observational methods provides a strategy for evaluating novel mouse models in behavioral domains relevant to the autism phenotype.
Asunto(s)
Ratones Endogámicos/fisiología , Conducta Social , Conducta Estereotipada/fisiología , Análisis de Varianza , Animales , Conducta Animal , Conducta Exploratoria , Habituación Psicofisiológica/fisiología , Masculino , Aprendizaje por Laberinto , Ratones , Movimiento , Aprendizaje Inverso/fisiología , Especificidad de la EspecieRESUMEN
A functional allele of the mouse catechol-O-methyltransferase (Comt) gene is defined by the insertion of a B2 short interspersed repeat element in its 3'-untranslated region (UTR). This allele has been associated with a number of phenotypes, such as pain and anxiety. In comparison with mice carrying the ancestral allele (Comt+), Comt B2i mice show higher Comt mRNA and enzymatic activity levels. Here, we investigated the molecular genetic mechanisms underlying this allelic specific regulation of Comt expression. Insertion of the B2 element introduces an early polyadenylation signal generating a shorter Comt transcript, in addition to the longer ancestral mRNA. Comparative analysis and in silico prediction of Comt mRNA potential targets within the transcript 3' to the B2 element was performed and allowed choosing microRNA (miRNA) candidates for experimental screening: mmu-miR-3470a, mmu-miR-3470b, and mmu-miR-667. Cell transfection with each miRNA downregulated the expression of the ancestral transcript and COMT enzymatic activity. Our in vivo experiments showed that mmu-miR-667-3p is strongly correlated with decreasing amounts of Comt mRNA in the brain, and lentiviral injections of mmu-miR-3470a, mmu-miR-3470b, and mmu-miR-667 increase hypersensitivity in the mouse formalin model, consistent with reduced COMT activity. In summary, our data demonstrate that the Comt+ transcript contains regulatory miRNA signals in its 3'-untranslated region leading to mRNA degradation; these signals, however, are absent in the shorter transcript, resulting in higher mRNA expression and activity levels.
Asunto(s)
Encéfalo/metabolismo , Catecol O-Metiltransferasa/genética , Regulación de la Expresión Génica/fisiología , Dolor/genética , Regiones no Traducidas 3' , Animales , Línea Celular Transformada , Femenino , Perfilación de la Expresión Génica , Regulación de la Expresión Génica/genética , Macrófagos , Masculino , Ratones , Ratones Endogámicos , MicroARNs/genética , Análisis de Secuencia por Matrices de Oligonucleótidos , Dolor/metabolismo , Dimensión del Dolor , Poliadenilación/genética , ARN Mensajero/metabolismo , TransfecciónRESUMEN
Catechol-O-methyltransferase (COMT) metabolizes catecholaminergic neurotransmitters. Numerous studies have linked COMT to pivotal brain functions such as mood, cognition, response to stress, and pain. Both nociception and risk of clinical pain have been associated with COMT genetic variants, and this association was shown to be mediated through adrenergic pathways. Here, we show that association studies between COMT polymorphic markers and pain phenotypes in 2 independent cohorts identified a functional marker, rs165774, situated in the 3' untranslated region of a newfound splice variant, (a)-COMT. Sequence comparisons showed that the (a)-COMT transcript is highly conserved in primates, and deep sequencing data demonstrated that (a)-COMT is expressed across several human tissues, including the brain. In silico analyses showed that the (a)-COMT enzyme features a distinct C-terminus structure, capable of stabilizing substrates in its active site. In vitro experiments demonstrated not only that (a)-COMT is catalytically active but also that it displays unique substrate specificity, exhibiting enzymatic activity with dopamine but not epinephrine. They also established that the pain-protective A allele of rs165774 coincides with lower COMT activity, suggesting contribution to decreased pain sensitivity through increased dopaminergic rather than decreased adrenergic tone, characteristic of reference isoforms. Our results provide evidence for an essential role of the (a)-COMT isoform in nociceptive signaling and suggest that genetic variations in (a)-COMT isoforms may contribute to individual variability in pain phenotypes.
Asunto(s)
Catecol O-Metiltransferasa/genética , Regulación de la Expresión Génica/genética , Umbral del Dolor/fisiología , Polimorfismo de Nucleótido Simple/genética , Trastornos de la Articulación Temporomandibular/genética , Encéfalo/metabolismo , Estudios de Casos y Controles , Catecol O-Metiltransferasa/metabolismo , Línea Celular Tumoral , Estudios de Cohortes , Femenino , Variación Genética , Humanos , Masculino , Neuroblastoma/patología , Dolor/etiología , Dolor/genética , Fenotipo , ARN Mensajero/metabolismo , Trastornos de la Articulación Temporomandibular/complicaciones , TransfecciónRESUMEN
Chronic pain is a highly prevalent and poorly managed human health problem. We used microarray-based expression genomics in 25 inbred mouse strains to identify dorsal root ganglion (DRG)-expressed genetic contributors to mechanical allodynia, a prominent symptom of chronic pain. We identified expression levels of Chrna6, which encodes the α6 subunit of the nicotinic acetylcholine receptor (nAChR), as highly associated with allodynia. We confirmed the importance of α6* (α6-containing) nAChRs by analyzing both gain- and loss-of-function mutants. We find that mechanical allodynia associated with neuropathic and inflammatory injuries is significantly altered in α6* mutants, and that α6* but not α4* nicotinic receptors are absolutely required for peripheral and/or spinal nicotine analgesia. Furthermore, we show that Chrna6's role in analgesia is at least partially due to direct interaction and cross-inhibition of α6* nAChRs with P2X2/3 receptors in DRG nociceptors. Finally, we establish the relevance of our results to humans by the observation of genetic association in patients suffering from chronic postsurgical and temporomandibular pain.
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Dolor Crónico/genética , Receptores Nicotínicos/genética , Receptores Purinérgicos P2X2/metabolismo , Receptores Purinérgicos P2X3/metabolismo , Animales , Regulación hacia Abajo , Transferencia Resonante de Energía de Fluorescencia , Ganglios Espinales/metabolismo , Humanos , Ratones , Ratones Mutantes , Antagonistas del Receptor Purinérgico P2X/farmacologíaRESUMEN
BACKGROUND: Frontotemporal dementia (FTD) is a clinically heterogeneous condition that can be associated with clinical manifestations of an extrapyramidal disorder or motor neuron disease. A range of histologic patterns has been described in patients with FTD. The most common familial form of this condition is caused by mutations in the microtubule-associated protein tau gene (MAP tau) and is associated with neuronal or glial tau inclusions. OBJECTIVES: To determine the clinical, anatomic, and pathological features of San Francisco family A and to map the mutation responsible for disease in this family. DESIGN: A systematic clinical, neuropsychologic, neuroimaging, and chromosome segregation analysis of San Francisco family A was performed. A pathological and biochemical assessment of a family member was made. SETTING: Family study. PATIENTS: San Francisco family A, with FTD, variable extrapyramidal symptoms, and prominent motor neuron disease. Afflicted family members do not have a MAP tau coding or splice regulatory sequence mutation, and the MAP tau is genetically excluded. MAIN OUTCOME MEASURES: Comparison of clinical, neuropsychologic, neuroimaging, and linkage findings of San Francisco family A with other familial forms of FTD and amyotrophic lateral sclerosis (ALS). RESULTS: The most probable location for the mutation responsible for this condition is on chromosome arm 17q, distal to the MAP tau. All previously identified susceptibility loci for FTD and ALS are excluded. Autopsy findings from an afflicted family member show distinctive tau and alpha-synuclein inclusions. Another unique feature is that the insoluble tau protein consists predominantly of the 4R/0N isoform. CONCLUSION: The condition affecting members of San Francisco family A is clinically, pathologically, and genetically distinct from previous familial forms of FTD and ALS.
Asunto(s)
Esclerosis Amiotrófica Lateral/genética , Demencia/genética , Cuerpos de Inclusión/metabolismo , Proteínas del Tejido Nervioso/metabolismo , Proteínas tau/metabolismo , Adulto , Anciano , Esclerosis Amiotrófica Lateral/metabolismo , Esclerosis Amiotrófica Lateral/patología , Western Blotting/métodos , Encéfalo/metabolismo , Encéfalo/patología , Cromosomas Humanos Par 17 , Demencia/metabolismo , Demencia/patología , Salud de la Familia , Femenino , Ligamiento Genético , Humanos , Escala de Lod , Imagen por Resonancia Magnética/métodos , Masculino , Persona de Mediana Edad , Mutación , Examen Neurológico/métodos , Pruebas Neuropsicológicas , Reacción en Cadena de la Polimerasa/métodos , Isoformas de Proteínas/metabolismo , Sinucleínas , alfa-Sinucleína , Proteínas tau/genéticaRESUMEN
The enzyme catechol-O-methyltransferase (COMT) metabolizes catecholamine neurotransmitters involved in a number of physiological functions, including pain perception. Both human and mouse COMT genes possess functional polymorphisms contributing to interindividual variability in pain phenotypes such as sensitivity to noxious stimuli, severity of clinical pain, and response to pain treatment. In this study, we found that the effects of Comt functional variation in mice are modality specific. Spontaneous inflammatory nociception and thermal nociception behaviors were correlated the most with the presence of the B2 SINE transposon insertion residing in the 3'UTR mRNA region. Similarly, in humans, COMT functional haplotypes were associated with thermal pain perception and with capsaicin-induced pain. Furthermore, COMT genetic variations contributed to pain behaviors in mice and pain ratings in humans in a sex-specific manner. The ancestral Comt variant, without a B2 SINE insertion, was more strongly associated with sensitivity to capsaicin in female vs male mice. In humans, the haplotype coding for low COMT activity increased capsaicin-induced pain perception in women, but not men. These findings reemphasize the fundamental contribution of COMT to pain processes, and provide a fine-grained resolution of this contribution at the genetic level that can be used to guide future studies in the area of pain genetics.
Asunto(s)
Catecol O-Metiltransferasa/genética , Dolor/genética , Polimorfismo de Nucleótido Simple/genética , Caracteres Sexuales , Adolescente , Adulto , Animales , Capsaicina/toxicidad , Estudios de Cohortes , Modelos Animales de Enfermedad , Femenino , Genotipo , Humanos , Masculino , Ratones , Ratones Endogámicos , Persona de Mediana Edad , Dolor/inducido químicamente , Umbral del Dolor/efectos de los fármacos , Umbral del Dolor/fisiología , Fármacos del Sistema Sensorial/toxicidad , Especificidad de la Especie , Adulto JovenRESUMEN
Ample data suggest that alcohol dependence represents a heritable condition, and several research groups have performed linkage analysis to identify genomic regions influencing this disorder. In the present study, a genome-wide linkage scan for alcohol dependence was conducted in a community sample of 565 probands and 1080 first-degree relatives recruited through the UCSF Family Alcoholism Study. The Semi-Structured Assessment for the Genetics of Alcoholism (SSAGA) was used to derive DSM-IV alcohol dependence diagnoses. Although no loci achieved genome-wide significance (i.e., LOD score > 3.0), several linkage peaks of interest (i.e., LOD score > 1.0) were identified. When the strict DSM-IV alcohol dependence diagnosis requiring the temporal clustering of symptoms served as the phenotype, linkage peaks were identified on chromosomes 1p36.31-p36.22, 2q37.3, 8q24.3, and 18p11.21-p11.2. When the temporal clustering of symptoms was not required, linkage peaks were again identified on chromosomes 1p36.31-p36.22 and 8q24.3 as well as novel loci on chromosomes 1p22.3, 2p24.3-p24.1, 9p24.1-p23, and 22q12.3-q13.1. Follow-up analyses were conducted by performing linkage analysis for the 12 alcohol dependence symptoms assessed by the SSAGA across the support intervals for the observed linkage peaks. These analyses demonstrated that different collections of symptoms often assessing distinct aspects of alcohol dependence (e.g., uncontrollable drinking and withdrawal vs. tolerance and drinking despite health problems) contributed to each linkage peak and often yielded LOD scores exceeding that reported for the alcohol dependence diagnosis. Such findings provide insight into how specific genomic regions may influence distinct aspects of alcohol dependence.
Asunto(s)
Alcoholismo/genética , Ligamiento Genético , Alcoholismo/diagnóstico , Manual Diagnóstico y Estadístico de los Trastornos Mentales , Predisposición Genética a la Enfermedad , Pruebas Genéticas/métodos , Estudio de Asociación del Genoma Completo , Genotipo , HumanosRESUMEN
BACKGROUND: A low level of response (LR) to alcohol seems to relate to a substantial proportion of the risk for alcoholism and to have significant heritability. METHODS: This report describes the results of a genome-wide segregation analysis for the first 139 pairs of full siblings by using an alcohol challenge protocol as a direct measure of LR. Subjects from 18 to 29 years old were selected if the original screen indicated they had an alcohol-dependent parent, reported a personal history of drinking but had no evidence of alcohol dependence, and had a full sibling with similar characteristics. Body sway and Subjective High Assessment Scale scores were measured at baseline and at regular intervals after the administration of a measured dose of alcohol. Participants and available parents were genotyped for 811 microsatellite markers, and resulting data were analyzed with a variance component method. RESULTS: Nine chromosome regions with logarithm of the odds ratio (LOD) between 2.2 and 3.2 were identified; several had previously been implicated regarding phenotypes relevant to alcoholism and the LR to alcohol. Several regions identified in the previous linkage study by using a retrospective self-report questionnaire were potentially confirmed by this study. The strongest evidence was on chromosomes 10, 11, and 22. CONCLUSIONS: Several chromosomal areas seem to relate to the low LR to alcohol as a risk factor for alcohol dependence.
Asunto(s)
Consumo de Bebidas Alcohólicas/genética , Etanol/administración & dosificación , Ligamiento Genético/genética , Escala de Lod , Adolescente , Adulto , Femenino , Ligamiento Genético/efectos de los fármacos , Humanos , Masculino , Linaje , FenotipoRESUMEN
We describe eight members from two large Amish kindreds who share a phenotype characterized by early-onset pigmentary retinopathy and myopia, global developmental delay and mental retardation, microcephaly, short stature, hypotonia, joint hyperextensibility, small hands and feet, common facial appearance, and friendly disposition. Several of the children had intermittent granulocytopenia. The phenotypic occurrence in three siblings coupled with the increased coefficient of inbreeding in the Amish suggested that this disorder is autosomal recessive and due to a single founder allele. Despite similarity to the clinical features of Cohen syndrome, experienced dysmorphologists attending the 23rd David W. Smith Workshop suggested the facial gestalt of the Amish children was inconsistent with this diagnosis. We mapped the locus responsible for these individuals' phenotype to chromosome 8q22-q23, which contains the recently discovered Cohen syndrome gene, COH1. Complete sequencing of the COH1 gene identified a likely disease-causing frameshift mutation and a missense mutation in the Amish patients. A comparison of features among different Cohen syndrome populations with shared linkage to the COH1 locus or known COH1 gene mutations may allow for the determination of improved clinical criteria on which to suspect the diagnosis of Cohen syndrome. We conclude that facial gestalt seems to be an unreliable indicator of Cohen syndrome between ethnic populations, although it is quite consistent among affected individuals within a particular ethnic group. Other features common to almost all individuals with proven COH1 mutations, such as retinal dystrophy, myopia, microcephaly, mental retardation, global developmental delay, hypotonia, and joint hyperextensibility appear to be better clinical indicators of this disorder.