The use of TaqMan RT-PCR assays for semiquantitative analysis of gene expression in CNS tissues and disease models

Abstract

TaqMan reverse transcription polymerase chain reaction (RT-PCR) is a recently developed technique which allows the measurement of an accumulating PCR product in real time. In the present study we have validated the use of TaqMan RT-PCR for mRNA localisation studies in human and rat tissues, and for the investigation of gene expression changes in CNS animal models. In human brain, D 2 receptor mRNA was enriched in caudate nucleus and putamen, whilst in rat brain, highest levels of D 2 receptor mRNA expression were observed in striatum and nucleus accumbens, consistent with the known distribution of this receptor in basal ganglia. In a rat model of permanent middle cerebral artery occlusion (pMCAO), endogenous interleukin-1 receptor antagonist (IL-1ra) mRNA was upregulated over 30-fold at 24 h post-lesion in both striatum and cortex ipsilateral to artery occlusion. Brain-derived neurotrophic factor (BDNF) mRNA was transiently upregulated 3.7-fold at 3 h, but not at 24 h or 3 days after induction of cortical spreading depression (CSD) in rats. Our observations in these two animal models using TaqMan RT-PCR were consistent with previous reports using other techniques. In conclusion, TaqMan RT-PCR assays provide a rapid and reliable method for semi-quantitative analysis of gene expression in the nervous system.