RESUMEN
The gradual loss of striatal dopamine and dopaminergic neurons residing in the substantia nigra (SN) causes parkinsonism characterized by slow, halting movements, rigidity, and resting tremor when neuronal loss exceeds a threshold of approximately 80%. It is estimated that there is extensive compensation for several years prior to symptom onset, during which vulnerable neurons asynchronously die. Recent evidence would argue that much of the compensatory response of the nigrostriatal system is multimodal including both pre-synaptic and striatal mechanisms. Although parkinsonism may have multiple causes, the classic syndrome, Parkinson's disease (PD), is frequently modeled in small animals by repeated administration of the selective neurotoxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). Because the MPTP model of PD recapitulates many of the known behavioral and pathological features of human PD, we asked whether the striatal cells of mice treated with MPTP in a semi-chronic paradigm enact a transcriptional program that would help elucidate the response to dopamine denervation. Our findings reveal a time-dependent dysregulation in the striatum of a set of genes whose products may impact both the viability and ability to communicate of dopamine neurons in the SN.
Asunto(s)
1-Metil-4-fenil-1,2,3,6-Tetrahidropiridina/farmacología , Cuerpo Estriado/efectos de los fármacos , Expresión Génica/efectos de los fármacos , Intoxicación por MPTP/metabolismo , Análisis de Varianza , Animales , Cuerpo Estriado/metabolismo , Modelos Animales de Enfermedad , Intoxicación por MPTP/genética , Masculino , Ratones , Ratones Endogámicos C57BL , Modelos Biológicos , Análisis de Secuencia por Matrices de Oligonucleótidos/métodos , Análisis de Componente Principal/métodos , Reproducibilidad de los ResultadosRESUMEN
We describe a simple method of using rolling circle amplification to amplify vector DNA such as M13 or plasmid DNA from single colonies or plaques. Using random primers and phi29 DNA polymerase, circular DNA templates can be amplified 10,000-fold in a few hours. This procedure removes the need for lengthy growth periods and traditional DNA isolation methods. Reaction products can be used directly for DNA sequencing after phosphatase treatment to inactivate unincorporated nucleotides. Amplified products can also be used for in vitro cloning, library construction, and other molecular biology applications.