RESUMO
Activation of AMPA receptors (AMPARs) in the nucleus accumbens is necessary for the reinstatement of cocaine-seeking behavior, an animal model of drug craving and relapse. AMPARs are tetrameric protein complexes that consist of GluA1-4 subunits, of which GluA2 imparts calcium permeability. Adenosine deaminase acting on RNA 2 (ADAR2) is a nuclear enzyme that is essential for editing GluA2 pre-mRNA at Q/R site 607. Unedited GluA2(Q) subunits form calcium-permeable AMPARs (CP-AMPARs), whereas edited GluA2(R) subunits form calcium-impermeable channels (CI-AMPARs). Emerging evidence suggests that the reinstatement of cocaine seeking is associated with increased synaptic expression of CP-AMPARs in the nucleus accumbens. However, the role of GluA2 Q/R site editing and ADAR2 in cocaine seeking is unclear. In the present study, we investigated the effects of forced cocaine abstinence on GluA2 Q/R site editing and ADAR2 expression in the nucleus accumbens. Our results demonstrate that 7 days of cocaine abstinence is associated with decreased GluA2 Q/R site editing and reduced ADAR2 expression in the accumbens shell, but not core, of cocaine-experienced rats compared with yoked saline controls. To examine the functional significance of ADAR2 and GluA2 Q/R site editing in cocaine seeking, we used viral-mediated gene delivery to overexpress ADAR2b in the accumbens shell. Increased ADAR2b expression in the shell attenuated cocaine priming-induced reinstatement of drug seeking and was associated with increased GluA2 Q/R site editing and surface expression of GluA2-containing AMPARs. Taken together, these findings support the novel hypothesis that an increased contribution of accumbens shell CP-AMPARs containing unedited GluA2(Q) promotes cocaine seeking. Therefore, CP-AMPARs containing unedited GluA2(Q) represent a novel target for cocaine addiction pharmacotherapies.
Assuntos
Adenosina Desaminase/metabolismo , Cocaína/administração & dosagem , Inibidores da Captação de Dopamina/administração & dosagem , Comportamento de Procura de Droga/efeitos dos fármacos , Regulação da Expressão Gênica/efeitos dos fármacos , Receptores de AMPA/metabolismo , Adenosina Desaminase/genética , Animais , Cálcio/metabolismo , Condicionamento Operante/efeitos dos fármacos , Desoxirribonucleases de Sítio Específico do Tipo II/administração & dosagem , Regulação da Expressão Gênica/fisiologia , Masculino , Núcleo Accumbens/efeitos dos fármacos , Núcleo Accumbens/metabolismo , Edição de RNA/efeitos dos fármacos , Edição de RNA/genética , RNA Mensageiro/metabolismo , Ratos , Ratos Sprague-Dawley , Receptores de AMPA/genética , Autoadministração , Transdução GenéticaRESUMO
Intensive scientific research devoted in the recent years to understand the molecular mechanisms or neurodegeneration in spinocerebellar ataxias (SCAs) are identifying new pathways and targets providing new insights and a better understanding of the molecular pathogenesis in these diseases. In this consensus manuscript, the authors discuss their current views on the identified molecular processes causing or modulating the neurodegenerative phenotype in spinocerebellar ataxias with the common opinion of translating the new knowledge acquired into candidate targets for therapy. The following topics are discussed: transcription dysregulation, protein aggregation, autophagy, ion channels, the role of mitochondria, RNA toxicity, modulators of neurodegeneration and current therapeutic approaches. Overall point of consensus includes the common vision of neurodegeneration in SCAs as a multifactorial, progressive and reversible process, at least in early stages. Specific points of consensus include the role of the dysregulation of protein folding, transcription, bioenergetics, calcium handling and eventual cell death with apoptotic features of neurons during SCA disease progression. Unresolved questions include how the dysregulation of these pathways triggers the onset of symptoms and mediates disease progression since this understanding may allow effective treatments of SCAs within the window of reversibility to prevent early neuronal damage. Common opinions also include the need for clinical detection of early neuronal dysfunction, for more basic research to decipher the early neurodegenerative process in SCAs in order to give rise to new concepts for treatment strategies and for the translation of the results to preclinical studies and, thereafter, in clinical practice.
Assuntos
Degeneração Neural/fisiopatologia , Degeneração Neural/terapia , Ataxias Espinocerebelares/fisiopatologia , Ataxias Espinocerebelares/terapia , Animais , Autofagia , Humanos , Canais Iônicos/metabolismo , Mitocôndrias/fisiologia , Degeneração Neural/genética , Degeneração Neural/patologia , RNA/metabolismo , Ataxias Espinocerebelares/genética , Ataxias Espinocerebelares/patologia , Transcrição GênicaRESUMO
A simpler, less expensive, and faster high-performance liquid chromatographic method was shown to be an alternative to urokinase potency determinations by the Ploug method. Post-elution recovery of the low-molecular-weight form was 104 +/- 2.4% as determined by the Ploug method. Two analysts reported relative standard deviations of 1.6% and 1.1% based on peak height determination of eight replicate injections of a single sample of low-molecular-weight material. Linearity at the same wavelength for low- and high-molecular-weight forms was 0.9999 and 0.9992, respectively, for peak height versus potency.