Investigation of Oxtr-expressing Neurons Projecting to Nucleus Accumbens using Oxtr-ires-Cre Knock-in prairie Voles (Microtus ochrogaster)

Abstract

Social bonds such as parent-infant attachment or pair bonds can be critical for mental and physical well-being. The monogamous prairie vole (Microtus ochrogaster) has proven useful for examining the neural substrates regulating social behaviors, including social bonding. Oxytocin (OXT) and oxytocin receptor (OXTR) play critical roles in alloparental care, pair bonding and consoling behavior in prairie voles. While OXTR in a few regions, such as the nucleus accumbnes (NAcc), prefrontal cortex (PFC) and anterior cingulate cortex (ACC), have been implicated in regulating these behaviors, the extent to which other OXT sensitive areas modulate social behaviors has not been investigated. The NAcc is a central hub for modulating OXTR dependent social behaviors. To identify neurons expressing Oxtr in prairie vole brain, we generated gene knock-in voles expressing Cre recombinase in tandem with Oxtr (Oxtr-ires-Cre) using CRISPR/Cas9 genome editing. We confirmed Oxtr and Cre mRNA co-localization in NAcc, validating this model. Next, we identified putative Oxtr-expressing neurons projecting to NAcc by infusing retrograde CRE-dependent EGFP AAV into NAcc and visualizing fluorescence. We found enhanced green fluorescent protein (EGFP) positive neurons in anterior olfactory nucleus, PFC, ACC, insular cortex (IC), paraventricular thalamus (PVT), basolateral amygdala (BLA), and posteromedial and posterolateral cortical amygdaloid area (PMCo, PLCo). The ACC to NAcc OXTR projection may represent a species-specific circuit since Oxtr-expressing neurons in the ACC of mice were reported not to project to the NAcc. This is the first delineation of Oxtr-expressing neural circuits in the prairie vole, and demonstrates the utility of this novel genetically modified organism for characterizing OXTR circuits involved in social behaviors.