Abstract

Sex-differences in selectively bred high- and low-responder rats: spatial gene expression in the dorsal hippocampus

Waselus M, Hebda-Bauer EK, Dai M, Meng F, Akil H, Watson SJ
50th Annual Meeting of the Society for Neuroscience, Virtual. 2021.

Historically, our laboratory has used radioactive in situ hybridization to localize and quantify differences in gene expression in various brain regions. Our selectively-bred lines of high- and low-responder rats (bHR and bLR, respectively) are advantageous in that we can use this model to study two “extreme” behavioral phenotypes (e.g., differences in locomotor activity in a novel environment, propensity to self-administer drugs of abuse, and anxiety-like behaviors). In situ hybridization has been used to measure gene expression in the brains of these rats which can then be mapped back on to behavioral differences. Unfortunately, traditionally used radioactive in situ hybridization studies are labor-intensive and costly, with only a finite amount of material available from a single subject thus limiting the number of genes examined. Recently, spatial transcriptomics, most recently the Visium Spatial Gene Expression solution developed by 10X Genomics (Pleasanton, CA), allows RNA-seq on a small region of interest within tissue sections, reducing the hands-on processing time and increasing the number of genes examined in a single study while retaining spatial resolution. Brains from the 68th generation of our selective breeding colony were used to examine both bHR/bLR- and sex differences in gene expression in the dorsal hippocampus of adult rats. One section per “condition” (bred line x sex) was collected from a bHR and bLR family per slide, allowing each slide to function as its own independent study. Adjacent sections were collected on a second slide to serve as a technical replicate. A second bHR and bLR family was used to examine bHR/bLR and sex differences, serving as a biological replicate. Our initial results indicate that all Visium samples were of high quality for use in downstream analyses. We first wanted to demonstrate that known markers of sex differences were different male and female rat brain, regardless of breeding background. To that end, we found two examples of Y-linked genes (i.e., Eif2s3y and Kdm5d) are expressed in the brains of male, but not female, rats. Furthermore, specific genes of interest are differentially expressed in the brains of adult bHR and bLR rats, consistent with other ongoing internal studies. Our conclusions thus far indicate that in our hands, Visium has the potential to be a powerful tool for elucidating differences in gene expression between not only male and female rats, but also selectively bred bHR and bLR rats. Further studies will allow us to combine single cell data generated by our group with spatial datasets to determine the precise localization of gene expression differences within larger brain structures.