Detecting neural activity-dependent immediate early gene expression in the brain
In this chapter, we present an in situ hybridization protocol with radioactive probe that has been successfully and easily used on detecting mRNA expression level and patterns, in multiple tissue types and developmental stages. To detect behaviorally regulated, i.e., motor or sensory, mRNA expression of immediate early genes (IEGs) within cells and tissues in vivo, in situ hybridization is a powerful method for discovering neural activity correlations and novel neural structures. Compared with nonradioactive probe methods such as digoxigenin (DIG) labeling, the radioactive probe hybridization method provides a semi-linear relation between signal intensity and targeted mRNA amounts for quantitative analysis. Furthermore, this method allows us high-throughput mRNA expression analysis for 100–200 sides with 400–1,000 tissue sections simultaneously. This method allows identifying the possible significance and function of interested genes in the nervous system under specific behaviors.
