Dynamics of stress-induced c-fos expression in the rat prelimbic cortex: lessons from intronic and mature RNA and protein analyses

Abstract

Despite the extensive use of c-fos as a marker of stress-induced neuronal activation, key aspects regarding its dynamics of expression remain poorly characterized. In the present study, we assessed in the prelimbic cortex of adult male rats the immediate transcriptional response of c-fos by measuring the heteronuclear (hn)RNA and mature (m)RNA expression by double fluorescent in situ hybridization as well as the c-Fos protein using immunofluorescence (FOS). We quantified in three different experiments the number of c-fos hnRNA+, mRNA+ and FOS+ neurons under basal conditions, immediately after different periods of immobilization stress (IMO), and after a recovery period. Our results indicate that stress induced a large increase in the number of positive neurons for all markers analyzed, each displaying a different time course. Moreover, our findings indicate that measuring the intensity of signal per neuron also provides relevant information. In addition, we report an increased number of FOS+ neurons after only 8–15 min of IMO, suggesting a surprisingly fast initiation of protein translation. Finally, the maturation from c-fos hnRNA+ to mRNA+ might depend on the duration and/or intensity of stress-induced activation. Our findings contribute to a better understanding of the dynamics of stress-induced c-fos expression and underscore the importance of examining multiple molecular components when using c-fos as a proxy of neuronal activation.