Patterns of Cellular Overlap in Expression of c‐Fos and Arc Activity‐Regulated Genes in the Mouse Brain Regions During Acquisition and Retrieval of Contextual Conditioned Fear

Abstract

Consolidation of long-term memory requires expression of immediate early genes (IEGs) encoding regulatory and effector proteins, such as inducible transcription factor c-Fos and activity-regulated cytoskeleton-associated protein Arc/Arg3.1. Since these IEGs are involved in various forms of behavioral and stimulus-induced plasticity, their products are often used to label active neurons, and their promoters are employed to tag and manipulate active engram cells. However, little is still known as to whether c-Fos and Arc proteins are induced identically and in the same neurons of different brain structures under various behavioral conditions. In the present work we compared expression patterns of c-Fos and Arc proteins in the mouse brain after acquisition and retrieval of contextual fear and analyzed cellular overlap of their expression in both conditions. Our hypothesis was that the activity patterns of two genes would not be identical though would substantially overlap according to similarities and differences in signaling cascades driving their expression. Both cohorts of mice were subjected to standard contextual fear conditioning with an electric footshock. One group of animals was sacrificed 90 minutes after the training session and the other was tested for retrieval of contextual memory on the next day and sacrificed 90 minutes later. The percentage of freezing in the conditioning context was measured as an index of memory. Brain sections were collected from hippocampus, amygdala and associative cortices of experimental mice, active control mice and home cage control mice. Double immunohistochemical staining with anti-Fos and anti-Arc antibodies was performed, and stained sections were scanned with a confocal microscope. The cell count showed behavioral induction of both c-Fos and Arc in all the examined brain structures. However, there was only a partial overlap of about 30% between the two labelled cell populations. Arc-positive cells predominated over c-Fos-positive cells in the hippocampus regardless of cognitive task, and in cingulate and parietal cortex Arc prevalence was stronger in after memory retrieval compared to learning. Moreover, within the above structures localization of c-Fos- and Arc-positive cells had different distribution across cortical layers and hippocampal subfields. Our results indicate that c-Fos and Arc proteins are not equivalent markers of stimulus-induced neural activity, as they are expressed in cell populations that differ by more than 60%. This data imposes a significant restraint on the use of these IEGs as interchangeable markers of neuronal activity/plasticity and calls for a further study of the differences in their regulation during behavior and learning.