Effect of CCK-8s injection (20 ug/kg BW i.p.) on mRNA translational activity in hypothalamic oxytocin neurons of mice fed standard chow or high-fat/high-sugar diet (RiboTag;TRAP)

Oxytocin-expressing paraventricular hypothalamic neurons (PVN^OT neurons) integrate afferent signals from the gut including cholecystokinin (CCK) to adjust whole-body energy homeostasis. However, the molecular underpinnings by which PVN^OT neurons orchestrate gut-to-brain feeding control remain unclear. Here, we show that mice undergoing selective ablation of PVNOT neurons fail to reduce food intake in response to CCK and develop hyperphagic obesity on chow diet. Notably, exposing wildtype mice to a high-fat/high-sugar (HFHS) diet recapitulates this insensitivity towards CCK, which is linked to diet-induced transcriptional and electrophysiological aberrations specifically in PVNOT neurons. Restoring OT pathways in DIO mice via chemogenetics or polypharmacology sufficiently re-establishes CCK?s anorexigenic effects. Lastly, by single-cell profiling, we identify a specialized PVN^OT neuronal subpopulation with increased ?-opioid signaling under HFHS diet, which restrains their CCK-evoked activation. In sum, we here document a novel (patho)mechanism by which PVN^OT signaling uncouples a gut-brain satiation pathway under obesogenic conditions. Overall design: We treated male Oxt-ires-Cre;RiboTag mice with either vehicle or CCK, collected hypothalami 2h post injection, pooled 2-3 tissues per sample, and affinity purified conditionally HA-tagged ribosomes (incl translating mRNA) specifically from oxytocin neurons. We then performed gene expression profiling analysis using data obtained from RNA-seq of immunoprecipitates versus inputs (n=4 per group) from standard chow diet fed mice and inputs only from high-fat/high-sugar diet fed mice (n=4 per group) Comparative gene expression profiling analysis of RNA-seq data for TRAP (translating ribosome affinity pufification) between vehicle and CCK-treated mice across immunopreciptates per diet. HTSeq quantification of transcript expression