Tadross, J. A. | bioRxiv Science

bioRxiv2021

Relaxin/insulin-like family peptide receptor 4 (Rxfp4) expressing hypothalamic neurons modulate food intake and preference in mice

Relaxin/insulin-like-family peptide receptor-4 (RXFP4), the cognate receptor for insulin-like peptide 5 (INSL5), has been implicated in feeding behaviour as Rxfp4 knockout mice display shorter meal durations and reduced high fat diet (HFD) intake. Here, we generated transgenic Rxfp4-Cre mice to explore Rxfp4 expression and physiology. Using this model, we identified Rxfp4 expression in the central nervous system, including in the ventromedial hypothalamus (VMH). Intra-VMH infusion of INSL5 increased HFD and highly palatable liquid meal intake (HPM) of ad libitum fed wildtype mice. Single-cell RNA-sequencing of VMH Rxfp4-expressing cells (RXFP4VMH) defined a cluster of Rxfp4-labelled neurons expressing Esr1, Tac1 and Oxtr, alongside known appetite-modulating neuropeptide receptors (Mc4r, Cckar and Nmur2). Viral tracing demonstrated RXFP4VMH neural projections to the bed nucleus of the stria terminalis, paraventricular hypothalamus, paraventricular thalamus and central nucleus of the amygdala. Utilising designer receptors exclusively activated by designer drugs (DREADDs), we found that whole body chemogenetic inhibition (Di) of Rxfp4-expressing cells, mimicking native INSL5-RXFP4 signalling, increased intake of HFD and HPM, whilst activation (Dq), either at whole body level or specifically within the VMH, reduced HFD and HPM intake and altered food preference. Ablating VMH Rxfp4-expressing cells recapitulated the lower HFD intake phenotype of Rxfp4 knockout mice, resulting in reduced body weight. These findings identify a discrete Rxfp4-expressing neuronal population as a key regulator of food intake and preference and reveal hypothalamic RXFP4 signalling as a target for feeding behaviour manipulation.

physiology↗

bioRxiv2021

A survey of the mouse hindbrain in the fed and fasted state using single-nucleus RNA sequencing

ObjectiveThe area postrema (AP) and the nucleus tractus solitaris (NTS), located in the hindbrain, are key nuclei that sense and integrate peripheral nutritional signals and, consequently, regulate feeding behaviour. While single cell transcriptomics have been used in mice to reveal the gene expression profile and heterogeneity of key hypothalamic populations, similar in-depth studies have not yet been performed in the hindbrain. MethodsUsing single-nucleus RNA sequencing, we provide a detailed survey of 16,034 cells within the AP and NTS of the mouse, in the fed and fasted state. ResultsOf these, 8910 are neurons that group into 30 clusters, with 4289 coming from mice fed ad libitum and 4621 from overnight fasted mice. 7124 nuclei are from non-neuronal cells, including oligodendrocytes, astrocytes and microglia. Interestingly, we identified that the oligodendrocyte population was particularly transcriptionally sensitive to an overnight fast. The receptors GLP1R, GIPR, GFRAL and CALCR, which bind GLP1, GIP, GDF15 and amylin respectively, are all expressed in the hindbrain and are major targets for anti-obesity therapeutics. We characterise the transcriptomes of these four populations and show that their gene expression profiles are not dramatically altered by an overnight fast. Notably, we find that roughly half of cells that express GIPR are oligodendrocytes. Additionally, we profile POMC expressing neurons within the hindbrain and demonstrate that 84% of POMC neurons express either PCSK1, PSCK2 or both, implying that melanocortin peptides are likely produced by these neurons. ConclusionWe provide a detailed single-cell level characterisation of AP and NTS cells expressing receptors for key anti-obesity drugs that are either already approved for human use or are in clinical trials. This resource will help delineate the mechanisms underlying the effectiveness of these compounds, and also prove useful in the continued search for other novel therapeutic targets.

neuroscience↗