Network Neuroscience Untethered: Brain-Wide Immediate Early Gene Expression for the Analysis of Functional Connectivity in Freely Behaving Animals

SIMPLE SUMMARY: Understanding the relationships between sets of regions across the brain is critical to understanding of cognitive function. The application of graph theoretical analyses to brain-wide immediate early gene expression density provides a means of studying these relationships in freely behaving animal models. Here, we provide overviews of the steps required to apply these techniques and acknowledge critical considerations which should be kept in mind when designing and conducting these experiments. ABSTRACT: Studying how spatially discrete neuroanatomical regions across the brain interact is critical to advancing our understanding of the brain. Traditional neuroimaging techniques have led to many important discoveries about the nature of these interactions, termed functional connectivity. However, in animal models these traditional neuroimaging techniques have generally been limited to anesthetized or head-fixed setups or examination of small subsets of neuroanatomical regions. Using the brain-wide expression density of immediate early genes (IEG), we can assess brain-wide functional connectivity underlying a wide variety of behavioural tasks in freely behaving animal models. Here, we provide an overview of the necessary steps required to perform IEG-based analyses of functional connectivity. We also outline important considerations when designing such experiments and demonstrate the implications of these considerations using an IEG-based network dataset generated for the purpose of this review.