Early-life prefrontal cortex inhibition and early-life stress lead to long-lasting behavioral, transcriptional, and physiological impairments

Early-life stress has been linked to multiple neurodevelopmental and neuropsychiatric deficits. Our previous studies have linked maternal presence/absence from the nest in developing rat pups to changes in prefrontal cortex activity. Furthermore, we have shown that these changes are modulated by serotonergic signaling. Here we test whether changes in prefrontal cortex activity during early-life affect the developing cortex leading to behavioral alterations in the adult. We show that inhibiting the prefrontal cortex of mouse pups leads to cognitive deficits in the adult comparable to those seen following maternal separation. Moreover, we show that activating the prefrontal cortex during maternal separation can prevent these behavioral deficits. To test how maternal separation affects the transcriptional profile of the prefrontal cortex we performed single-nucleus RNA sequencing. Maternal separation lead to differential gene expression almost exclusively in inhibitory neurons. Among others, we found changes in GABAergic and serotonergic pathways in these interneurons. Interestingly, both maternal separation and early-life prefrontal cortex inhibition led to changes in physiological responses in prefrontal activity to GABAergic and serotonergic antagonists that were similar to the responses of more immature brains. Prefrontal activation during maternal separation prevented these changes. These data point to a crucial role of prefrontal cortex activity during early-life in behavioral expression in adulthood Overall design: C57Bl/6J mice were subjected to the maternal separation protocol described above. The PFC of 4 control and 4 MS male mice was dissected at P13; another set of 4 control and 4 MS mice were allowed to grow until P70 at which point we dissected the PFC. Tissue was dissected using a tissue puncher and flash frozen in dry ice. Samples were homogenized, and the nuclei were purified using an iodixanol cushion. Each sample was stained with HTO-tagged antibodies. Pools of eight hashed samples were processed simultaneously. We performed library preparations using the 10X Chromium platform and sequenced them on the Illumina NovaSeq. Each run consisted of two P13 controls, two adult controls, two P13 MS and two adult MS samples. One adult mouse from the MS group was removed from the analyses because it was a female misidentified as a male.